SOCIETY OF ECOLOGICAL CHEMISTRY AND ENGINEERING

ECOLOGICAL CHEMISTRY AND ENGINEERING A

CHEMIA I IN¯YNIERIA EKOLOGICZNA A

Vol. 18 No. 12 OPOLE 2011 EDITORIAL COMMITTEE Witold Wac³awek (Society of Ecological Chemistry and Engineering, PL) – Editor-in-Chief Marina V. Frontasyeva (Joint Institute for Nuclear Research, Dubna, RU) – heavy metals and radionuclides Vasil Simeonov (University of Sofia, BG) – monitoring Maria Wac³awek (University, Opole, PL) – alternative energy sources Barbara Wiœniowska-Kielian (University of Agriculture, Kraków, PL) – agricultural chemistry

PROGRAMMING BOARD Witold Wac³awek (Society of Ecological Chemistry and Engineering, PL) – Chairman Jerzy Bartnicki (Meteorological Institute – DNMI, Oslo-Blindern, NO) Mykhaylo Bratychak (National University of Technology, Lviv, UA) Bogus³aw Buszewski (Nicolaus Copernicus University, Toruñ, PL) Eugenija Kupcinskiene (University of Agriculture, Kaunas, LT) Bernd Markert (International Graduate School [IHI], Zittau, DE) Nelson Marmiroli (University, Parma, IT) Jacek Namieœnik (University of Technology, Gdañsk, PL) Lucjan Paw³owski (University of Technology, Lublin, PL) Krzysztof J. Rudziñski (Institute of Physical Chemistry PAS, Warszawa, PL) Manfred Sager (Agency for Health and Food Safety, Vienna, AT) Mark R.D. Seaward (University of Bradford, UK) Pavlina Simeonova (Bulgarian Academy of Sciences, Sofia, BG) Petr Škarpa (Mendel University of Agriculture and Forestry, Brno, CZ) Piotr Tomasik (University of Agriculture, Kraków, PL) Roman Zarzycki (University of Technology, £ódŸ, PL) Ma³gorzata Rajfur (University, Opole, PL) – Secretary

STATISTICAL EDITORS W³adys³aw Kamiñski (Technical University, £ódŸ, PL) Zbigniew Ziembik (Opole University, Opole, PL)

LANGUAGE EDITOR Ian Barnes (University of Wuppertal, Wuppertal, DE)

EDITORIAL OFFICE Opole University ul. kard. B. Kominka 6, 45–032 OPOLE, PL phone: +48 77 455 91 49 email: [email protected]

SECRETARY Ma³gorzata Rajfur phone: +48 77 401 60 42 email: [email protected]

Copyright © by Society of Ecological Chemistry and Engineering, Opole

Ecological Chemistry and Engineering A / Chemia i In¿ynieria Ekologiczna A is partly financed by Ministry of Science and Higher Education, Warszawa

ISSN 1898–6188 CONTENTS

Marcin KOZAK, W³adys³aw MALARZ, Andrzej KOTECKI, Waldemar HELIOS and Joanna GÓRA – Follow-up Effect of Hilling on Growth and Yielding of Miscanthus (Miscanthus x giganteus Greef et Deu.) ...... 1599 El¿bieta SACA£A – Miscanthus – Unusual Grass: Biochemical and Physiological Characteristic: A Review ...... 1615 Anna GORCZYCA and Marek J. KASPROWICZ – Initial Research on the Effect of the Nanogro Plant Growth Stimulator on Fusarium culmorum (W.G. Smith) Sacc. . . 1625 Marcin SIDORUK, Andrzej ROCHWERGER, El¿bieta SKORBI£OWICZ and Miros³aw SKORBI£OWICZ – Effect of Catchment Area Use on Lead and Zinc Accumulation in the Bottom Deposits of Lakes Ardung and Bukwald ...... 1633 Zdzis³aw CIEÆKO, Miros³aw WYSZKOWSKI and El¿bieta ROLKA – Aluminium Concentration in Plants Depending on Soil Contamination with Cadmium ...... 1641 Janina GOSPODAREK – Effect of Soil Contamination with a Mixture of Heavy Metals on Broad Bean (Vicia faba L.) Seed Quality ...... 1651 Pawe³ WOLSKI, Iwona ZAWIEJA and Lidia WOLNY – Impact of Temperature on Viscosity of Sewage Sludge After Conditioning ...... 1659 Teresa RAUCKYTE-¯AK and S³awomir ¯AK – Changeability of the Speciation Forms of Heavy Metals in Soil Subject to Many Years of Fertilization with Wastewaters from Vegetable Fat Production ...... 1667 Beata GRYGIERZEC – Effect of Nitrogen Fertilization on Seed Production of Lolium perenne L. Turfgrass Cultivars ...... 1675 Joanna KOSTECKA and Grzegorz P¥CZKA – Kitchen Wastes as a Source of Nitrogen and Other Macroelements According to Technology of Vermiculture .... 1683 Teresa RAUCKYTE-¯AK and Bo¿ena SZEJNIUK – Influence of 1,3,4-Thiadiazole Derivatives on the Biological Activity of the Selected Environmental Bacteria ..... 1691 Krystyna PRZYBULEWSKA, Anna STOLARSKA and Daria G£¥BOWSKA – Change of Proline Content in Selected Soil Fungi as Affected by Osmotic Stress ...... 1705 Joanna JARMU£-PIETRASZCZYK, Marta KAMIONEK and Ewelina MALINOWSKA – Occurrence of Entomopathogenic Fungi and Nematodes on Pastures in Central and Northern Poland ...... 1711 Wiera MICHALCEWICZ, S³awomir STANKOWSKI, Ma³gorzata GA£CZYÑSKA and Marzena GIBCZYÑSKA – Successive Influence of Fluidal Ashes on General Number of Bacteria, Actinomycetes and Fungi in Pot Experiments .... 1715 Katarzyna GLEÑ – Effect of Foliar Fertilizers and Their Mixtures on Phytopathogenic Fusarium Fungi ...... 1721 Ma³gorzata NABRDALIK and Katarzyna GRATA – Influence of the Culture Conditions on Lipolytic Activity of Bacillus cereus and Bacillus mycoides ...... 1727 Adam RADKOWSKI – Effect of Microelement Fertilization on the Quality and Nutritional Value of the Meadow Sward Hay. Part II. The Content of Macroelements ...... 1737 Anna KWIECIÑSKA and Krystyna KONIECZNY – Recovery of Industrial Water from Pig Liquid Manure by Means of Membrane Techniques ...... 1743 Karolina MIELCZAREK, Jolanta BOHDZIEWICZ and Anna KWARCIAK- -KOZ£OWSKA – Application of Polysulfone Membranes for Coke-Making Wastewater Treatment ...... 1751 Justyna HACHO£ and El¿bieta BONDAR-NOWAKOWSKA – Ecological Risk Classification in the Regulated and Conserved Watercourses ...... 1763

INDEXES

Contents of Volume 18 of „Ecological Chemistry and Engineering A” ...... 1777 Author Index of Volume 18 of „Ecological Chemistry and Engineering A” ...... 1787 Subject Index ...... 1793 Indeks rzeczowy ...... 1801 Index of Latin, Polish and English Species Names of Microorganisms, Plants and Animals and their Anatomical Parts ...... 1809 Index of Acronyms ...... 1813 Wykaz akronimów ...... 1815

VARIA

Central European Conference ECOpole ’11. Short Conference Report ...... 1819 Invitation for ECOpole ’12 Conference ...... 1825 Zaproszenie na Konferencjê ECOpole ’12 ...... 1827 Acknowledgement of Reviewers ...... 1829 Guide for Authors on Submission of Manuscripts ...... 1831 Zalecenia dotycz¹ce przygotowania manuskryptów ...... 1833 SPIS TREŒCI

Marcin KOZAK, W³adys³aw MALARZ, Andrzej KOTECKI, Waldemar HELIOS i Joanna GÓRA – Nastêpczy wp³yw obredlania na rozwój i plonowanie miskanta olbrzymiego (Miscanthus x giganteus Greef et Deu.) ...... 1599 El¿bieta SACA£A – Miscanthus – trawa niezwyk³a: charakterystyka biochemiczno- -fizjologiczna: przegl¹d literaturowy ...... 1615 Anna GORCZYCA i Marek J. KASPROWICZ – Wstêpne badania nad wp³ywem stymulatora wzrostu roœlin Nanogro na Fusarium culmorum (W.G. Smith) Sacc. .... 1625 Marcin SIDORUK, Andrzej ROCHWERGER, El¿bieta SKORBI£OWICZ i Miros³aw SKORBI£OWICZ – Wp³yw u¿ytkowania zlewni na akumulacjê o³owiu i cynku w osadach dennych na przyk³adzie jezior Ardung i Bukwa³d ...... 1633 Zdzis³aw CIEÆKO, Miros³aw WYSZKOWSKI i El¿bieta ROLKA – Zawartoœæ glinu w roœlinach w zale¿noœci od zanieczyszczenia gleby kadmem ...... 1641 Janina GOSPODAREK – Oddzia³ywanie ska¿enia gleby mieszanin¹ metali ciê¿kich na jakoœæ nasion bobu (Vicia faba L.)...... 1651 Pawe³ WOLSKI, Iwona ZAWIEJA i Lidia WOLNY – Wp³yw temperatury na lepkoœæ kondycjonowanych osadów œciekowych ...... 1659 Teresa RAUCKYTE-¯AK i S³awomir ¯AK – Zmiennoœæ form specjacyjnych metali ciê¿kich w glebie poddanej wieloletniemu nawo¿eniu œciekami z produkcji t³uszczów roœlinnych ...... 1667 Beata GRYGIERZEC – Nawo¿enie azotem w produkcji nasiennej gazonowych odmian Lolium perenne L...... 1675 Joanna KOSTECKA i Grzegorz P¥CZKA – Odpady kuchenne jako Ÿród³o azotu i innych makropierwiastków zale¿nie od technologii prowadzenia wermikultury .... 1683 Teresa RAUCKYTE-¯AK i Bo¿ena SZEJNIUK – Wp³yw pochodnych 1,3,4-tiadiazoli na aktywnoœæ wybranych bakterii œrodowiskowych ...... 1691 Krystyna PRZYBULEWSKA, Anna STOLARSKA and Daria G£¥BOWSKA – Zmiana zawartoœci proliny u wybranych grzybów glebowych pod wp³ywem stresu osmotycznego ...... 1705 Joanna JARMU£-PIETRASZCZYK, Marta KAMIONEK and Ewelina MALINOWSKA – Wystêpowanie grzybów i nicieni entomopatogennych na pastwiskach w centralnej i pó³nocnej Polsce ...... 1711 Wiera MICHALCEWICZ, S³awomir STANKOWSKI, Ma³gorzata GA£CZYÑSKA i Marzena GIBCZYÑSKA – Nastêpczy wp³yw popio³ów fluidalnych na ogóln¹ liczbê bakterii, promieniowców i grzybów w badaniach wazonowych ...... 1715 Katarzyna GLEÑ – Wp³yw nawozów dolistnych i ich mieszanin na grzyby fitopatogenne z rodzaju Fusarium ...... 1721 Ma³gorzata NABRDALIK i Katarzyna GRATA – Wp³yw warunków œrodowiska na aktywnoœæ lipolityczn¹ szczepów Bacillus cereus i Bacillus mycoides ...... 1727 Adam RADKOWSKI – Wp³yw nawo¿enia mikroelementami na jakoœæ i wartoœæ pokarmow¹ siana runi ³¹kowej. Cz. II. Zawartoœæ makroelementów ...... 1737 Anna KWIECIÑSKA i Krystyna KONIECZNY – Odzysk wody przemys³owej z gnojowicy trzody chlewnej z wykorzystaniem technik membranowych ...... 1743 Karolina MIELCZAREK, Jolanta BOHDZIEWICZ i Anna KWARCIAK-KOZ£OWSKA – Membrany polisulfonowe w oczyszczaniu œcieków koksowniczych ...... 1751 Justyna HACHO£ i El¿bieta BONDAR-NOWAKOWSKA – Klasyfikacja ryzyka ekologicznego w ciekach regulowanych i konserwowanych ...... 1763

INDEXES

Contents of Volume 18 of „Ecological Chemistry and Engineering A” ...... 1777 Author Index of Volume 18 of „Ecological Chemistry and Engineering A” ...... 1787 Subject Index ...... 1793 Indeks rzeczowy ...... 1801 Index of Latin, Polish and English Species Names of Microorganisms, Plants and Animals and their Anatomical Parts ...... 1809 Index of Acronyms ...... 1813 Wykaz akronimów ...... 1815

VARIA

Central European Conference ECOpole ’11. Short Conference Report ...... 1819 Invitation for ECOpole ’12 Conference ...... 1825 Zaproszenie na Konferencjê ECOpole ’12 ...... 1827 Acknowledgement of Reviewers ...... 1829 Guide for Authors on Submission of Manuscripts ...... 1831 Zalecenia dotycz¹ce przygotowania manuskryptów ...... 1833 ECOLOGICAL CHEMISTRY AND ENGINEERING A

Vol. 18, No. 12 2011

Marcin KOZAK1*, W³adys³aw MALARZ1, Andrzej KOTECKI1, Waldemar HELIOS1 and Joanna GÓRA1

FOLLOW-UP EFFECT OF HILLING ON GROWTH AND YIELDING OF MISCANTHUS (Miscanthus x giganteus Greef et Deu.)

NASTÊPCZY WP£YW OBREDLANIA NA ROZWÓJ I PLONOWANIE MISKANTA OLBRZYMIEGO (Miscanthus x giganteus Greef et Deu.)

Abstract: In 2005–2007 in Pawlowice near Wroclaw, Poland, field experiments were conducted on the follow-up effect of hilling of Miscanthus in autumn after seeding. The split-plot experiment was set for the following three variable factors: I. Harvest date: a – autumn harvest after the vegetation period ends (11.07.2005, 14.12.2006; b – winter harvest before the vegetation starts (10.03.2006, 09.03.2007); II. Autumn treatment of rhizomes after seeding: a – with hilling; b – without hilling; III. N fertilization: 100, 150 and 200 kg N × ha–1. In the first years after the experiment had been set up, the morphological features, dry matter yield, water and ash contents were related to the age of the plantation. Among the investigated factors, harvest dates had the most significant influence on yielding. Winter harvest resulted in a lower by 18.4 % dry matter yield and a decrease in water content in green matter by 23.8 % and in ash content by 43 %. However, it increased energy value in green matter by 52 %. In the second and the third year of the cultivation, out of 1 ha field of Miscanthus, it is possible to obtain biomass yield with a mean energy value of 294 GJ, which corresponds to 7.03 toe. It is possible to obtain high dry matter yields of Miscanthus with autumn hilling and applying 150 kg N × ha–1. Keywords: Miscanthus, harvest date, hilling, N fertilization

Introduction

Energy development strategies of the developed countries focus more and more on obtaining renewable energy sources. In Poland, according to “The strategy for develop- ment of renewable energy” passed by the Polish Parliament, the renewable energy

1 Department of Crop Production, Wroclaw University of Environmental and Life Sciences, pl. Grunwaldzki 24a, 50–363 Wroc³aw, Poland, phone: +48 71 320 16 41, fax: +48 71 320 16 39, email: [email protected] * Corresponding author. 1600 Marcin Kozak et al sources should make for 7.5 % of all the energy in 2010 and reach 14 % in 2020. In order to achieve these levels, it will be necessary to produce biomass on energy plantations located on arable lands additionally to firewood and agricultural by-products. Potentially useful species for energy production in Polish environment include willow (Salix sp.), Miscanthus (Miscanthus x giganteus Greef et Deu), Virginia mallow (Sida hermaphrodita Rusby) and Jerusalem artichoke (Helianthus tuberosus L.). Miscanthus x giganteus is a hybrid of a tetraploid M. sinenss and a diploid M. sacchariflorus. It was imported from Japan to Denmark in 1930 by Aksel Olsen and it gave start to cultivars grown at present in Europe. Since 1983 field experiments have been conducted with this species in the Northern Europe. The plant has been proved to be an effective energy crop and produce over 20 Mg d.m. per ha–1 × year–1 [1]. Miscanthus can be harvested twice: in autumn from October to November (and with favourable conditions in the first decade of December) when the vegetation period ends or in winter from February to March before the vegetation starts. Harvest dates affect the yield and the quality of the combusted biomass [1, 2]. Dry matter content in the harvested biomass in autumn is between 35–45 %, and in the winter harvest it is from 60 to 70 % [3]. The lower dry matter yield in winter is caused by some leaves falling down. The reduced yield is compensated by the increase in cellulose content which is low in leaves. The other reason for a lower yield of the aerial parts of the plants is the fact that in autumn the plants transport nutrients to the rhizomes which increase in mass and change in their chemical content. Since these are the rhizomes that determine winter survival, the plants gather in them osmotic active compounds which reduce the plants’ susceptibility to low temperatures [4]. The yield of biomass in the autumn harvest is higher than in the spring harvest, the differences can be from 14 to 30 % [5]. The disadvantage of the autumn harvest is not only high moisture of the obtained biomass but also higher removal (accumulation and extraction) of the nutrients from the fields [6]. During the winter period the plants shed leaves, which makes potassium drain to soil [7]. As a consequence, sodium and potassium contents are reduced in Miscanthus straw and later in ashes [5]. Biomass of the highest quality for energy can be obtained in a winter–spring harvest. A later harvest results in biomass that is on average 30 % drier than in autumn. Winter harvest reduces also biomass chlorine content. The presence of chlorine in the biomass can cause emission of hydrogen chloride and dioxins in combustion [5, 8]. A drawback in growing Miscanthus is its little tolerance of low temperature in the first year after seeding. During the first winter after starting the plantation, rhizomes which are planted too shallow and those which are not fully formed are destroyed by frost and high moisture. Literature does not provide information on similar problems with winter survival of Miscanthus in the second and the following years of the cultivation [1]. After the second year of growth, Miscanthus survives well temperature drops below – 20 oC, even without a snow cover [4]. In the Danish and Irish experiments the majority of Miscanthus plants grown through micropropagation (in vitro) did not survive the first winter after seeding, while the plants obtained from rhizomes managed well through the winter period. Numerous Follow-up Effect of Hilling on Growth and Yielding of Miscanthus... 1601 experiments indicate that the method of breeding of Miscanthus can affect its ability to survive in winter conditions [9]. Miscanthus needs relatively little fertilisation thanks to use and transport of nutrients in the plants from rhizomes and accumulation of large amounts of nutrients in rhizomes by the end of the vegetation period. It has been determined that a dose of 60 kg N × ha–1 is sufficient for an appropriate development of rhizomes. The leaves usually fall down and stay on the fields so nutrients absorption is relevant only for rhizomes. Miscanthus is capable to make a good use of nitrogen because it translocates it from rhizomes to shoots in spring and is again able to cumulate it in them at the end of the vegetation period [1, 10]. By the end of the vegetation period, 21–46 % of N, 36–50 % of P, 14–30 % of K and 27 % Mg accumulated in the aerial parts of the plants is translocated into the rhizomes [11]. The aim of the study was to determine the effect of hilling in Miscanthus giganteus in autumn after its seeding on growth, yielding and chemical content of the yield as well as on the energy value of this crop.

Materials and methods

In 2005–2007 in Pawlowice near Wroclaw, Poland, field experiments were conduct- ed on the follow-up effect of autumn hilling of Miscanthus after seeding. The split-plot experiment was set for three following variable factors: I. Harvest dates: a – autumn harvest after the vegetation period ended – 11.07.2005 and 14.12.2006; II. Autumn treatment of rhizomes after seeding: a – with hilling, b – without hilling; III. N Fertilization: 100, 150 and 200 kg per ha–1. The experiments were set on a very light river alluvial soil with loose sand and sandy gravel. In 2004–2006 soil pH was acid to very acid with the following concentrations of microelements: P – very high, K – medium to high, Mg – very low. Important features of Miscanthus growing in the year of seeding: – harvest date: 07.05.2004, – depth of rhizome seeding: 5–10 cm, – row density: 70 cm, – plant density in a row: 45 cm, – number of rhizomes planted in a plot: 16 pieces, – fertilization [kg × ha–1]: N – 60, × –1 – fertilization in all the years of the experiment [kg ha ]: P2O5 – 60 (26 – P); K2O– 100 (83 – K), – size of a plot: 5.04 m2, – harvest date: 03.12.2004. In 2004–2006 weed control was carried out – Roundup 360 SL at the dose of 4dm3 × ha–1 was applied focally. The plots were sporadically infested with White goosefoot (Chenopodium album L., Shepherd’s purse (Capsella bursa-pastoris L.) and Common barnyard grass (Echinochloa crus-galli L.). From the vegetation start in 2005 and 2006 observations and measurements were conducted on the growth and height of plants as well as diseases and insect infestations 1602 Marcin Kozak et al were monitored. After the vegetation period had ended, the shoots were numbered on every plot, the results were calculated per 1 m2. From 10 shoots from every plot the following features were measures: plants’ height, flag-leaf length, and stem diameter (on the height of 10 cm above the soil surface). Autumn and winter harvests were done manually with use of a chainsaw for hedges and a circular saw. After harvest, Miscanthus green matter yield was measured from every plot. Chemical analyses were conducted in the laboratory of the Department of Crop Production at the Wroclaw University of Environmental and Life Sciences. The following parameters were determined: – dry matter – through drying of minced plant material for 4 hours at 105 oC, – crude ash – by combusting plant material in 600 oC in an electric oven, – total nitrogen content with Kjeldahl’s method, – mineral compounds contents: K, Ca (flame photometry), P, Mg (coulometry). Based on the results obtained from the chemical analyses, water percentage content in the plant material and dry matter yield were calculated. Additionally, the contents of crude ash and of the investigated macronutrients were determined. Energy value of biomass was measured in the Institute of Agricultural Engineering, Wroclaw University of Environmental and Life Sciences. The analysis was conducted with use of a semi-automatic calorimeter Precyzja-BIT KL-10 which is designed to measure heat in solid fuels combustion such as turf, lignite, coal, coal and lignite pellets, coke and non-explosive, combustive organic substances. The method of measure is compatible with the requirements of the official Polish Standard for measuring heat of combustion and calculating of calorific value of solid fuels from 1981 [12]. On the basis of the research the relationship between biomass energy value and moisture was determined (Figs. 1 and 2).

20 000 17 969 18 000

16 000

14 000

12 000

–1

× 10 000

[MJ t ] 8 000

6 000 5 389

4 000

2 000 y = –179.71x + 17969

0 0 10 20 30 40 50 60 70 80 Moisture [%]

Fig. 1. The relationship between energy value and biomass moisture content (autumn harvest) Follow-up Effect of Hilling on Growth and Yielding of Miscanthus... 1603

20 000 18 303 18 000

16 000

14 000

12 000

–1

× 10 000

[MJ t ] 8 000

6 000 5 489

4 000

2 000 y = –183.06x + 18303

0 0 10 20 30 40 50 60 70 80 Moisture [%]

Fig. 2. The relationship between energy value and biomass moisture content (winter harvest)

In 2004–2006 the Department of Plant Protection, Wroclaw University of Environ- mental and Life Sciences, monitored diseases and insect infestation in Miscanthus. Low numbers of phytophagous organisms were observed in all the years of the experiment. A higher infestation was noted by aphids: the bird cherry-oat aphid Rhopalosiphum padi L. and rose-grain aphid Metopolophium dirhodum Walk, and thrips: Frankliniella tenuicornis Uzel., and Haplothrips aculeatus Fabricius. In 2005–2006 on Miscanthus leaves was sparsely recorded one fungi species: Alternaria alternata (Fr.) Keissl.

Results and discussion

Before the autumn harvest 03.12.2004 it was calculated that for 1 m2, Miscanthus produced 14 shoots with a mean length of 112 cm and the stem diameter of 6.7 mm, and a 66 cm long flag-leaf. Green matter yield was 3.8 t × ha–1 with 64 % water content, and dry matter yield was 1.37 t × ha–1. In 2005 and 2006 the vegetation started on April 20, and ended on November 10, 20, respectively. In these two years of the experiment, the mean monthly temperatures from April to October, except for August, were higher than multiyear means. The sum of precipitations was similar to the mean values of a multiyear. It needs to be underlined that the pattern of precipitation was unfavourable (Table 1). So far the research has indicated that Miscanthus starts growing in spring when the soil temperature reaches 10–12 oC. Air temperature of 5–10 oC is a threshold from which leaves start growing [1, 13]. Despite a low transpiration ration of 300 dm3 of water per kg d.m., Miscanthus has high water requirements because of a high production of biomass [2]. According to Beale et al [14] Miscanthus plants use from 80 to 330 g of water per1gofdrymatter. 1604 Marcin Kozak et al Table 1 Temperature Precipitation C] and precipitation sum [mm] in 2004–2007 o Mean monthly air temperature [ I II III IV V VI VII VIII IX X XI XII Month 2004200520062007 –2.9 2.1 5.9 –6.02004 –1.52005 4.9 4.8 –1.92006 1.7 2.72007 9.8 0.6 9.8 36.6 13.2 6.5 9.9 41.7 14.3 32.8 16.7 10.9 23.5 14.3 39.1 16.9 54.9 52.0 18.6 16.2 39.3 18.5 9.3 19.7 21.5 59.0 19.6 19.2 22.1 23.4 25.5 17.7 39.1 48.8 14.4 19.2 51.1 17.3 121.0 15.2 43.9 10.5 2.7 18.9 15.9 16.1 36.3 66.1 9.9 50.3 12.9 56.6 4.9 11.0 109.3 33.0 69.2 3.3 12.0 51.0 8.3 2.0 6.7 25.8 166.7 92.4 0.8 20.2 2.8 4.3 51.4 17.6 52.8 5.4 1.0 77.7 57.9 46.1 26.3 15.8 68.3 21.7 95.9 35.2 53.9 21.0 Multiyear means for 1970–2000 –1.0 0.1 3.7Multiyear sums for 1970–2000 8.1 30.5 13.9 24.8 16.7 33.2 18.5 31.9 17.7 49.9 13.3 64.9 8.8 75.4 3.6 63.5 0.5 44.7 35.5 33.9 36.3 Year Follow-up Effect of Hilling on Growth and Yielding of Miscanthus... 1605

There was no correlation noted between the investigated factors and the growth of Miscanthus during the vegetation period. Therefore, mean measures from the particular years of the experiment were used to describe the plant growth as a function of time. From the vegetation start to mid-June, Miscanthus reached 80 % of its full height (Figs. 3 and 4). In a two-year experiment conducted in Greece, where the plants were watered every 6–7 days, Miscanthus shoots were growing on average 3 cm per 24 hours from May to the end of June, reaching the height of 170 cm. In the next months the shoots

[cm] 250 233 y = – 0.0063x2 + 2.3945x 2 215 R = 0.9897 218 . 200 188 190

143 150 160

116 100 95

61 50

0 20 27 4 11 18 25 1 8 15 22 29 6 13 20 27 3 10 17 24 31 7 14 21 28 5 12 19 26 2 9 16 April May June July August September October November

Fig. 3. Growth of Giant Miscanthus (Miscanthus x giganteus Greef et Deu.) in 2005

[cm] 350 y = – 0.009x2 + 3.4271x 315 R2 = 0.9917 298 300 318 322

250 227 252 205 200

150 165

100 112

85 50

0 20 27 4 11 18 25 1 8 15 22 29 6 13 20 27 3 10 17 24 31 7 14 21 28 5 12 19 26 2 9 16 April May June July August September October November

Fig. 4. Growth of Giant Miscanthus (Miscanthus x giganteus Greef et Deu.) in 2006 1606 Marcin Kozak et al grew from 0.5 to 1 cm per 24 h until they reached their maximal height in August. The full length of shoots was 233 cm in the first year after seeding and 323 cm in the second year [15]. The number of shoots before harvest and the morphological features of the plants were affected mostly by N fertilization, and later to a lesser and lesser degree by hilling and harvest dates (Table 2). Autumn hilling, compared to the control, considerably increased the height of plants, the length of the flag with leaf and the stem diameter. Because of dynamic growth of Miscanthus in the second and the third year after seeding, significant differences were recorded in the number of shoots per 1 m2, the height of plants and the length of the flag-leaf between the years of the experiment.

Table 2

Number of shoots before harvest and morphological features of Miscanthus plants (Miscanthus x giganteus Greef et Deu.) in 2005–2006 (means for factors 2005–2006)

Number Flag-leaf Harvest Type N rate Plant height Stem diameter of shoots length date of growing [kg × ha–1] [cm] [mm] per1m2 [cm] Autumn 39 280 46.0 9.4 Winter 40 276 46.1 9.2 LSD (a = 0.05) NSD NSD NSD 0.15 with hilling 39 288 48.2 9.5 without hilling 40 268 43.8 9.2 LSD (a = 0.05) NSD 6 1.0 0.25 100 37 266 44.5 9.1 150 41 278 46.3 9.2 200 41 290 47.3 9.6 LSD (a = 0.05) 2 6 1.6 0.19 2005 25 233 55.1 9.3 Years 2006 54 322 37.0 9.3 LSD (a = 0.05) 4 4 1.3 NSD NSD – no significant difference.

In the environment of Greece, the number of shoots per 1 m2 from May to June was: 80–85 in the first year and 90–100 in the second year. Later it gradually decreased down to 75 shoots [15]. Jezowski et al [16] stated that Miscanthus yielding depends mostly on the growth stage of the plants, their branching and the diameter of the clump. Miscanthus green and dry matter yields as well as energy yield and water content depend mostly on the harvest date. In the autumn harvest, the dry matter yield was higher by 23 % than in the winter harvest, but it also contained 23.8 % more water and its energy value per 1 kg of green matter was 34 % lower (Table 3). Hilling increased, among other things, dry matter yield by 75 %. The highest dry matter yields were obtained with 150 kg N × ha–1 fertilization. Follow-up Effect of Hilling on Growth and Yielding of Miscanthus... 1607 ] –1 ha Table 3 × ] [toe –1 energy value ha × ] –1 ha × yield [t ] [GJ –1 ha × ] [GJ energy value –1 t × [GJ Green matter Dry matter Miscanthus: autumn harvest: 17.97 GJ; winter harvest: 18.30 GJ. [%] water content n1tof Greef et Deu.) in 2005–2006 (means for factors in 2005–2006) ] –1 ha × yield [t ] –1 ha × 100150200 27.2 29.8 29.2 40.7 41.0 39.4 10.77 10.72 11.02 279 305 300 15.4 16.8 16.5 279 305 300 6.66 7.28 7.16 20052006 20.7 36.8 42.6 38.1 10.43 11.24 199 390 11.0 21.5 199 390 4.75 9.31 N rate Miscanthus x giganteus [kg ( Green and dry matter yields, water content in green matter and energy value in Miscanthus Type of growing with hillingwithout hilling 29.6 27.8 40.3 40.4 10.85 10.82 305 284 16.8 15.7 305 284 7.28 6.78 = 0.05)= 0.05) 1.4 3.1 1.2 NSD 0.57 NSD 12 6 0.6 0.3 12 6 0.27 0.15 = 0.05) 1.4 3.1 0.57 12 0.6 12 0.27 = 0.05) 1.3 NSD NSD 14 0.8 14 0.33 a a a a Harvest date AutumnWinterLSD ( LSD ( 36.9 20.6 52.3 28.5 8.58 13.09 322 267 17.9 14.6 322 267 7.68 6.38 Years LSD ( LSD ( NSD – no significant difference; dry matter calorie content i 1608 Marcin Kozak et al

Many researchers [1, 2] claim that the quality of Miscanthus biomass for combustion and the yield are determined by the harvest date. The autumn harvest yield is higher than from the spring harvest, and the losses can range from 14 to 30 % [5]. The disadvantage of the autumn harvest is not only high moisture of the obtained biomass but also higher removal (accumulation) of the nutrients from the fields [6]. According to several researchers [1, 11] N fertilization at the rate of 50–70 kg × ha–1 × year–1 is sufficient for Miscanthus’s requirements when applied in spring at the beginning of shooting. Kaack and Schwarz [17] argue that high doses of N in growing M. x giganteus (over 75 kg of N × ha–1) can result in an increased lodging, on fields not sheltered from wind. The harvest dates and the age of the plantation affected chemical content of Miscanthus biomass. Harvested in winter, Miscanthus contained 40 % less ash and 25–48 % less of all the investigated macronutrients than the crops harvested in autumn. This was due to a translocation of nutrients from shoots to rhizomes and their draining with rain during the winter period.

Table 4

Chemical composition of Miscanthus biomass [%] (Miscanthus x giganteus Greef et Deu.) (means for factors in 2005–2006)

Harvest Type N rate Crude ash NPKCaMg date of treatment [kg × ha–1] Autumn 3.16 0.31 0.06 0.43 0.23 0.08 Winter 1.79 0.23 0.04 0.22 0.12 0.06 LSD (a = 0.05) 0.14 0.02 0.01 0.02 0.03 0.01 with hilling 2.49 0.27 0.05 0.32 0.18 0.07 without hilling 2.46 0.27 0.05 0.33 0.17 0.07 LSD (a = 0.05) NSD NSD NSD NSD NSD NSD 100 2.55 0.27 0.05 0.33 0.17 0.07 150 2.41 0.27 0.05 0.32 0.18 0.07 200 2.46 0.27 0.05 0.32 0.18 0.07 LSD (a = 0.05) NSD NSD NSD NSD NSD NSD 2005 3.04 0.28 0.07 0.48 0.26 0.07 Years 2006 1.91 0.26 0.03 0.17 0.09 0.07 LSD (a = 0.05) 0.14 NSD 0.01 0.02 0.03 NSD NSD – no significant difference.

According to the studies conducted by Lewandowski et al [18] a delayed harvest decreased on average ash content in Miscanthus by 28 % in Portugal and the Great Britain, by 42 % in Germany, and by 50 % in Sweden and 54 % in Denmark. All the agrotechnical factors and the age of the stand affected the accumulation of crude ash and the investigated macronutrients in Miscanthus biomass (Table 5). The winter harvest lowered crude ash content by 50 % and nutrient content by 40–58 % compared with the autumn harvest. Follow-up Effect of Hilling on Growth and Yielding of Miscanthus... 1609 Table 5 Greef et Deu.) Miscanthus x giganteus ]( –1 ha × Crude ash N P K Ca Mg ] –1 ha × 100150200100 513150 566200 541 496 53.1 458 57.6 538 59.9 50.5 7.87 53.7 9.86 54.0 9.07 7.92 57.3 8.83 62.7 8.89 65.9 60.6 29.8 64.2 35.1 67.1 38.6 29.9 14.2 37.1 16.7 34.5 13.9 13.1 13.4 13.4 100150200100 248150 264200 240 238 30.7 239 35.2 242 32.2 31.6 4.70 33.9 4.22 32.8 3.92 5.06 29.3 4.80 29.6 4.49 27.1 28.3 13.2 27.9 16.0 28.3 14.5 14.7 9.3 13.7 7.5 14.6 10.1 9.0 8.3 8.2 N rate [kg with hilling without hilling with hilling without hilling Accumulation of crude ash and macrocomponents in Miscanthus biomass [kg = 0.05) 35 NSD 0.5 NSD 1.7 1.5 a Harvest date Type of growing Autumn Winter LSD ( 1610 Marcin Kozak et al Table 5 contd. Means for factors Crude ash N P K Ca Mg ] –1 ha × 100150200 374 382 390 41.5 45.1 44.7 6.38 6.93 6.59 43.9 46.1 47.1 21.9 25.5 25.6 11.4 11.5 11.4 20052006 340 423 30.8 56.8 7.58 5.69 53.6 37.7 29.5 19.1 7.8 15.1 N rate [kg with hillingwithout hilling 395 369 44.8 42.8 6.60 6.66 45.3 46.1 24.6 24.1 11.9 10.9 = 0.05)= 0.05) NSD 2.1 12 0.25 1.6 1.6 0.17 0.9 1.1 0.8 0.6 NSD = 0.05) 6 0.8 NSD 0.7 0.4 NSD = 0.05) 12 1.6 0.17 1.1 0.6 0.6 a a a a Harvest date Type of growing LSD ( Years LSD ( LSD ( AutumnWinterLSD ( 519 245 54.8 32.7 8.74 4.53 63.0 28.4 34.2 14.5 14.1 8.7 NSD – no significant difference. Follow-up Effect of Hilling on Growth and Yielding of Miscanthus... 1611

In the experiments carried out by Clifton-Brown et al [19] on a 16-year old Miscanthus stand, it was recorded that the contamination of nutrients in the autumn harvest was as follows: 145 ± 9.4 kg N × ha–1,23± 1.1 kg P × ha–1 and 111 ± 9.9 kg K × ha–1. When the harvest date was moved to March, the concentration of nutrients was lowered: 51 ± 6.1 kg N × ha–1, 8.3 ± 0,7 kg P × ha–1 and 42 ± 7.9 kg K × ha–1. The mean yields for 15 years were as follows: 13.4 ± 1.1 kg d.m. × ha–1 year–1 for autumn harvests and 9.0 ± 0.7 kg d.m. × ha–1 year–1 in winter harvests.

Conclusions

1. In the first years of the experiment on Miscanthus, morphological features, d.m. yield, water and ash contents depended on the age of the stand. 2. Harvest dates proved to have the most significant effect on Miscanthus yielding among all the investigated factors. Compared with the autumn harvest, the winter harvest lowered d.m. yield by 18.4 % and water content by 23.4 % in green matter. With winter harvest crude ash content also decreased by 43 %, while green biomass energy value increased by 52 %. 3. It is possible to obtain a mean biomass energy yield of 294 GJ (which corresponds to 7.03 toe*) from 1 ha of Miscanthus field in the second and the third year of the cultivation. 4. It is possible to obtain high d.m. yields in Miscanthus with autumn hilling and by applying 150 kg N × ha–1. 5. Although the winter harvest results in lower biomass yields, it is more favourable in terms of quality of the plant material for energy production than the autumn harvest.

References

[1] Lewandowski I., Clifton-Brown J.C., Scurlock J. M.O. and Huisman W.: Miscanthus: European experience with a novel energy crop. Biomass and Bioenergy 2000, 19(4), 209–227. http://dx.doi.org/10.1016/S0961-9534(00)00032-5. [2] Je¿owski S.: Energia w trawie. Czysta Energia 2004, 5(33), 15–16. [3] Kozak M.: Mo¿liwoœci uprawy i wykorzystania Miskanta olbrzymiego na cele energetyczne w Polsce. Cz. II, Ekonatura, Wroc³aw 2006, 3, 20–22. [4] Roszewski R.: Miskant olbrzymi – Miscanthus sinensis giganteus, Nowe roœliny uprawne na cele spo¿ywcze, przemys³owe i jako odnawialne Ÿród³a energii. SGGW, Warszawa 1996, 123–135. [5] Clifton-Brown J.C. and Lewandowski I.: Screening Miscanthus genotypes in field trials to optimize biomass yield and quality in Southern Germany. Eur. J. Agron. 2002, 16, 97–110. http://dx.doi.org/10.1016/S1161-0301(01)00120-4. [6] Lewandowski I. and Heinz A.: Delayed harvest of miscanthus – influences on biomass quantity and quality and environmental impacts of energy production. Eur. J. Agron. 2003, 19, 45–63. http://dx.doi.org/10.1016/S1161-0301(02)00018-7. [7] Kahle P., Beuch S., Boelcke B., Leinweber P. and Schulten H.-R.: Cropping of Miscanthus in Central Europe: biomass production and influence on nutrients and soil organic matter. Eur. J. Agron. 2001, 15, 171–184. http://dx.doi.org/10.1016/S1161-0301(01)00102-2.

* toe – ton of oil equivalent (fuel with caloric value of 41.87 GJ × t–1). 1612 Marcin Kozak et al

[8] Lewandowski I. and Kicherer A.: Combustion quality of biomass: practical relevance and experiments to modify the biomass quality of Miscanthus x giganteus. Eur. J. Agron. 1997, 6, 163–177. http://dx.doi.org/10.1016/S1161-0301(96)02044-8. [9] Lewandowski I.: Propagation method as an important factor in the growth and development of Miscanthus x giganteus. Ind. Crops and Prod. 1998, 8, 229–245. http://dx.doi.org/10.1016/S0926-6690(98)00007-7. [10] Beale C.V. and Long S.P.: Seasonal dynamics of nutrient accumulation and partitioning in the perennial C4-grasses Miscanthus x giganteus and Spartina cynosuroides. Biomass and Bioenergy 1997, 12(6), 419–428. http://dx.doi.org/10.1016/S0961-9534(97)00016-0. [11] Himken M., Lammel J., Neukirchen D., Czypionka-Krause U. and Olfs H.W.: Cultivation of Miscanthus under West European conditions: Seasonal changes in dry matter production, nutrient uptake and remobilization. Plant and Soil 1997, 189, 117–126. http://dx.doi.org/10.1023/A:1004244614537. [12] Polska Norma. PN-G-04513:1981 Paliwa sta³e. Oznaczanie ciep³a spalania i wartoœci opa³owej. 1981. [13] Clifton-Brown J.C. and Jones M.B.: The thermal response of leaf extension rate in genotypes of the C4-grass Miscanthus: an important factor in determining the potential productivity of different genotypes. J. Exp. Bot. 1997, 48 (313), 1573–1581. http://dx.doi.org/ 10.1093/jxb/48.8.1573. [14] Beale C.V., Morison J.I.L. and Long S.P.: Water use efficiency of C4 perennial grasses in a temperate climate. Agricult. Forest Meteorol. 1999, 96, 103–115. http://dx.doi.org/10.1016/S0168-1923(99)00042-8. [15] Danalatos N.G., Archontoulis S.V. and Mitsios I.: Potential growth and biomass productivity of Miscanthus x giganteus as affected by plant density and N-fertilization in central Greece. Biomass and Bioenergy 2007, 31(2–3), 145–152. http://dx.doi.org/10.1016/j.biombioe.2006.07.004. [16] Je¿owski S., G³owacka K. and Bocianowski J.: Zmiennoœæ wybranych klonów traw olbrzymich z rodzaju Miscanthus pod wzglêdem plonowania w pierwszych latach uprawy. Zesz. Probl. Post. Nauk Roln. 2007, 517, 339–348. [17] Kaack K. and Schwarz K.U.: Morphological and mechanical properties of Miscanthus in relation to harvesting, lodging, and growth conditions. Ind. Crops and Prod. 2001, 14, 145–154. http://dx.doi.org/10.1016/S0926-6690(01)00078-4. [18] Lewandowski I., Clifton-Brown J.C., Andersson B., Basch G., Christian D.G., Jorgensen U., Jones M.B., Richie A.B., Schwarz K.U., Tayebi K. and Teixeira F.: BIOFUELS, Environment and Harvest Time Affects the Combustion Qualities of Miscanthus Genotypes. Agron. J. 2003, 95, 1274–1280. http://dx.doi.org/10.2134/agronj2003.1274. [19] Clifton-Brown J.C., Breuer J. and Jones M.B.: Carbon mitigation by the energy crop Miscanthus. Global Change Biol. 2007, 13, 2296–2307. http://dx.doi.org/10.1111/j.1365-2486.2007.01438.x.

NASTÊPCZY WP£YW OBREDLANIA NA ROZWÓJ I PLONOWANIE MISKANTA OLBRZYMIEGO (Miscanthus x giganteus Greef et Deu.)

Katedra Szczegó³owej Uprawy Roœlin Uniwersytet Przyrodniczy we Wroc³awiu

Abstrakt: W latach 2005–2007 w Paw³owicach ko³o Wroc³awia prowadzono badania polowe nad nastêpczym wp³ywem obredlania miskanta olbrzymiego jesieni¹, po posadzeniu. Doœwiadczenie za³o¿ono w uk³adzie „split–plot” na trzy czynniki zmienne, którymi w kolejnoœci by³y: I. Terminy zbioru: a – jesienny po zahamowaniu wegetacji – 11.07.2005 r., 14.12.2006 r., b – zimowy przed ruszeniem wegetacji – 10.03.2006 r., 09.03.2007 r., II. Jesienna pielêgnacja k³¹czy po posadzeniu a – z obredlaniem, b – bez obredlania, III. Nawo¿enie w kg N × ha–1: 100, 150 i 200. W pocz¹tkowych latach po za³o¿eniu doœwiadczenia z miskantem olbrzymim cechy morfologiczne, plon suchej masy, zawartoœæ wody i popio³u zale¿a³y od wieku plantacji. Spoœród badanych czynników agrotechnicznych najwiêkszy wp³yw na poziom plonu i jego jakoœæ mia³ termin zbioru. Zimowy zbiór, w porównaniu z jesiennym, spowodowa³ zmniejszenie: plonu suchej masy o 18,4 %, zawartoœci wody w œwie¿ej masie o 23,8 % i popio³u surowego 43 % oraz wzrost wartoœci Follow-up Effect of Hilling on Growth and Yielding of Miscanthus... 1613 energetycznej œwie¿ej masy o 52 %.Z1hauprawy miskanta olbrzymiego w drugim i trzecim roku uprawy mo¿na uzyskaæ plon biomasy o œredniej wartoœci energetycznej 294 GJ, co odpowiada 7,03 toe. Du¿e plony suchej masy miskanta olbrzymiego mo¿na uzyskaæ przy jesiennym obredlaniu plantacji i zastosowaniu 150 kg N × ha–1. S³owa kluczowe: miskant olbrzymi, terminy zbioru, obredlanie, nawo¿enie azotem

ECOLOGICAL CHEMISTRY AND ENGINEERING A

Vol. 18, No. 12 2011

El¿bieta SACA£A1

MISCANTHUS – UNUSUAL GRASS: BIOCHEMICAL AND PHYSIOLOGICAL CHARACTERISTIC: A REVIEW

MISCANTHUS – TRAWA NIEZWYK£A: CHARAKTERYSTYKA BIOCHEMICZNO-FIZJOLOGICZNA: PRZEGL¥D LITERATUROWY

Abstract: Miscanthus × giganteus (Giant Miscanthus) is one of the most promising plants cultivated for biomass production. In spite of its origin from south-east Asia and being warm adapted plant it grows well and produces high biomass in temperate latitudes. Miscanthus × giganteus is a C4 plant and hence this study presents a brief description of C4 photosynthesis and the enzymes involved in this process. On the basis of data from current literature, the biochemical bases of relatively high tolerance of miscanthus to cold temperatures (0–15 oC) were evaluated. Moreover, it was reviewed numerous ecophysiological features of Miscanthus × giganteus were reviewed (high productivity, low fertiliser and pesticides requirements, possibility to use in phytoremediation) which showed that it is a proecological and environmentally friendly crop. This causes that Miscanthus × giganteus might be recognize as a leading crop in non-food cultivations.

Keywords: miscanthus, C4 photosynthesis, chilling tolerance, phytoremediation, allelopathy

Introduction

Plants from genus of Miscanthus originate from south-east Asia and are widely distributed in Asia and Pacific Islands. Miscanthus belongs to Poaceae family and tribe of Andropogoneae [1]. Miscanthus is rhizomatous perennial grass reaching height of 5 m and offers very high biomass yield. In 1935 it was introduced into Europe and initially was cultivated in parks and gardens as ornamental plants. Over the past 20 years, there is increasing interest in Miscanthus as a raw material for industry (paper production, insulation material) and renewable source of energy. Heating value of Miscanthus is about 17–19 MJ × kg–1 and is similar to energetic value of wood fuel and it accounts for about 60 % of that of hard coal [2, 3].

1 Department of Plant Nutrition, Wroclaw University of Environmental and Life Sciences, ul. Grun- waldzka 53, 50–357 Wroc³aw, Poland, email: [email protected] 1616 El¿bieta Saca³a

Plants from Miscanthus genus are frequently so-called Chinese Grass or Elephant Grass. The most famous representatives of these grasses are: Miscanthus sinensis, Miscanthus sacchariflorus and Miscanthus × giganteus (Giant Miscanthus). Miscanthus × giganteus is a sterile triploid (2n = 3x = 57) arising through natural crossing of tetraploid Miscanthus sacchariflorus (2n = 4x = 76) and diploid (2n = 2x = 38) Miscanthus sinensis [4]. Generated hybrid is sterile, does not produce seeds and hence it must be propagated vegetatively by rhizome division or in vitro cultures. Miscanthus × giganteus displays a good balance of traits from each parent, combining rapid growth with a tolerance to low temperatures [5]. Miscanthus × giganteus (hereafter referred to as miscanthus) is a robust woody perennial which may be cultivated at one site for 15–20 years. It reaches full productivity over the first 3 years after planting. Miscanthus appears in numerous genotypes [3, 6]. Miscanthus is closely related to three important agricultural crops: sugarcane (Saccharum officinarum), sorghum (Sorghum bicolor) and maize (Zea mays). All these crop species are C4 plants and exhibit high productivity particularly in warmer and well radiated regions.

Miscanthus –C4 plant (biochemical characteristic)

C4 plants have evolved mechanisms to improve photosynthetic efficiency and decrease water loss in hot, dry environments. In Miscanthus, as in other C4 plants, CO2 fixation is a two-step process. CO2 is initially fixed in the cytosol of mesophyll cells surrounding the bundle sheath [7]. This reaction is catalyzed by phosphoenolopyruvate carboxylase (PEPC; EC 4.1.1.31). PEPC is homotetrameric enzyme that has high affinity to CO2 and is activated by divalent cation [8]. PEPC can fixe CO2 at very low its b concentration and may use CO2 produced in respiration. PEPC catalyses the -carbo- – xylation of phosphoenolopyruvate using HCO3 as substrate in a reaction that yields oxaloacetate (a four carbon dicarboxylic compound) and phosphate (Pi): – ® phosphoenolopyruvate (PEP) + HCO3 oxaloacetate (OAA) + Pi. Then oxaloacetate is converted to malate by NADPH-malate dehydrogenase:

Oxaloacetate + NADPH + H+ ® malate + NADP+.

Formed malate is transported by plazmodesmata to the bundle sheath cells, where it is decarboxylated in reaction catalysed by NADP-malic enzyme (NADP-ME): + ® + malate + NADP CO2 + pyruvate + NADPH + H .

Released CO2 is fixed by Rubisco (ribulose-1,5-bisphosphate carboxylase/oxygenase; EC 4.1.1.39) and converted to carbohydrate in the Calvin cycle. Pyruvate (C3 acid) diffuses back to the mesophyll cells to regenerate phosphoenolopyruvate – primary acceptor of CO2:

pyruvate + ATP + Pi ® phosphoenolopyruvate + AMP + PPi. Miscanthus – Unusual Grass: Biochemical and Physiological Characteristic: A Review 1617

This reaction is very important because it leads to regeneration of primary CO2 acceptor and allows to remaining its concentration at suitable level. Reaction occurs in mesophyll chloroplasts and is catalyzed by pyruvate orthophosphate dikinase (PPDK, EC 2.7.9.1). Enzyme action consumes two high-energetic bounds of ATP and requires orthophosphate. PPDK activity is regulated by light. Fully active PPDK is a tetramer that may dissociate at low temperature and it leads to a loss of its activity. Divalent cations (Mn2+,Mg2+,Ca2+), phosphoenolopyruvate, polyols (glycerol, sorbitol) and proline protect PPDK against cold inactivation [9–11]. It is worth to note that among the C4 plants there are three subtypes characterized according to the type of decarboxylating enzyme in the bundle sheath cells. Miscanthus belongs to described above the NADP-malic enzyme (NADP-ME) pathway. Plants using this pathway are characterized by very high nitrogen use efficiency (NUE). The physiological significance of two-step CO2 fixation process is increasing CO2 concentration in cells of bundle sheath (it is estimated 10-fold over atmospheric concentration). This leads to elimination of oxygenase activity of Rubisco and carbon loss in photorespiration process. As a result, in favourable environments, C4 plants are the most productive crops. They show high photosynthetic rates (both interception of solar radiation and CO2 fixation) and higher water and nitrogen use efficiency. The field trials performed with Miscanthus × giganteus and Triticosecale (C3 plant) showed that – NUE, defined as the ratio of biomass yield to N supply (sum of soil NO3 and N fertilization) were 0.35 and 0.14 Mg dry biomass per kg N, respectively for miscanthus and triticale [12]. C4 plants have high rates of photosynthesis and productivity but this is realized in favourable conditions – high radiation and warm, humid environments. In temperate zones, low temperature is a major constraint limiting rate of photosynthesis and consequently productivity of C4 plants. Miscanthus × giganteus appears to be unusual o among C4 species and cultivated at 52 northern latitude produces 30 tonnes of dry matter per hectare per year, exceeding the most productive C3 crops [13].

Cold tolerance of Miscanthus × giganteus

Low temperature is a major factor that influences plant metabolism and physiology and affects its productivity. The most of C4 species influenced by chilling temperatures o (0–15 C) react by apparent reduction of CO2 assimilation and consequently, utilization of absorbed solar energy is limited. This leads to photoinhibition and photodamage of photosynthetic apparatus. Miscanthus × giganteus is relatively tolerant to chilling temperatures (avoids these damages) and similarly to maize may be cultivated in temperate climates. Nevertheless, in comparison with maize, miscanthus considerably better tolerates suboptimal conditions, particularly too low temperatures in spring and o autumn [14]. Maize growing at 14 C shows 90 % reduction in CO2 uptake comparing with plants grown at 25 oC, whereas in Miscanthus × giganteus this negative response does not happen [15, 16]. Some researchers have postulated that Rubisco is the most likely candidate for limiting C4 photosynthesis at chilling temperatures [17]. However experiments conducted by Wang et al [18] on Rubisco from Miscanthus × giganteus 1618 El¿bieta Saca³a grown at 14 and 25 oC and Zea mays grown at 25 oC showed that there were no sign- ificant differences in enzyme catalytic proprieties. Rubisco did not differ significantly, either between the two species or between growth temperature. These results suggest that higher cold tolerance of miscanthus than maize does not result from Rubisco activity [18]. Wang et al [11] concluded that pyruvate orthophosphate dikinase (PPDK) is responsible for greater photosynthetic capacity in miscanthus than other C4 species cultivated under suboptimal temperature. In experiments of Wang et al [11], lowering of growth temperature from 25 to 14 oC caused different reaction in miscanthus and maize. On the first day at low temperature, PPDK protein declined slightly in miscanthus but then accumulated above the initial level and was nearly doubled after 7 days, whereas Rubisco level did not change significantly. In contrast to miscanthus, PPDK in maize leaves declined throughout in chilling period and Rubisco level was also significantly reduced. Naidu et al [19] obtained similar results. In miscanthus low temperature caused accumulation of PPDK protein while level of Rubisco remained unaffected. Whereas in maize leaves there was recorded decrease in quantity and activity of both crucial enzymes – PPDK and Rubisco. Concluding, it can be stated that increases in either protein content and PPDK activity in leaves of C4 plants growing in chilling conditions may be one of the mechanisms increasing their tolerance to low temperatures. As mentioned above, low temperatures can disrupt balance between absorbed excitation energy of light and its utilization by photosynthetic apparatus [20]. Low temperatures decrease rates of CO2 uptake and assimilation, and in turn, plant demand for ATP and NADPH is markedly reduced. Consequently, it leads to secondary effects associated with formation and accumulation of reactive oxygen species (ROS). The ·– · most important ROS are: superoxide (O2 ), hydroxyl radical (OH ), hydrogen peroxide 1 (H2O2) and singlet oxygen ( O2). These molecules are potentially harmful and may attack cellular components (macromolecules and membranes) and cause serious damage of photosynthetic apparatus (photoinhibition), ultimately leading to severe cellular damage. Plants have the capacity to eliminate ROS and antioxidant molecules such as ascorbic acid, carotenoids, glutathione play important role in this protective system. Carotenoids, lipid-soluble pigments presented in chloroplasts, have the ability to dissipation of surplus excitation energy and act as quencher of its excess and hence they are the main compounds involved in preventing photoinhibition. A particular class of carotenoids – the xananthopylls – play a key role in protection against oxidative damage. Zeaxanthin, the de-epoxidised form, better dissipates excitation energy and is derived from the epoxidised pigments violaxantin. The rapid changes between the two forms, brought about by special enzymes, constitute the “xantophyll cycle”. It is worth to note that in chilling conditions miscanthus avoids photooxidative damages by maintaining a high rate of CO2 uptake and assimilation, and in turn increased utilization of absorbed light energy [13]. On the other hand, in this plant very efficiently operates described above mechanism of non-photochemical quenching of excitation energy. This correlates with large increase in xanthophylls content and its de-epoxidation [16]. In comparison with the other C4 plants, Miscanthus × giganteus has a lower temperature optimum for light-saturated photosynthesis [15]. Miscanthus – Unusual Grass: Biochemical and Physiological Characteristic: A Review 1619

It is worth noting that Miscanthus × giganteus shows high cold susceptibility, especially to freezing temperature, in the first year after planting. Periods of frost, typical during early spring and winter, may result in poor plants’ survival and are potential obstacles to the establishment of miscanthus in northern regions of Europe [21–23]. Plazek et al [24] suggested that frost susceptibility of miscanthus is mainly caused by sensitivity of shoot apical meristems to frost. Farrell at al [23] have investigated genotypic variation in the base temperature (Tb) for shoot emergence and in the lethal temperature for shoots in four Miscanthus genotypes. In all genotypes, lowering temperature increased the time of shoot emergence. Tb for Miscanthus × giganteus accounted 8.5 oC and was slight higher compared with Miscanthus sinensis. All examined genotypes exhibited considerable leaf damage following exposure to –8 oC and they might be classed as freezing sensitive [23]. However, taking into account the information that tropical grasses fall within the temperature range –1.8 oCto –4.3 oC [25], it might be concluded that plants from Miscanthus genus are relatively tolerant to non severe frost. In the case of Miscanthus × giganteus, the lethal temperature at which 50 % of the shoots were killed was estimated as –8 oC [23].

Other promising features of Miscanthus × giganteus

Miscanthus × giganteus has several positive ecophysiological traits causing that is considered as environmentally benignant plant: – High productivity. At temperate climate conditions miscanthus yield reaches 20–30 tonnes dry matter per hectar per year (since the second or third year after planting when the crop is well established) and it may be higher – 40 tonnes dry matter year–1 ha–1 – in southern Europe (Mediterranean regions) with irrigation [12, 26, 27]. The first side-by-side large-scale field trials conducted on Zea mays and Miscanthus × giganteus in the U.S. Corn Belt (Illinois) showed that miscanthus was 59 % more productive than modern lines of grain maize bred for high productivity [28]. Nevertheless, it is interesting that examinations concerning some important parameters of photosynthesis (rate of both phosphoenolopyruvate carboxylation and phospho- enolopyruvate regeneration, quantum efficiency of CO2 assimilation) demonstrated that they were higher in maize than in miscanthus. However, these biochemical and photochemical traits of maize were not enough to match up to miscanthus in productivity. Dohleman and Long [28] state that higher productivity of miscanthus in comparison with maize is related to larger leaf area and its longer growing season. Consequently, miscanthus intercepts more PAR (photosynthetically active radiation) and longer continues photosynthesis. Length of growing season (average for two years) for maize was 125 days whereas for miscanthus it was 199 days (60 % longer than for maize). Dohleman et al [29] conducted field trials to find the reasons for hugely higher biomass production of Miscanthus × giganteus in comparison with Panicum virgatum (switchgrass; such as miscanthus is C4 perennial grass considered as bioenergy crop). They concluded that there are numerous factors determined greater productivity of miscanthus: almost 40 % higher photosynthetic rates, significantly higher both nitrogen 1620 El¿bieta Saca³a and water use efficiencies, lower respiration, larger leaf canopy and its more vertical standing (more radiation penetrates into the deeper canopy). – Low fertiliser requirements. The most field trials conducted at different sites showed that mineral fertilisation (particularly nitrogen supply) does not improve significantly yield of miscanthus especially in the case of full established plantations [26, 30–34]. In initial seasons of planting fertilisation may lead to an increase in above-ground biomass accumulation [35, 36]. Field experiments in southern regions of Europe demonstrated that nitrogen fertilisation significantly increase dry mass of aboveground biomass when water was not limiting (trials with irrigation) [36, 37]. Experiments conducted by Kalembasa et al [35] on five clones of Miscanthus, after two years of cultivation, showed that N60P50K100 fertilisation cause decrease in dry mass of underground parts (rhizomes + roots). Amougou et al [33] stated that underground biomass was not significantly affected by nitrogen fertilization (120 kg N ha–1 year–1) but it tended to decrease under N fertilization, particularly on 2-year-old miscanthus stand. It is noteworthy that long-term trials (14 years) conducted in different locations in Europe also demonstrate that nitrogen fertilisation does not improve growth and biomass productivity of Miscanthus × giganteus [32]. This unpredictable response of miscanthus to nitrogen application results from very efficient minerals’ movement within the plant during the growing season. Rhizomes act as a storage organ and in autumn nitrogen and other nutrients are translocated from aboveground parts (stem + leaves) to rhizome. In subsequent spring season, nutrients stored in rhizome are remobilized and transported into new growing shoots. This causes that miscanthus is partly independent of the actual nutrient supply from soil [26, 38]. Kalembasa et al [39] investigated 5 clones of Miscanthus in second year of cultivation, at five different term of growing season (from June to October) and they demonstrated that total nitrogen content was the highest in June and lowered significantly in subsequent months. At the end of growing season (in October) N concentration in dry matter lowered by 5.5-fold on average for all clones comparing with its concentration in June. Effective minerals’ translocation is very profitable phenomenon also in aspect of biomass utilization and its combustion quality. Low concentration of nutrients in harvestable biomass results in reduction of undesirable compounds released during combustion such as SO2,NOx, HCl, dioxin. Delayed harvest date (over winter) reduces moisture, ash and nutrient contents in yield and hence is an effective method to improve the biomass quality for combustion [2, 22, 40]. – Poor needs for crop protection. In well established plantations (from year two or three onwards), miscanthus very effectively competes with weeds. Mechanical or chemical weed control is necessary during crop establishment [26, 32, 34]. In subsequent years, fast growing plants and leaf litter layer on the soil create conditions that significantly limit growth of other plants and hence application of herbicides is not necessary every year. It is likely that Miscanthus × giganteus such as other represent- atives of Poaceae family shows high allelopathic activity [41]. It was documented that Miscanthus transmorrisonensis and Miscanthus floridulus (species growing in Taiwan) produce phenolic compounds that inhibit growth of other plants [1, 41, 42]. Miscanthus Miscanthus – Unusual Grass: Biochemical and Physiological Characteristic: A Review 1621 is characterized by relatively high resistence to pests and diseases and chemical products against them are not necessary [26, 32, 34]. – Potential possibility to using in phytoremediation. Miscanthus ssp. may grow on acidic soils where increases bioavailability of heavy metals and in turn raises their phytotoxicity [43, 44]. Under low pH conditions aluminium (Al), the third most abundant element in earth’s crust, is a major soil mineral constituent that may restrict plant growth. It has been reported that Al promotes growth of Miscanthus sinesis and is considered as beneficial element for this plant [45]. Besides, it was shown that Miscanthus sinesis is also tolerant to other heavy metals: chromiun and zinc [44]. Aluminium presented in plant tissues may enhance herbivore defence and promote phosphorus uptake [45]. Arduini et al [46] have shown that Miscanthus × giganteus is not particularly tolerant to cadmium. Their experiments conducted in a controlled environment demonstrated that biomass production is declined by 50 % with cadmium concentrations ranging from 0.75 to 2.25 mg × dm–3 in nutrient solution and Cd concentration in roots was more than 10-fold higher than that of the shoot. However, these authors suggest that even Cd-sensitive species like miscanthus could effectively accumulate cadmium and widen the range of species using in phytoremediation. Plantations of Miscanthus ssp. may prevent and reduce soil erosion as well as stabilize and extract contaminations from soil [47, 48].

Conclusions

Miscanthus × giganteus in spite being C4 plant adapted to warmer climate, is relatively tolerant to chilling temperatures and in temperate zones its growth is fast and biomass production very high. Miscanthus × giganteus displays a good combination between high productivity and efficiencies in using of water, nitrogen and solar radiation. Besides, it shows numerous ecophysiological traits causing that it might be recognize as leading crop in non-food cultivations.

References

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[7] Furbank R.T. and Taylor W.C.: Regulation of photosynthesis in C3 and C4 plants: A molecular approach. Plant Cell 1995, 7, 797–807. 1622 El¿bieta Saca³a

[8] Chollet R., Vidal J. and Oleary M.H.: Phosphoenolpyruvate carboxylase: a ubiquitous, highly regulated enzyme in plants. Ann. Rev. Plant Physiol. Plant Mol. Biol. 1996, 47, 273–298. [9] Krall J.P., Edwards G.E. and Andreo C.S.: Protection of pyruvate, Pi dikinase from maize against cold lability by compatible solutes. Plant Physiol. 1989, 89, 280–285. [10] Salahas C., Cormas E. and Zervoudakis G.: Cold inactivation of phosphoenolpyruvate carboxylase and pyruvate orthophosphate dikinase from the C4 perennial plant Atriplex halimus. Russ. J. Plant Physiol. 2002, 49, 211–215. [11] Wang D., Portis Jr. A.R., Moose S.P. and Long S.P.: Cool C4 photosynthesis: pyruvate Pi dikinase expression and activity corresponds to the exceptional cold tolerance of carbon assimilation in Miscanthus × giganteus. Plant Physiol. 2008b, 148, 557–567. [12] Lewandowski I. and Schmidt U.: Nitrogen, energy and land use efficiencies of miscanthus, reed canary grass and triticale as determined by the boundary line approach. Agric. Ecosyst. Environ. 2006, 112, 335–346. [13] Beale C.V., Bint D.A. and Long S.P.: Leaf photosynthesis in the C4-grass Miscanthus× giganteus, growing in the cool temperate climate of southern England. J. Exp. Bot. 1996, 47, 267–273. [14] Beale C.V. and Long S.P.: Can perennial C4 grasses attain high efficiencies of radiant energy conversion in cool climates? Plant Cell Environ. 1995, 18, 641–650. [15] Naidu S.L. and Long S.P.: Potential mechanism of low-temperature tolerance of C4 photosynthesis in Miscanthus × giganteus: an in vivo analysis. Planta. 2004, 220, 145–155. [16] Farage P.K., Blowers D.A., Long S.P. and Baker N.R.: Low growth temperatures modify the efficiency of light use by photosystem II for CO2 assimilation in leaves of two chilling-tolerant C4 species, Cyperus longus L. and Miscanthus × giganteus. Plant Cell Environ. 2006, 29, 720–728. [17] Kubien D.S., von Caemmerer S., Furbank R.T. and Sage R.: C4 photosynthesis at low temperature. A study using transgenic plants with reduced amounts of rubisco. Plant Physiol. 2003, 132, 1577–1585. [18] Wang D., Naidu S.L., Portis Jr. A.R., Moose S.P. and Long S.P.: Can the cold tolerance of C4 photosynthesis in Miscanthus × giganteus relative to Zea mays be explained by differences in activities and termal properties of Rubisco? J. Exp. Bot. 2008, 59, 1779–1787. [19] Naidu S.L., Moose S.P., Al-Shoaibi A.K., Raines A.K. and Long S.P.: Cold tolerance of C4 photosynthesis in Miscanthus × giganteus: adaptation in amounts and sequence of C4 photosynthetic enzymes. Plant Physiol. 2003, 132, 1688–1697. [20] Beck E.H., Fettig S., Knake C., Hartig K. and Bhattarai T.: Specific and unspecific responses of plants to cold and drought stress. J. Biosci. 2007, 32, 501–510. [21] Allen D.J. and Ort D.R.: Impacts of chilling temperature on photosynthesis in warm-climate plants. Trends Plant Sci. 2001, 6, 36–42. [22] Clifton-Brown J.C. and Lewandowski I.: Screening Miscanthus genotypes in field trials to optimise biomass yield and quality in Southern Germany. Eur. J. Agron. 2002, 16, 97–110. [23] Farrell A.D., Clifton-Brown J.C., Lewandowski I. and Jones M.B.: Genotypic variation in cold tolerance influences the yield of Miscanthus. Ann. Appl. Biol. 2006, 149, 337–345. [24] P³a¿ek A., Dubert F. and Marzec K.: Cell membrane permeability and antioxidant activities in the rootstocks of Miscanthus × giganteus as an effect of cold and frost treatment. J. Appl. Bot. Food Quality 2009, 82, 158–162. [25] Ivory D.A. and Whiteman P.C.: Effects of environment and plant factors on foliar freezing resistance in tropical grasses. II. Comparison of frost resistance between cultivars of Cenchrus ciliaris, Chloris gayana and Setaria anceps. Austral. J. Agricul. Res. 1978, 29, 261–266. [26] Lewandowski I., Clifton-Brown J.C., Scurlock J.M.O. and Huisman W.: Miscanthus: European experience with a novel energy crop. Biomass Bioenergy 2000, 19, 209–227. [27] Zub H.W. and Brancourt-Hulmel M.: Agronomic and physiological performances of different species of Miscanthus, a major energy crop. A review. Agron. Sustain. Devel. 2010, 30, 201–214. [28] Dohleman F.G. and Long S.P.: More productive than maize in the midwest: How does Miscanthus do it? Plant Physiol. 2009, 150, 2104–2115. [29] Dohleman F.G., Heaton E.A., Leakey A.D.B. and Long S.P.: Does greater leaf-level photosynthesis explain the longer solar energy conversion efficiency of Miscanthus relative to switchgrass? Plant Cell Environ. 2009, 32, 1525–1537. [30] Himken M., Lammel J., Neukirchen D., Czypionka-Krause U. and Olfs H.-W.: Cultivation of Miscanthus under West European conditions: Seasonal changes in dry matter production, nutrient uptake and remobilization. Plant Soil 1997, 189, 117–126. Miscanthus – Unusual Grass: Biochemical and Physiological Characteristic: A Review 1623

[31] Donalatas N.G., Archontoulis S.V. and Mitsios I.: Potential growth and biomass productivity of Miscanthus × giganteus as affected by plant density and N-fertilization in central Greece. Biomass Bioenergy. 2007, 31, 145–152. [32] Christian D.G., Riche A.B. and Yates N.E.: Growth, yield and mineral content of Miscanthus × giganteus grown as biofuel for 14 successive harvests. Ind. Crop. Prod. 2008, 28, 320–327. [33] Amougou N., Bertrand I., Machet J.-M. and Recous S.: Quality and decomposition in soil of rhizome, root and senescent leaf from Miscanthus × giganteus, as affected by harvest date and N fertilization. Plant Soil 2011, 338, 83–97. [34] Kotecki A.: Cultivation of Miscanthus × giganteus. Wyd. Uniwersytetu Przyrodniczego we Wroc³awiu, 2010 [in Polish with English abstracts]. [35] Kalembasa D., Malinowska E., Jaremko D. and Je¿owski S.: The influence of NPK fertilization on yield structure of the Miscanthus ssp. grasses. Biul. IHAR 2004, 234, 205–211 [in Polish with English summary]. [36] Cosentino S.L., Patanè C., Sanzone E., Copani V. and Foti S.: Effects of soil content and nitrogen supply on the productivity of Miscanthus × giganteus Greef et Deu. in a Mediterranean environment. Ind. Crop. Prod. 2007, 25, 75–88. [37] Ercoli L., Mariotti M., Masoni A. and Bonari E.: Effect of irrigations and nitrogen fertilization on biomass yield and efficiency of energy use in crop production of Miscanthus. Field Crops Res. 1999, 63, 3–11. [38] Christian D.G., Poulton P.R., Riche A.B., Yates N.E. and Todd A.D.: The recovery over several seasons of 15N-labelled fertilizer applied to Miscanthus × giganteus ranging from 1 to 3 years old. Biomass Bioenergy 2006, 30, 125–133. [39] Kalembasa D., Je¿owski S., Pude R. and Malinowska E.: The content of carbon, hydrogen and nitrogen in different development stage of some clones of Miscanthus. Polish J. Soil Sci. 2005, 38, 169–177. [40] Lewandowski I. and Heinz A.: Delayed harvest of miscanthus – influences on biomass quantity and quality and environmental impacts of energy production. Eur. J. Agron. 2003, 19, 45–63. [41] Sànchez-Moreiras A.M., Weiss O.A. and Reigosa-Roger M.J.: Allelopathic evidence in the Poaceae. Bot. Rev. 2004, 69, 300–319. [42] Chou Ch.-H. and Lee Y.-F.: Allelopathic dominance of Miscanthus transmorrisonensis in an alpine grassland community in Taiwan. J. Chem. Ecology 1991, 17, 2267–2281. [43] Watanabe T., Jansen S. and Osaki M.: Al-Fe interactions and growth enhancement in Melastoma malabathricum and Miscanthus sinensis dominating acid sulphate soils. Plant Cell Environ. 2006, 29, 2124–2132. [44] Ezaki B., Nagao E., Yamamoto Y., Nakashima S. and Enomoto T.: Wild plants, Andropogon viriginicus L. and Miscanthus sinensis Anders, are tolerant to multiple stresses including aluminium, heavy metals and oxidative stresses. Plant Cell Rep. 2008, 27, 951–961. [45] Pilon-Smits E.A.H., Quinn C.F., Tapken W., Malagoli M. and Schiavon M..: Physiological functions of beneficial elements. Curr. Opin. Plant Biol. 2009, 12, 267–274. [46] Arduini I., Ercoli L., Mariotti M. and Masoni A.: Response of miscanthus to toxic cadmium applications during period of maximum growth. Environ. Exp. Bot. 2006, 55, 29–40. [47] Rowe R.L., Street N.R. and Taylor G.: Identifying potential environmental impacts of large-scale deployment of dedicated bioenergy crops in UK. Renew. Sustain. Energ. Rev. 2009, 13, 271–290. [48] Smeets E.M.W., Lewandowski I.M. and Faaij A.P.C.: The economical and environmental performance of miscanthus and switchgrass production and supply chains in a European setting. Renew. Sustain. Energ. Rev. 2009, 13, 1230–1245.

MISCANTHUS – TRAWA NIEZWYK£A: CHARAKTERYSTYKA BIOCHEMICZNO-FIZJOLOGICZNA: PRZEGL¥D LITERATUROWY

Katedra ¯ywienia Roœlin, Wydzia³ Przyrodniczo-Technologiczny Uniwersytet Przyrodniczy we Wroc³awiu

Abstrakt: Miscanthus × giganteus (Miskant olbrzymi) jest jedn¹ z bardziej obiecuj¹cych tzw. roœlin alternatywnych uprawianych z przeznaczeniem na cele energetyczne. Pomimo ¿e pochodzi z po³udniowo- 1624 El¿bieta Saca³a

-wschodniej Azji i jest roœlin¹ ciep³olubn¹, to bardzo dobrze roœnie i charakteryzuje siê wysok¹ produk- tywnoœci¹ w strefie umiarkowanych szerokoœci geograficznych. Miscanthus × giganteus jest roœlin¹ typu C4, dlatego w pracy przedstawiono krótki opis procesu fotosyntetycznego wi¹zania CO2 w tzw. szlaku C4 oraz uczestnicz¹cych w nim enzymów. Korzystaj¹c z najnowszych danych literaturowych, przeanalizowano podstawy biochemiczne stosunkowo du¿ej odpornoœci tej roœliny na niskie temperatury (0–15 oC). Opisano równie¿ szereg ekofizjologicznych w³aœciwoœci Miscanthus × giganteus (wysoka produktywnoœæ, niewielkie wymagania nawozowe, brak koniecznoœci stosowania pestycydów, mo¿liwoœæ wykorzystania w procesach fitoremediacji), które sprawiaj¹, ¿e jest on okreœlany jako roœlina ekologiczna i szczególnie przyjazna œrodowisku. Opisane cechy roœliny sprawiaj¹, ¿e mo¿na j¹ uznaæ za lidera wœród roœlin uprawianych na cele nie¿ywnoœciowe.

S³owa kluczowe: miskantus, tolerancja na ch³ód, C4 fotosynteza, fitoremediacja, allelopatia ECOLOGICAL CHEMISTRY AND ENGINEERING A

Vol. 18, No. 12 2011

Anna GORCZYCA1 and Marek J. KASPROWICZ2

INITIAL RESEARCH ON THE EFFECT OF THE NANOGRO PLANT GROWTH STIMULATOR ON Fusarium culmorum (W.G. Smith) Sacc.

WSTÊPNE BADANIA NAD WP£YWEM STYMULATORA WZROSTU ROŒLIN NANOGRO NA Fusarium culmorum (W.G. Smith) Sacc.

Abstract: An in vitro experiment determined the effect of the Nanogro plant growth and development stimulator, newly introduced in Poland, on Fusarium culmorum (W.G. Smith) Sacc. phytopathogenic fungus. A modification of linear growth and sporulation of F. culmorum mycelium caused by Nanogro was observed after the contact with the fungus spores, vegetative mycelium and the added medium. In the experiments where limited linear growth of Nanogro was observed as a result, stimulation of mycelium sporulation occurred the most frequently. The application of Nanogro in practice will not contribute to limiting the harmfulness of this pathogen. Keywords: F. culmorum, Nanogro, linear growth, sporulation

Research on agro-homeopathy has been conducted worldwide and preparations with ultra low content of elements or chemical compounds positively affecting growth and development of plants have been introduced to the agro-market. Scientific research on agro-homeopathy revealed a positive effect of ultra-low concentrations of the substances selected for the experiments on the growth of various plants, particularly during germination [1–7]. Like medical homeopathy, also agro-homeopathy has both supporters and opponents [8]. Nanogro, a plant growth and development stimulator, was first marketed in Poland in 2007 as one of the few agro-homeopathic products. The preparation consists of oligosaccharide granules (pellets) saturated with metal sulphates (Fe, Co, Al, Mg, Mn, Ni and Ag) in nanomolar concentrations. Nanogro is recommended for treatment and

1 Department of Agricultural Environment Protection, University of Agriculture in Krakow, al. A. Mic- kiewicza 21, 31–120 Kraków, Poland, phone: +48 12 662 44 00, email: [email protected] 2 Department of Physics, University of Agriculture in Krakow, al. A. Mickiewicza 21, 31–120 Kraków, Poland, phone: +48 12 662 43 91, email: [email protected] 1626 Anna Gorczyca and Marek J. Kasprowicz watering agronomic and horticultural plants, which according to its producer causes an increase in yields, more exuberant plant development, shortens vegetation and improves plant viability and vigour. Fungi of the Fusarium genus are dangerous cosmopolitan pathogens of crops. Diseases caused by these fungi lead not only to a decline in yield and its parameters but also worsens its quality due to the presence of micotoxins dangerous for humans and animals. The species dominant in Europe is Fusarium culmorum (W.G. Smith) Sacc. [9–12]. The research was conducted to assess the effect of Nanogro on the vegetative mycelium and spores of Fusarium culmorum.

Material and methods

F. culmorum strain purchased from the collection of the Plant Protection Institute (the Plant Pathogen Bank), isolated from cabbage was used for the analyses. The experiment was conducted in vitro to test the effect of Nanogro on the concentrated spore suspension and vegetative mycelium of F. culmorum. Nanogro in the concentrations recommended by the producer (10 granules per 10 dm3 of water) and three times increased (30 granules per 10 dm3 of water) was added to the fungus spore suspension prepared in sterile distilled water. The control was the suspension with the same spore concentration without the Nanogro supplement. The spore suspension was shaken in 300 cm3 flasks for 81 hours and after 1, 2, 3, 6, 9 and 81 hours some amount of suspension, adequate for further culturing was collected. F. culmorum spore suspension was inoculated on Petri dishes with solid PDA medium in 7 replications. The culture was maintained at the temperature of 21 oC. Linear growth of the cultured mycelium was measured. Once the culturing was completed, spore suspensions were prepared of the mycelia discs with a 50 mm diameter and shaken in 100 cm3 of distilled water. After filtration the spores number was measured in the obtained suspensions using the spectrophotometric method. The subsequent in vitro experiment aimed at an assessment of the effect of Nanogro on F. culmorum vegetative mycelium. Mycelial discs witha5mmdiameter from the Department’s own collection were inoculated on Petri dishes with solid PDA medium. Nanogro solutions were prepared in sterile distilled water (10, 20 and 30 granules per 10 dm3 of water). Subsequently, 30 mm3 (ml) of the solution was dripped with a micropipette onto the discs inoculated on the Petri dishes. 7 replications were made for each Nanogro concentration, whereas the fungus culture from discs dripped with sterile distilled water constituted the control. The culture was maintained and mycelial linear growth was measured until sporulation was obtained, which was assessed as presented above. The effect of Nanogro supplied to PDA medium in the concentration of 10 granules per 10 dm3 was also tested. As in the other experiments, assessed were mycelia linear growth and its sporulation after culturing. The control was the culture maintained on the standard PDA medium. The obtained results were verified statistically. Initial Research on the Effect of the Nanogro Plant Growth Stimulator... 1627

Results and discussion

Figures 1 and 2 present the results obtained in the experiment assessing the effect of Nanogro on F. culmorum spores. After 1, 2, 3, 6 and 9-hour spore contact with the preparation in the concentration recommended by the producer (10 granules per 10 dm3)

90 90 80 80 control control 70 N1 70 N1 N2 N2 60 60

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10 T=9h 10 T=81h 0 0 0 10 203040 50 60 70 80 90 100 110 120 130 0 10 203040 50 60 70 80 90 100 110 120 130 Time of growth [h] Time of growth [h]

Fig. 1. The effect of Nanogro on F. culmorum spores expressed by linear growth in consecutive culturing after contact (T) with preparation in the concentrations of 10 granules per 10 dm3 (N1) and 30 granules per 10 dm3 (N2) 1628 Anna Gorczyca and Marek J. Kasprowicz

0.20

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Fig. 2. The effect of Nanogro on F. culmorum spores expressed by mycelial sporulation obtained in consecutive culturing after contact with the preparation (1, 2, 3, 6, 9 and 81 h) in concentration of 10 granules per 10 dm3 (N1) and 30 granules per 10 dm3 (N2), c – control

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Fig. 3. The effect of Nanogro on F. culmorum vegetative mycelium growth, applied Nanogro concentrations: 10 granules per 10 dm3 (N1), 20 granules per 10 dm3 (N2) and 30 granules per 10 dm3 (N3) Initial Research on the Effect of the Nanogro Plant Growth Stimulator... 1629

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Fig. 4. The effect of Nanogro applied on vegetative mycelium on sporulation obtained in consecutive culturing. Applied Nanogro concentrations: 10 granules per 10 dm3 (N1), 20 granules per 10 dm3 (N2) and 30 granules per 10 dm3 (N3)

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control

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Fig. 5. The effect of Nanogro supplied to the medium in concentration of 10 granules per 10 dm3 on F. culmorum linear growth 1630 Anna Gorczyca and Marek J. Kasprowicz

0.18

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Fig. 6. The effect of Nanogro supplied to the medium in the concentration of 10 granules per 10 dm3 on F. culmorum sporulation a slight reduction of linear growth of mycelium cultured from spores was revealed in comparison with the control (Fig. 1), which was not registered for the higher concentration (30 granules per 10 dm3). The longest (81 h) spore contact with the preparation in both applied concentrations significantly limited the fungus growth in the consequent culturing. The analyzed mycelia sporulation after culturing (Fig. 2) evidences that despite limiting mycelia linear growth, Nanogro affects the strong stimulation of F. culmorum sporulation. The results obtained from the experiment on the effect of Nanogro on F. culmorum vegetative mycelium show that only in the concentration of 20 granules per 10 dm3 of water the preparation significantly limits the linear growth of the pathogen mycelium in comparison with the control (Fig. 3). Like in the experiments on spores, Nanogro in the concentration limiting the linear growth caused a markedly greater mycelium sporula- tion (Fig. 4). On the other hand, Nanogro supplied to the medium slightly stimulated linear growth of F. culmorum (Fig. 5) in comparison with the control and significantly limited mycelium sporulation obtained in culturing on this medium (Fig. 6). The obtained results demonstrated that Nanogro modifies linear growth and sporulation of F. culmorum – a dangerous crop pathogen. However, no fungistatic effect of Nanogro was registered, whereas practical application of this preparation will contribute to limiting this pathogen harmfulness.

References

[1] Betti L., Brizzi M., Nani D. and Peruzzi M.: Brit. Homeopath. J. 1994, 83, 195–201. [2] Bornoroni C.: Berlin J. Res. Homeopath. 1991, 1, 275–278. Initial Research on the Effect of the Nanogro Plant Growth Stimulator... 1631

[3] Hamman B., Koning G. and Lok Him K.: Homeopathy 2003, 92, 140–144. [4] Jones R.L. and Jenkins M.D.: Brit. Homeopath. J. 1981, 70, 120–128. [5] Jones R.L. and Jenkins M.D.: Brit. Homeopath. J. 1983, 72, 143–147. [6] Pelikan W. and Unger G.: Brit. Homeopath. J. 1971, 60, 232–266. [7] Pongratz W., Endler P.C.: Ultra High Dilution, Physiology and Physics, Kluwer Academic Publishers, Dordrecht 1994, 19–26. [8] Scofield A.M.: Biol. Agric. Hortic. 1984, 2, 1–50. [9] Gang G., Miedaner T., Schuhmacher U., Schollenberger M. and Geiger H.H.: Phytopathology 1998, 88, 879–884. [10] Hestbjerg H., Felding G. and Elmholt S.: J. Phytopathol. 2002, 150, 308–316. [11] Parry D.W., Jenkinson P. and McLeod L.: Plant Pathol. 1995, 44, 207–238. [12] Wagacha J.M. and Muthomi J.W.: Crop Protection 2007, 26, 877–885.

WSTÊPNE BADANIA NAD WP£YWEM STYMULATORA WZROSTU ROŒLIN NANOGRO NA Fusarium culmorum (W.G. Smith) Sacc.

1 Katedra Ochrony Œrodowiska Rolniczego 2 Katedra Chemii i Fizyki Uniwersytet Rolniczy im. Hugona Ko³³¹taja w Krakowie

Abstract: W doœwiadczeniu in vitro okreœlono wp³yw nowego w Polsce stymulatora wzrostu i rozwoju roœlin Nanogro na grzyb fitopatogenny Fusarium culmorum (W.G. Smith) Sacc. Stwierdzono modyfikacjê wzrostu liniowego i zarodnikowania grzybni F. culmorum wywo³ywane przez Nanogro po kontakcie z zarodnikami grzyba, grzybni¹ wegetatywn¹, jak i dodanego do pod³o¿a hodowlanego. W doœwiadczeniach, gdzie pod wp³ywem Nanogro obserwowano ograniczenie wzrostu liniowego, wystêpowa³a najczêœciej stymulacja sporulacji grzybni. Stosowanie Nanogro w praktyce nie przyczyni siê do ograniczenia szkodliwoœci tego patogenu. S³owa kluczowe: F. culmorum, Nanogro, wzrost liniowy, zarodnikowanie

ECOLOGICAL CHEMISTRY AND ENGINEERING A

Vol. 18, No. 12 2011

Marcin SIDORUK1, Andrzej ROCHWERGER1, El¿bieta SKORBI£OWICZ2 and Miros³aw SKORBI£OWICZ2

EFFECT OF CATCHMENT AREA USE ON LEAD AND ZINC ACCUMULATION IN THE BOTTOM DEPOSITS OF LAKES ARDUNG AND BUKWALD

WP£YW U¯YTKOWANIA ZLEWNI NA AKUMULACJÊ O£OWIU I CYNKU W OSADACH DENNYCH NA PRZYK£ADZIE JEZIOR ARDUNG I BUKWA£D

Abstract: The study concerns with the effect of catchment area use on lead and zinc accumulation in the bottom deposits of lakes. It was carried out in two water bodies in the Olsztyn Lakeland. The catchment areas of the investigated lakes are used for various purposes, ranging from forests to agricultural production. Lake Ardung (N 53o45¢,E20o55¢) is situated in the eastern part of the Masurian Lakeland, approximately 25 km east of Olsztyn. The lake has an area of 26.2 ha and a maximum depth of 3.6 m. The lake’s catchment area of 1539 ha is covered by farmland in 2 %, grassland in 2 %, fallow land overgrown with shrubs in 11.4 % and forests in 84.6 %. Lake Bukwald (N 53o58¢,E20o16¢) is located in the vicinity of the village of Bukwald, municipality of Dywity, around 20 km north of Olsztyn. The lake’s catchment area of 1156.8 ha comprises arable land in 60 %, forests and afforested areas in 31 % and wasteland in the remaining part. The studied water bodies were characterized by low concentrations of the analyzed elements. The average lead and zinc levels reached 28.3 mg/kg d.m. and 32.3 mg/kg d.m., respectively in Lake Ardung, and 33.3 mg/kg d.m. and 91.9 mg/kg d.m., respectively in Lake Bukwald. The total zinc and lead accumulation in the bottom deposits of the investigated water bodies, in terms of the surface area of the lakes and their catchments, was significantly higher in Lake Bukwald than in Lake Ardung. Keywords: lakes, bottom deposits, catchment area, trace elements, lead, zinc

Substance runoff from the catchment area into surface waters is determined by various factors, in particular land relief, soil cohesion and fertility, the type of land use, water relations and climate conditions, mostly the volume and distribution of precipita-

1 Department of Land Reclamation and Environmental Management, University of Warmia and Mazury in Olsztyn, pl. £ódzki 2, 10–756 Olsztyn, Poland, phone: +48 89 523 43 51, email: [email protected] 2 Department of Technology in Environmental Engineering and Protection, ul. Wiejska 45E, 15–351 Bia³ystok, Poland, email: [email protected] 1634 Marcin Sidoruk et al tion [1]. Vast quantities of substances are also supplied by atmospheric precipitation which contributes to the leaching of chemical components from the soil [2, 3]. Bottom deposits are a combination of crystalline and amorphous minerals with a different grain size, a various content of organic matter and mineral or organic colloidal substances [4, 5]. Bottom deposits are formed by the sedimentation of allochtonic material created outside the sedimentation area as well as autochthonic material formed in the place of sedimentation [6]. The bottom deposits of aquatic ecosystems vary immensely from coarse-grained, nearly mineral deposits to fine- -grained, mostly organic deposits in the deep strata of lakes. Deposits comprise mineral substances (silica, silicate, aluminosilicate, carbonate) as well as organic substances of various origin (catchment area, littoral and pelagial zones) and various degree of decomposition. Deposits found at deeper strata are generally characterized by greater thickness, fine-grained structure and a higher organic matter content [7]. The trace element content of bottom deposits is conditioned by numerous natural and anthropogenic factors. It is largely determined by the geological structure of the catchment area, its geomorphologic characteristics and climate conditions which are responsible for rock weathering, the mobilization, migration and accumulation of elements in the environment. In undeveloped areas, high concentrations of potentially harmful trace elements in bottom deposits are attributed mainly to different types of human activity in the catchment area, mostly agricultural production [8–10].

Materials and methods

In the study the effect of catchment area use on lead and zinc accumulation in the bottom deposits of lakes was carried out in two water bodies in the Olsztyn Lakeland. The catchment areas of the investigated lakes are used for various purposes, ranging from forests to agricultural production. Lake Ardung (N 53o45¢,E20o55¢) is situated in the eastern part of the Masurian Lakeland, approximately 25 km east of Olsztyn. The lake has an area of 26.2 ha and a maximum depth of 3.6 m. The lake’s catchment area of 1539 ha is covered by farmland in 2 %, grassland in 2 %, fallow land overgrown with shrubs in 11.4 % and forests in 84.6 %. Lake Ardung is located in the catchment area of the Lyna River in the watershed of Lyna (tributary of the Prego³a River) and Omulew (tributary of the Narew River in the Vistula Basin) river systems. This area is characterized by a large number of small lakes, ponds and swamps. It is weakly populated, and the predominant types of human activity include farming, forestry and tourism. The parent rock of catchment area soils comprises sandy glaciofluvial deposits that fill the channel surrounded by more cohesive clay formations [11]. The catchment area features mostly podzolic soils in natural pine forests. Depression areas comprise mostly hydrogenic soils developed in the course of several drainage schemes. The lake’s present catchment area was formed during land improvement projects carried out in the mid 19th and the early 20th century. As part of those schemes, the lake’s water horizon was lowered to expose shallow waters. Today, Lake Ardung is a remnant of a multi-sectional lake with the original area of around 250 ha surrounding the town of Nerwik. The lake was periodically separated Effect of Catchment Area Use on Lead and Zinc Accumulation... 1635 to create 224 ha of farmland. The original lake basin which today forms Lake Ardung spanned an area of 62 ha. Lake Bukwald (N 53o58¢,E20o16¢) is located in the vicinity of the village of Bukwald, municipality of Dywity, around 20 km north of Olsztyn. Lake Bukwald is a flow through water body fed by four streams in its north-western and the south-western parts. Water is evacuated from the lake via a single watercourse in the village of Bukwald. The lake has an area of 36.2 ha and a maximum depth of 12.4 m. It is supplied from a hilly area marked by significant altitude variations – the difference between the highest and the lowest point in the catchment area is 23.5 m. Lake Bukwald’s catchment area of 1156.8 ha comprises arable land in 60 %, forests and afforested areas in 31 % and wasteland in the remaining part. There is a predominance of light to medium-heavy loams in the north, and light and heavy loamy sands turning into light loams and slightly loamy sands in the south. Catchment area soils fall into quality classes IVa and IVb and, locally, IIIa and IIIb. Surface samples from the bottom deposits of the studied lakes were collected for physical and chemical analyses in June 2008. Sampling sites were distributed along five perpendicular transects, three sites per transect (Fig. 1). The location of sampling sites was chosen based on barimetric charts to produce the most comprehensive overview of the morphological and chemical properties of deposits, subject to the lake-bottom topography, the shape of the lake basin, flow rate, tributaries, etc.

Fig. 1. Location of deposit sampling sites in Lake Ardung and Lake Bukwald

Deposit samples were collected using the Ekman grab (for collecting samples with a surface area larger than 250 cm2). Three samples were collected from each site, and they were averaged into one sample. The deposits and sedimentation water sampled from each site were placed in air-tight glass containers. The physical properties of samples were analyzed by the Troels-Smith method immediately after collection [12]. At the laboratory of the Department of Land Improvement and Environmental Management at the University of Warmia and Mazury in Olsztyn, the following physical and chemical parameters were determined in fresh deposit samples directly after collection: – solid residue – by the gravimetric method at 105 oC [g/kg d.m.], 1636 Marcin Sidoruk et al

– ignition losses – as an indicator of organic substance content, calculated as the difference between solid residue and residue after ignition [g/kg d.m.], Pb and Zn levels were determined by atomic absorption spectroscopy (AAS) in deposit samples at the laboratory of the Department of Technology in Environmental Engineering and Protection at the Bialystok University of Technology.

Results and discussion

The bottom deposits of the analyzed water bodies were characterized by significant morphological diversity. Deposits from Lake Ardung (Nig. 4) were marked by a much greater degree of darkness than samples from Lake Bukwald (Nig. 2). Structural layering was not determined in samples from both lakes (Strf. 0), and the analyzed deposits were completely homogenous. Owing to continuous contact with water, the surface layers of bottom deposits were strongly hydrated (Sicc. at level 2–3), and they contained highly decomposed plant matter (Humo at level 3–4, H7–H10 on the von Post scale of humification). The investigated deposits had homogenous, greasy structure. Littoral zone samples contained fragments of hydrophyte roots with partially decompos- ed plant matter and small quantities of mollusk shells, while profundal zone samples were more homogenous (with an absence of plant remains or macrophyte roots). The analyzed deposits had the characteristic features of sapropel formations [12]. Significant differences were observed in the water content of deposit samples. The average dry matter content was 16.3 % in samples from Lake Ardung and 20.9 % in deposits from Lake Bukwald (Table 1). More profound differences in the dry matter content of deposits were found between sampling sites in Lake Ardung. The highest moisture content of 88.7 % was observed in samples from the central part of transect A3, and the lowest moisture content of 65.4 % was reported in transect A1, ie on the side of the inflow to the lake near the river head-streams. In Lake Bukwald, the highest water content of bottom deposits was determined in samples from the central part of transect B2 (86.8 %), and the lowest – on the northern side of transect B4 (66.3 %).

Table 1 Physical and chemical properties of bottom deposits in lakes

Lake Ardung Lake Bukwald

Parameter Unit mean mean (min – max) (min – max) 16.3 20.9 Dry matter [%] (11.3–34.6) (13.2–33.7) 18.9 14.0 Organic content [% d.m.] (14.9–26.0) (8.1–19.9) 28.3 33.3 Pb [mg/kg d.m.] (20.0–35.0) (24.0–56.0) 32.3 91.1 Zn [mg/kg d.m.] (19.7–68.8) (45.9–112.9) Effect of Catchment Area Use on Lead and Zinc Accumulation... 1637

The organic content of deposits is formed by remnants of aquatic organisms that fall to the bottom of the lake, organic suspension, the precipitation and coagulation of organic substances dissolved in water. The organic matter content of deposit surface layers is determined mostly by the productive output of the trophogenic zone, the quantity of allochtonic matter and sedimentation time [13–17]. The average organic matter content of sediment samples from the analyzed lakes was 14.0 % in Lake Bukwald and 18.9 % in Lake Ardung. The reported values are characteristic of post-glacial lakes whose organic matter content ranges from 10 % to 70 % [16, 18]. The results of the study point to variations not only between the water bodies, but also in the spatial distribution of organic matter in the examined lakes. In Lake Ardung, the lowest organic matter concentrations (14.9 %) were noted in deposits from the southern part of the water body on the side of the outflow in transect A5 and in deposits from the side of the inflow to the lake (15.7 %), while the highest organic content at 26 % was reported in the central part of transect A2 (deepest waters). Similar results were noted in Lake Bukwald where the lowest organic matter concentrations were found in samples collected on the side of the outflow (8.1 %) and near the inflow (13.8 %), and the highest organic matter content was determined in deposits from the profundal zone of the lake (19.9 %). Organic matter produced by the precipitation and coagulation of organic substances dissolved in water has a capillary structure, and its specific gravity is similar to the specific gravity of water [12, 14, 15]. The above facilitates the lifting and the transport of substance particles, and it explains the limited organic matter content of samples collected on the side of the outflows and inflows because the deposits accumulating in those areas are depleted by moving water which washes out lighter particles. Runoff and feeder sections of the lake are shallower and more abundant in oxygen which contributes to the mineralization of deposits. The highest organic matter content was noted in the deepest zones of the lake owing to the gravitational force which pushes deposits deeper down the lake basin. The studied deposits were characterized by an average lead content of 28.3 mg/kg d.m. in Lake Ardung and 33.3 mg/kg d.m. in Lake Bukwald. The geochemical background of lead in the bottom deposits of lakes in north-eastern Poland is 11 mg Pb/kg d.m., and the lead content of unpolluted deposits should not exceed 20 mg/kg d.m. [8]. In Lake Bukwald, the highest Pb levels were noted on the southern side of transect B4 (56.0 mg Pb/kg d.m.), while the lowest lead concentrations were found in deposits on the southern side of transect B1 (24.0 mg Pb/kg d.m.). In the mid-forest Lake Ardung, Pb levels were lower, in the range of 20.0 mg/kg d.m. to 35.0 mg/kg d.m. (Table 1). The average zinc content of the analyzed deposits was low, ranging from 32.3 mg/kg d.m. in Lake Ardung to 91.1 mg/kg d.m. in Lake Bukwald. The geochemical background of zinc in soils usually does not exceed 50.0 mg/kg dm, while the average values noted in lakes are generally under 100 mg/kg d.m. [19]. The average Zn content of the investigated deposits approximated geochemical background levels. Zinc concentrations in the bottom deposits from Lake Ardung were within the 19.7–68.8 mg/kg d.m. range. The Zn content of samples from Lake Bukwald, whose catchment area is occupied by a housing estate and farmland, was determined at 45.9–112.9 mg/kg 1638 Marcin Sidoruk et al d.m. (Table 1). According to the system for classifying the purity of bottom deposits proposed by Bojakowska and Sokolowska [20], the Zn and Pb concentrations noted in this study meet first class purity standards. Agricultural production in the catchment area of Lake Bukwald, including the use of pesticides, contributes to high Pb values in deposit samples from the lake. Lead concentrations were further increased by the long-term use of tetraethyl lead as an antiknock agent in the fuel powering farming machines as well as atmospheric deposition. According to Nicholson [21], atmospheric deposition is the main source of lead in agricultural areas, and it accounts for as much as 77 % of its total supply. A comparison of lead and zinc concentrations in deposit samples shows that they were significantly lower in the littoral zones than in the profundal zones of the analyzed lakes. The low Pb and Zn content of bottom deposits in littoral zones could be attributed to bioaccumulation. Deposits in deeper parts of the lake are characterized by high concentrations of organic substances and fine-grained mineral fractions that bind metals [22]. The results of the analysis indicate that the highest concentrations of trace elements in Lake Ardung were found on the side of the outflow and in the profundal zone of the lake, while the lowest Pb and Zn levels were observed on the side of the inflow near the river head-streams. In Lake Bukwald, lower lead and zinc levels were also noted in deposit samples collected on the side of the inflow, while the highest Pb and Zn concentrations were reported in transects on the side of the outflow. An analysis of total lead and zinc load in the bottom deposits of the examined water bodies points to significantly higher accumulation levels in Lake Bukwald at 1372.6 kg Zn and 502.3 kg Pb. In Lake Ardung, zinc accumulation was 80 % lower (279.5 kg Zn) and Pb accumulation was approximately 50 % lower (247.2 kg Pb) than in Lake Bukwald (Table 2).

Table 2

Accumulation of trace elements in the 0–20 cm layer of bottom deposits in the studied lakes

Lake Indicator Ardung Bukwald Pb Zn Pb Zn kg/lake 247.2 279.5 502.3 1 372.6 kg /ha lake 9.43 10.7 13.88 37.9 kg/ha catchment area 0.14 0.16 0.43 1.2

An analysis of Zn and Pb accumulation in terms of the catchment area unit points to higher deposition in Lake Bukwald at 1.19 kg Zn/ha and 0.43 kg Pg/ha. In the mid-forest Lake Ardung, accumulation values reached 0.16 kg Zn/ha and 0.14 kg Pb/ha. In terms of the surface area of the lakes, the bottom deposits of Lake Bukwald also accumulated more Zn and Pb (13.88 kg Pb/ha, 37.9 kg Zn/ha) than the deposits of Lake Ardung (9.43 kg Pb/ha, 10.7 kg Zn/ha) (Table 2). Effect of Catchment Area Use on Lead and Zinc Accumulation... 1639

A comparison of lead accumulation values in the bottom deposits of the examined lakes indicates that lead concentrations are largely determined by the type of catchment area use. A higher level of trace elements accumulation was noted in the bottom deposits of Lake Bukwald, whose catchment area is used for agricultural production, than in the deposits of the mid-forest Lake Ardung.

Conclusions

1. Variations in the spatial distribution of lead and zinc were determined in the bottom deposits of the investigated lakes. Higher levels of the analyzed trace elements were noted in deposit samples collected from shallower lake sections on the side of the outflow. 2. The level of Zn and Pb accumulation in the bottom deposits of the examined water bodies was determined mostly by the type of catchment area use and the applied fertilization rates. In the bottom deposits of Lake Bukwald, whose catchment area is used for farming production and human settlement, the total accumulation of the analyzed elements was much higher (1372.6 kg Zn and 502.3 kg Pb) than in the bottom deposits of the mid-forest Lake Ardung (279.5 kg Zn and 247.2 kg Pb). 3. The total lead and zinc accumulation in bottom deposits in terms of the surface area of the lakes was significantly higher in Lake Bukwald (1372.6 kg Zn and 502.3 kg Pb) than in Lake Ardung (279.5 kg Zn and 247.2 kg Pb). The above differences resulted mainly from the type of activity performed in the catchment areas of the investigated water bodies.

References

[1] Sidoruk M. and Skwierawski A.: Effect of land use on the calcium, sodium, potassium and magnesium contents in water flowing into the Bukwa³d Lake. Ecol. Chem. Eng. 2006, 13, 337–343. [2] Mosiej J.: Przyrodniczo-techniczne uwarunkowania gospodarowania wod¹ w dolinie rzeki Ner. Wyd. SGGW. Rozprawy Naukowe i Monografie. 1999. [3] Wiœniewski R.J. and Nowicka B.: Ocena stanu i przyrodnicze uwarunkowania ochrony wód powierzch- niowych. Materia³y do monografii przyrodniczej regionu gdañskiego. Gdañsk 2003, 53–65. [4] Pasternak K.: Bottom sediments of the polluted dam reservoir AT Otmuchów. Acta Hydrobiol. 1970, 12, 377–380. [5] Nocoñ W.: Zawartoœæ metali ciê¿kich w osadach dennych rzeki K³odnicy. J. Elementol. 2006, 11(4), 457–466. [6] Miranda L.E., Hargreaves J.A. and Raborn S.W.: Predicting and managing risk of unsuitable dissolved oxygen in a eutrophic lake. Hydrobiologia 2001, 457, 177–185. [7] Kajak Z.: Hydrologia – Limnologia. Ekosystemy wód œródl¹dowych. Wyd. PWN, Warszawa 2001. [8] Bojakowska I. and Gliwicz T.: Wyniki geochemicznych badañ osadów wodnych Polski w latach 2000–2002. Biblioteka Monitoringu Œrodowiska, Warszawa 2003. [9] Cieszewski D., and Malik I.: Zapis XX-wiecznej historii zanieczyszczenia rzeki Ma³ej Panwi metalami ciê¿kimi w jej osadach. Przegl. Geol. 2003, 51(20), 142–147. [10] Skorbi³owicz E.: Ocena jakoœci œrodowiska wodnego wybranych rzek powiatu Siemiatycze. Woda – Œrodowiska – Obszary Wiejskie 2004, 4(11), 429–444. [11] Koc J., Nowicki Z., Gliñska K. and £achacz A.: Kszta³towanie siê jakoœci wód w warunkach malej antropopresji na przyk³adzie zlewni strugi Ardung (Pojezierze Olsztyñskie). Zesz. Nauk. Komitetu „Cz³owiek i Œrodowisko” 2000, 25, 155–166. [12] Tobolski K.: Przewodnik do oznaczania torfów i osadów jeziornych. PWN, Warszawa 2000. 1640 Marcin Sidoruk et al

[13] Mc Coll R.H.S.: Chemistry of sediments In relation to trophic condition of Wight Rotoura Lakes. J. Mar. Freshwater Res. 1977, 11, 371–380. [14] Januszkiewicz T. and Samulowska B.: Chemizm wspó³czesnych osadów dennych jeziora Wad¹g k. Olsztyna. Zesz. Nauk. ART Olsztyn 1978, 187, 31–58. [15] Rybak J.I.: Przegl¹d badañ nad osadami. Ekol. Pol. 1989, 15, 19–30. [16] Kentzer A.: Fosfor i jego biologiczne dostêpne frakcje w osadach jezior ró¿nej trofii. Wyd. Uniwersytetu M. Kopernika, Toruñ 2001, 5–9. [17] Kowalczewska-Madura K.: Materia organiczna w osadach dennych jeziora Swarzêdzkiego. Jeziora i sztuczne zbiorniki wodne. Uniwersytet Œl¹ski, Sosnowiec 2004, 125–131. [18] Siwek H., W³odarczyk M., Brzostowska-Œelechowska D. and Wachowiak M.: Wp³yw wybranych parametrów fizyczno-chemicznych osadu na zawartoœæ nieorganicznych form fosforu w osadach dennych ma³ych zbiorników polimiktycznych. Acta Agrophys. 2009, 13, 497–503. [19] Lis J. and Pasieczna A.: Atlas geochemiczny Polski 1: 2 500 000. Pañstw. Inst. Geol., Warszawa 1995. [20] Bojakowska I. and Soko³owska G.: Geochemiczne klasy czystoœci osadów wodnych. Przegl. Geolog. 1998b, 46, 49–54. [21] Nicholson F.A., Smith S.R., Alloway B.J., Carlton-Smith C. and Chambers B.J.: An inventory of heavy metals inputs to agricultural soils in England and Wales. Sci. Total Environ. 2003, 311, 205–219. [22] Szafran K.: Metale ciê¿kie w osadach dennych trzech p³ytkich jezior ³êczyñsko-w³odawskich. Acta Agrophys. 2003, 1, 329–337.

WP£YW U¯YTKOWANIA ZLEWNI NA AKUMULACJÊ O£OWIU I CYNKU W OSADACH DENNYCH NA PRZYK£ADZIE JEZIOR ARDUNG I BUKWA£D

1 Katedra Melioracji i Kszta³towania Œrodowiska, Uniwersytet Warmiñsko-Mazurski w Olsztynie 2 Katedra Technologii w In¿ynierii i Ochronie Œrodowiska, Politechnika Bia³ostocka

Abstrakt: Do badañ maj¹cych na celu okreœlenie wp³ywu u¿ytkowania zlewni jezior na akumulacjê o³owiu i cynku w ich osadach dennych wytypowano dwa zbiorniki po³o¿one na obszarze Pojezierza Olsztyñskiego. Zlewnie badanych jezior obejmuj¹ obszary o zró¿nicowanym zagospodarowaniu – od obszarów leœnych po u¿ytki rolne. Jezioro Ardung (N 53o45¢,E20o55¢) po³o¿one jest we wschodniej czêœci Pojezierza Mazurskiego ok. 25 km na wschód od Olsztyna. Powierzchnia jeziora wynosi 26,2 ha, natomiast jego maksymalna g³êbokoœæ 3,6 m. Na obszarze zlewni jeziora o powierzchni 1539 ha grunty orne stanowi¹ 2 %, ³¹ki i pastwiska 2 %, od³ogi w znacznym stopniu zakrzewione 11,4 % i 84,6 % lasy. Jezioro Bukwa³d (N 53o58¢,E20o16¢) po³o¿one jest w okolicach wsi Bukwa³d w gminie Dywity oko³o 20 km na pó³noc od Olsztyna. Ca³kowita zlewni jeziora Bukwa³d wynosi 1156,8 ha, z czego 60 % stanowi¹ grunty orne, 31 % to lasy i tereny zalesione, pozosta³¹ czêœæ stanow¹ nieu¿ytki. Badane osady charakteryzowa³y siê niskim stê¿eniem badanych pierwiastków i w jeziorze Ardung œrednie stê¿enie Pb wynosi³o 28,3 mg Pb/kg s.m., natomiast Zn – 32,3 mg Zn/kg s.m., zaœ w jeziorze Bukwa³d by³o to 33,3 mg Pb/kg s.m. oraz 91,1 mg Zn/kg s.m. Tak¿e ca³kowita akumulacja cynku i o³owiu w osadach badanych zbiorników zarówno w odniesieniu do powierzchni lustra wody, jak i zlewni by³a zdecydowanie wiêksza w osadach jeziora Bukwa³d ni¿ œródleœnego jeziora Ardung. S³owa kluczowe: jeziora, osady denne, zlewnia, pierwiastki œladowe, o³ów, cynk ECOLOGICAL CHEMISTRY AND ENGINEERING A

Vol. 18, No. 12 2011

Zdzis³aw CIEÆKO1, Miros³aw WYSZKOWSKI and El¿bieta ROLKA

ALUMINIUM CONCENTRATION IN PLANTS DEPENDING ON SOIL CONTAMINATION WITH CADMIUM

ZAWARTOŒÆ GLINU W ROŒLINACH W ZALE¯NOŒCI OD ZANIECZYSZCZENIA GLEBY KADMEM

Abstract: The aim of the study has been to determine the effect of soil contamination with cadmium (10, 20, 30 and 40 mg Cd × ha–1 soil) on the concentration of aluminium in aboveground parts and roots of oats, maize, yellow lupine and radish. In order to neutralise cadmium, the following neutralising agents were introduced to soil: compost, brown coal, lime and bentonite. Apart from the plant species and type of organs, other factors which largely affected the concentration of aluminium were a rate of cadmium and type of a neutralising substance. Roots contained much more aluminium than aboveground parts of plants. The highest levels of aluminium were found in roots of yellow lupine and maize whereas the smallest concentrations of this metal were determined in grain and roots of oats. Soil contamination with cadmium caused bigger changes in the concentration of aluminium in aboveground parts of plants than in their roots, especially in the case of maize and yellow lupine. Cadmium applied at 20 mg (maize and yellow lupine roots) or 40 mg Cd × ha–1 soil (aboveground parts of maize and yellow lupine) caused increased levels of cadmium in plant tissues. Any further increase in the rates of the pollutant caused depression in the content of aluminium in roots of the above crops. In the case of aboveground parts and roots of radish and grain of oats, less aluminium was observed in all cadmium contaminated objects. The range of effects produced by the test neutralising substances on the concentration of aluminium was varied. The neutralising agents tended to depress the content of aluminium in plant tissues. Brown coal, bentonite and lime caused larger changes in the content of aluminium than compost. The concentration of aluminium was correlated with yields of the crops. For most of the plants, these correlations were negative in the case of aboveground parts (except radish) and positive in roots (except yellow lupine). The concentration of aluminium in particular plant organs was correlated with a number of macro- and microelements, with the correlations being usually positive for manganese, iron, cobalt, lithium, copper and zinc but negative for sulphur and boron. Keywords: cadmium contamination, compost, brown coal, lime, bentonite, plants, aluminium content

Contamination of natural environment with heavy metals has many negative consequences to the nature. Among particularly harmful effects are modifications of the soil environment, especially changes in physicochemical, microbiological and bio-

1 Department of Environmental Chemistry, University of Warmia and Mazury, pl. £ódzki 4, 10–727 Olsztyn, Poland, phone: +48 89 523 35 66, fax: +48 89 523 39 76, email: [email protected] 1642 Zdzis³aw Cieæko et al chemical properties of soils [1–3]. Such alterations disrupt the uptake of many nutrients by plants, which in turn deteriorates quality of crops [4]. Crops which contain excessive concentrations of heavy metals cannot be used as animal fodder or human foodstuff. Cadmium is one of the most dangerous heavy metals – found in soil, it not only increases its levels in the plants but also changes concentrations of other elements in plant tissues. The latter effect is caused by antagonistic and syngeristic interactions [5, 6]. The response of plants to an identical level of soil contamination with cadmium can be highly varied – from very small changes, in some species, that are invisible to the naked eye, to failed emergence or necrosis of seedlings of other plant species [1]. The risk of cadmium and other heavy metals passing to successive food chain links must be minimised by undertaking protective measures, which reduce the influence of this metal on plants. Liming is the most popular treatment for this purpose, but other methods can also be helpful. Considering the above, a study has been conducted in order to assess the effect of soil contamination with cadmium on concentration of aluminium on aboveground parts and roots of oats, maize, yellow lupine and radish. Cadmium introduced to soil was neutralising with compost, brown coal, lime and bentonite.

Material and methods

A greenhouse experiment was conducted at the University of Warmia and Mazury in Olsztyn. Acidic soils of the granulometric composition of light loamy sand were used for the trials. A more detailed description of the soils can be found in our earlier publication [7]. Polyethylene pots were filled with 9 or 10 kg of soil. The soil was contaminated with cadmium at the following rates: 10, 20, 3 and 40 mg Cd × kg–1 soil. The effect produced by cadmium was tested on four plants: oats, maize, yellow lupine and radish. In order to reduce the influence of cadmium on plants, the following were applied in the trials with oats: compost, brown coal and lime. In the trials involving the other crops, bentonite was used in addition to the above neutralising agents. Compost and brown coal were introduced to soil at a rate of 4 % while bentonite was added at 2 % relative to the whole mass of soil in a treatment. Lime was added in an amount corresponding to 1 unit of hydrolytic acidity of soil. Simultaneously, the soils were fertilized with NPK, in the amounts adjusted to the crop species. The elements were introduced to soil in the following compounds and mixtures: cadmium as CdCl2, nitrogen as CO(NH2)2, phosphorus as Ca(H2PO4)2 +H3PO4 + CaSO4 and potassium as KCl. Cadmium, the neutralising agents and the mineral fertilizers were carefully mixed with the soil when the trials were being established. The experiment was run with 4 replications. During the growing season, the soil moisture level was maintained at 60 % capillary water capacity. The test plants were harvested in the stage of full maturity (oats and radish), the inflorescence stage (yellow lupine) and the cob formation stage (maize). During the harvest, samples of aboveground and underground parts of plants were collected. Plant roots were rinsed with water to remove soil. Samples of the plant material were fragmented, dried and ground. The concentration of aluminium in the Aluminium Concentration in Plants Depending on Soil Contamination with Cadmium 1643 plant material was determined by atomic absorption spectrophotometry (AAS) using a Unicam 939 Solar spectrophotometer. The statistical calculations were done using the software package Statistica [8].

Results

Apart from the plant species and plant organ, other factors which strongly affected the concentration of aluminium in plant tissues were the rate of cadmium and the type of a neutralising agent applied. The roots contained much more aluminium than the aboveground parts of plants. Most aluminium was determined in roots of yellow lupine and maize; the smallest levels of Al were found in grains and straw of oats. The soil pollution with cadmium caused more profound changes in the content of aluminium in aboveground rather than underground parts of plants, especially in maize and yellow lupine (Tables 1–2).

Table 1

Effect of cadmium on aluminium (Al) content in oats and maize [mg per kg of d.m.]

Oat Maize Cd contamination × –1 aboveground [mg kg of soil] grain straw roots roots parts 0 32.1 63.3 1385.4 59.9 2821.8 10 25.6 54.8 1474.8 94.5 3100.3 20 23.7 60.9 1330.6 90.4 3488.9 30 28.4 64.7 1506.3 65.6 3280.2 40 26.2 65.5 1569.2 239.7 2962.2 Average 27.2 61.8 1453.3 110.0 3130.7 r –0.440** 0.527** 0.662** 0.707** 0.278** r – correlation coefficient; * significant at p = 0.05; ** significant at p = 0.01.

Table 2

Effect of cadmium on aluminium (Al) content in yellow lupine and radish [mg per kg of d.m.]

Cd contamination Yellow lupine Radish × –1 [mg kg of soil] aboveground parts roots aboveground parts roots 0 205.1 4945.2 295.4 548.3 10 327.6 5404.4 284.6 509.5 20 647.8 5972.2 283.9 424.8 30 939.7 4995.4 273.6 370.4 40 1265.3 5130.7 244.5 418.2 Average 677.1 5289.6 276.4 454.2 r 0.991** –0.014 –0.917** –0.869** r – correlation coefficient; * significant at p = 0.05; ** significant at p = 0.01. 1644 Zdzis³aw Cieæko et al

The cadmium rates of 20 mg (maize and yellow lupine roots) and 40 mg Cd × ha–1 (aboveground parts of maize and yellow lupine) increased the levels of aluminium in plant tissues. Under these rates of the soil pollutant, the content of aluminium rose by 24 % (r = 0.278) in maize roots, 21 % in yellow lupine roots, 300 % (r = 0.707) in aboveground parts of maize and up to 517 % (r = 0.991) in aboveground parts of yellow lupine. Any further increase in the rates of cadmium introduced to soil depressed the concentration of aluminium in roots of these two crops. Regarding the aboveground parts and roots of radish as well the oats grain, all cadmium-polluted objects revealed depressed aluminium concentrations in plant tissues. Under the highest cadmium rate, the difference varied from 17 % (r = –0.917) in aboveground parts to 24 % (r = –0.869) in roots of radish. The smallest fluctuations in the content of cadmium were demonstrated in oats straw and roots. The range of effects produced by the neutralising agents on the concentration of aluminium was varied (Table 3).

Table 3

Effect of neutralization substances on aluminium (Al) content in plants [in mg per kg of d.m.]

Cd contamination Without Brown Compost Lime Bentonite Average [mg × kg–1 of soil] additions coal Oats Grain 31.1 24.5 26.1 27.1 — 27.2 Straw 69.3 61.3 59.5 57.3 — 61.8 Roots 1622.0 1466.0 1333.7 1391.2 — 1453.3 Maize Aboveground parts 55.9 38.4 57.8 228.4 169.6 110.0 Roots 3275.6 2870.5 2898.9 2921.2 3687.4 3130.7 Yellow lupine Aboveground parts 1226.4 802.8 394.4 485.3 476.7 677.1 Roots 5266.5 4638.2 4278.9 5279.0 6827.0 5289.6 Radish Aboveground parts 410.4 288.5 204.5 288.4 273.4 276.4 Roots 631.9 411.2 376.6 433.7 554.3 454.2

The neutralising agents tended to depress the concentration of aluminium in plants. Brown coal, bentonite and lime caused bigger changes in the content of aluminium in plants than did compost. Brown coal reduced the aluminium level by 68 % in yellow lupine aboveground parts, 50 % in radish aboveground parts, 40 % in radish roots, 19 % in yellow lupine roots, and in oats roots, grain and straw – by 18, 16 and 14 %, respectively. Bentonite depressed the concentration of aluminium by 61 % in above- ground parts of yellow lupine, 33 % in radish aboveground parts and 12 % in the roots of the former crop. The relationships determined for yellow lupine roots and, especially, for aboveground parts of maize were reverse. Also liming caused a contrary effect on aluminium in aboveground parts of maize. Lime and bentonite produced, respectively, a Aluminium Concentration in Plants Depending on Soil Contamination with Cadmium 1645

Aboveground parts Roots Oat

43 7.5 40 y = –0.398x + 69.978 R2 = 0.400 7.0 37 34 6.5

g per pot]

ld [ 31 e 6.0

Yi Yield [g per pot] y = 0.004x + 0.497 28 R=0.2162 25 5.5 79 82 85 88 91 94 97 1300 1400 1500 1600

[mg Al× kg–1 d.m.] [mg Al× kg–1 d.m.]

Maize 280 36 270 y = –0.236x + 283.997 R2 = 0.744 260 34 250

gperpot]

ld [ 240 32

e

Yi Yield [g per pot] y = 0.003x + 23.480 230 R2 = 0.070 220 30 50 100 150 200 250 2800 3000 3200 3400 3600

[mg Al× kg–1 d.m.] [mg Al× kg–1 d.m.]

Yellow lupine 120 10 100 80 9 60

g per pot]

ld [ 40 8 e y = –0.102x + 124.889

Yi Yield [g per pot] y = –0.002x + 17.491 2 20 R = 0.687 R=0.0522 0 7 0 300 600 900 1200 4800 5200 5600 6000

[mg Al× kg–1 d.m.] [mg Al× kg–1 d.m.]

Radish 90 90

80 80 70

g per pot] 60 ld [ 70 e y = 0.715x – 130.601 y = 0.087x – 37.300

Yi 2 Yield [g per pot] 50 R = 0.948 R=0.1172 40 60 240 250 260 270 280 290 300 350 400 450 500 550

[mg Al× kg–1 d.m.] [mg Al× kg–1 d.m.]

Fig. 1. Relationship between the content of aluminium (Al) and yield of plants 1646 Zdzis³aw Cieæko et al * ** ** ** ** * 0.36 0.55 Table 4 ** ** 0.53 0.27 0.55 0.14 0.05 –0.27 –0.74 ** ** * ** 0.12 –0.06 0.08 0.51 0.01 –0.05 –0.45 0.18 0.96 ** ** * ** 0.20 0.40 0.21 0.30 0.80 0.26 0.54 0.04 0.22 –0.04 0.33 –0.06 0.06 0.26 –0.21 ** ** ** ** ** ** 0.13 0.50 0.11 0.46 0.89 0.40 –0.47 –0.39 ** ** ** ** ** ** 0.57 0.300.96 0.53 0.85 0.99 0.47 –0.10 0.58 ** ** ** ** ** ** ** 0.47 0.61 0.70 0.89 0.93 0.94 0.44 0.18 0.74 * ** ** ** ** ** ** ** 0.34 0.65 0.84 0.000.82 0.10 0.13 –0.12 –0.29 0.97 ** ** ** ** ** ** 0.83 0.04 0.74 ** * Content in plants –0.23 0.64 –0.32 0.64 –0.00 0.91 –0.21–0.29 0.18 0.89 ** * ** * ** –0.24 0.20–0.62 0.60 –0.08 0.11–0.67 0.17 0.97 –0.31 ** ** ** ** ** –0.48 –0.61 –0.17 –0.57 * ** ** 0.62 –0.13 0.53 ** ** 0.01 0.44 0.090.50 0.08 –0.43 –0.22 –0.37 ** ** ** ** –0.27 0.13 0.22 –0.48 –0.50 –0.38 ** ** ** 0.09 –0.22 –0.14 0.14 –0.06 –0.30 –0.11 0.24–0.90 0.43 ** ** ** t p = 0.01. 0.22 –0.61 –0.84 ** ** N P K Mg Ca Na S B Mo Mn Fe Co Li Cd Pb Cu Ni Zn 0.21 –0.06 0.72 0.09 0.58 –0.17 0.06–0.13 –0.23 –0.36 –0.40 –0.95 –0.47 significant a ** ** ** ** ** ** ** Yield Relationships (r) between the aluminium (Al) content versus the yield and the content of macro- and microelements in plants t p = 0.05; in plants Al content significant a Yellow lupine aboveground parts –0.59 Oat grain –0.42 Oat rootsMaize above- ground partsMaize 0.46 roots –0.45 0.05Yellow lupine –0.14roots –0.25Radish above- –0.15ground parts –0.15Radish roots –0.25 0.36 0.12 0.03 –0.13 0.44 –0.18 –0.17 0.21 0.08 0.87 –0.58 Oat straw –0.28 –0.16 0.20 0.26 –0.07 –0.18 0.30 –0.24 –0.25 0.05 0.51 * Aluminium Concentration in Plants Depending on Soil Contamination with Cadmium 1647 four-fold and three-fold increase in aluminium found in aboveground parts of maize. Liming was also responsible for a 60 % decline in the aluminium concentration in yellow lupine aboveground parts, 30–31 % decrease in aboveground and underground parts of radish and 13–17 % drops in all parts of oats. The influence of compost on the concentration of aluminium in all the crops, and particularly in aboveground parts of yellow lupine and in aboveground and underground parts of radish, was significantly negative. The regression curves and equations derived from the data suggest that the concentration of aluminium in plant tissues was correlated with the plant yields. Most of the test plants showed negative correlations between these two factors for the aboveground parts (except radish) and positive correlations for the root tissues (except yellow lupine) (Fig. 1). As regards the aboveground parts of radish and roots of yellow lupine, contrary correlations were observed. The concentration of aluminium in particular plant organs was correlated with many macro- and micronutrients, with the correlations being stronger in roots than in aboveground parts. Besides, the correlations involving mircoelements were stronger than those with macroelements (Table 4). Significant correlations were the least frequent in the case of maize. The content of aluminium was most often positively correlated with the content of manganese, iron, cobalt, lithium, cadmium, copper and zinc. Most negative correlations were determined between aluminium and sulphur or boron.

Discussion

The literature has very few references on the above issues. Guo Tian Rong et al [9] reported higher levels of aluminium in barley caused by cadmium. By introducing various substances to soil, concentrations of aluminium in plants can be modified. Alfaro et al [10] demonstrated that liming can depress the content of aluminium in grasses and legumes with a wide range of differences (from 35 to 61 %). Mongia et al [11] proved analogous relationships in rice grain and straw. Wallace [12] studied such interactions in maize; Patriam-Rai et al [13] investigated beans, Hahn and Marschner [14] analysed aluminium content in spruce roots and, finally, Kiepul [15] studied a variety of plant species in this respect.

Conclusions

1. Concentration of aluminium in plants depends not only on the plant species, type of plant organs but, to a large extent, on cadmium soil pollution (rates of the pollutants) and type of a neutralising agent introduced to the soil. Roots of the test plants contained much more aluminium than their aboveground parts did. The highest levels of aluminium were determined in yellow lupine and maize roots whereas the smallest amounts of this metal were discovered in oats grain and straw. 2. Contamination of soil with cadmium caused bigger changes in the concentration of aluminium in aboveground parts than in roots of the test plants, especially in maize and yellow lupine. Cadmium applied at 20 mg (maize and yellow lupine roots) or 40 mg 1648 Zdzis³aw Cieæko et al

Cd × ha–1 (aboveground parts of maize and yellow lupine) caused increased con- centrations of aluminium. Any further increase in the cadmium rates resulted in depressed amounts of aluminium in roots of these two crops. Concerning the aboveground parts and roots of radish as well as oats grain, the content of aluminium was found to have been depressed as a consequence of cadmium pollution of the soil. 3. The range of effects produced by the test neutralising agents on the concentrations of aluminium was varied. The neutralising agents tended to depress the content of aluminium in plants. Brown coal, bentonite and lime caused larger changes in the levels of aluminium in plant tissues than compost. 4. The content of aluminium was correlated with yields of the crops. In most of the test plants, we noticed negative correlations for the aboveground parts (except radish) and positive ones for roots (except yellow lupine). The concentration of aluminium in particular plant organs was correlated with many micro- and macroelements. These correlations were predominantly positive in the case of manganese, iron, cobalt, lithium, copper and zinc, but negative for the concentrations of sulphur and boron.

References

[1] Kabata-Pendias A. and Pendias H.: Biogeochemia pierwiastków œladowych. PWN, Warszawa 1999. [2] Paivoke A.E.A. and Simola L.K.: Ecotoxicol. Environ. Safety 2001, 49, 111–121. [3] Wyszkowska J. and Wyszkowski M.: Polish J. Environ. Stud. 2003, 12(4), 479–485. [4] Wyszkowski M.: Polish J. Natur. Sci. 2002, 12(3), 21–35. [5] Cieæko Z., Wyszkowski M., Krajewski W. and Zabielska J.: Sci. Total Environ. 2001, 281(1–3), 37–45. [6] Gorlach E., Gambuœ F., Jamrozy G. and Tomek A.: Zesz. Probl. Post. Nauk Roln. 1996, 434, 89–98. [7] Cieæko Z., Kalembasa S., Wyszkowski M. and Rolka E.: Plant, Soil Environ. 2004, 50(7), 283–294 [8] StatSoft, Inc. 2006. STATISTICA (data analysis software system), version 7.1. www.statsoft.com. [9] Guo Tian Rong, Zhou Mei Xue, Wu Fei Bo and Chen Jin Xin.: Pedosphere 2007, 17(4), 505–512. [10] Alfaro V.M., Teuber K.N., Dumont L.J.C. and Medone V.F.: Agricult. Techn. Santiago 1998, 58(3), 173–180. [11] Mongia A.D., Singh N.T., Mandal L.N. and Guha A.: J. Indian Soc. Soil Sci. 1998, 46(1), 61–66. [12] Wallace A.: Soil Sci. 1989, 147(6), 451–453. [13] Patiram-Rai R.N., Singh K.P. and Prasad R.N.: J. Indian Soc. Soil Sci. 1990, 38(3), 499–503. [14] Hahn G. and Marschner H.: Plant Soil 1998, 199(1), 23–27. [15] Kiepul J.: Pamiêt. Pu³aw. 1992,101, 201–208.

ZAWARTOŒÆ GLINU W ROŒLINACH W ZALE¯NOŒCI OD ZANIECZYSZCZENIA GLEBY KADMEM

Katedra Chemii Œrodowiska Uniwersytet Warmiñsko-Mazurski w Olsztynie

Abstrakt: Celem badañ by³o okreœlenie wp³ywu zanieczyszczenia gleby kadmem (10, 20, 30 i 40 mg Cd × kg–1 gleby) na zawartoœæ glinu w czêœciach nadziemnych i korzeniach owsa, kukurydzy, ³ubinu ¿ó³tego i rzodkiewki. Do neutralizacji kadmu wprowadzonego do gleby zastosowano: kompost, wêgiel brunatny, wapno i bentonit. Na zawartoœæ glinu w roœlinach, oprócz gatunku i organu roœlin, du¿y wp³yw mia³a dawka kadmu, jak równie¿ rodzaj zaaplikowanej substancji neutralizuj¹cej. Korzenie zawiera³y zdecydowanie wiêcej glinu ni¿ czêœci nadziemne roœlin. Najwiêcej glinu stwierdzono w korzeniach ³ubinu ¿ó³tego i kukurydzy, a najmniej w ziarnie i s³omie owsa. Zanieczyszczenie gleby kadmem spowodowa³o wiêksze zmiany w zawartoœci glinu w czêœciach nadziemnych ni¿ w korzeniach roœlin, szczególnie w przypadku kukurydzy i ³ubinu ¿ó³tego. Dawki kadmu w wysokoœci 20 mg (korzenie kukurydzy i ³ubinu ¿ó³tego) lub 40 mg Cd × kg–1 Aluminium Concentration in Plants Depending on Soil Contamination with Cadmium 1649 gleby (czêœci nadziemne kukurydzy i ³ubinu ¿ó³tego) powodowa³y zwiêkszenie zawartoœci glinu. Dalszy wzrost dawek kadmu wywo³ywa³ zmniejszenie zawartoœci glinu w korzeniach tych roœlin. W przypadku czêœci nadziemnych i korzeni rzodkiewki oraz ziarna owsa odnotowano zmniejszenie zawartoœci glinu we wszystkich obiektach zanieczyszczonych kadmem. Zakres oddzia³ywania testowanych substancji neutra- lizuj¹cych na zawartoœæ glinu by³ zró¿nicowany. Dodatki neutralizuj¹ce na ogó³ obni¿a³y zawartoœæ glinu w roœlinach. Wêgiel brunatny, bentonit i wapno wywo³ywa³y wiêksze zmiany w zawartoœci glinu w roœlinach ni¿ kompost. Zawartoœæ glinu wykazywa³a skorelowanie z plonem roœlin. U wiêkszoœci roœlin wyst¹pi³y relacje ujemne w czêœciach nadziemnych (oprócz rzodkiewki), a dodatnie w korzeniach (z wyj¹tkiem ³ubinu ¿ó³tego). Zawartoœæ glinu w poszczególnych organach roœlin by³a skorelowana z licznymi makro- i mikro- elementami, w tym przewa¿nie dodatnio z zawartoœci¹ manganu, ¿elaza, kobaltu, litu, kadmu, miedzi i cynku, a ujemnie z zawartoœci¹ siarki i boru. S³owa kluczowe: zanieczyszczenie kadmem, kompost, wêgiel brunatny, wapno, bentonit, roœliny, zawartoœæ glinu

ECOLOGICAL CHEMISTRY AND ENGINEERING A

Vol. 18, No. 12 2011

Janina GOSPODAREK1

EFFECT OF SOIL CONTAMINATION WITH A MIXTURE OF HEAVY METALS ON BROAD BEAN (Vicia faba L.) SEED QUALITY

ODDZIA£YWANIE SKA¯ENIA GLEBY MIESZANIN¥ METALI CIʯKICH NA JAKOŒÆ NASION BOBU (Vicia faba L.)

Abstract: The work aimed at an assessment of soil contamination with mixtures of heavy metals with zinc and nickel on two levels of pollution on the broad bean seed yield, degrees of injuries caused by broad bean beetle, and the germination energy and ability. The assessment of germination energy and ability of broad bean seeds was tested in laboratory, according to generally used standards. Soil contamination with mixtures of zinc and nickel with cadmium, copper and lead on III level of pollution acc. to the IUNG classification led to a significant decline in broad bean seed yield or its total loss, but the decrease in yield was lower than when the soil was contaminated by zinc or nickel used separately. Soil contamination with mixtures of zinc with nickel, zinc with copper and zinc with lead and nickel with copper on II level of pollution in the IUNG classification also caused a notable decrease in broad bean seed yield. Soil pollution with the tested mixtures of heavy metals did not affect the degree of seed injuries due to Bruchus rufimanus or their germination energy. Soil contamination with a mixture of zinc and nickel in II class of pollution in the IUNG classification leads to a worsening of the seed quality (the percentage of dead seeds increased and the condition of obtained seedlings worsened). The above-mentioned features were also negatively affected by the soil pollution with a mixture of nickel and cadmium on III level of pollution in the IUNG classification. Keywords: heavy metals, soil pollution, Bruchus rufimanus Boh.

One of the ways of agricultural management of heavy metal polluted soils is their destination for seed crops. Soil concentrations of heavy metals such as zinc and nickel on the level of medium pollution in the IUNG classification [1] cause considerable decline in or total loss of broad bean seed yield. On the other hand, copper, lead and cadmium do not adversely affect the quantity or quality of broad bean seed yield and in the case of seeds injured by broad bean beetle (Bruchus rufimanus Boh.) even stimulate their germination ability. Soils contaminated with these elements may be thus designed

1 Department of Agricultural Environment Protection, Agricultural University of Krakow, al. A. Mic- kiewicza 21, 31–120 Kraków, Poland, phone: +48 12 662 44 00, fax +48 12 633 44 43, email: [email protected] 1652 Janina Gospodarek for seed plantation of this crop [2]. A similar beneficial effect of lead and cadmium on broad bean plants was observed in the case when the soil pollution level was equal to I class of pollution in the IUNG classification [3]. Heavy metals which occur in soil jointly often have a different effect on a plant than when present separately [4]. Because among the five heavy metals (Pb, Cd, Cu, Zn and Ni) studied so far and applied separately, zinc and nickel revealed the strongest (negative) effect both on broad bean plant growth and the degree of injuries due to pests, it seems justified to study the effect of soil contamination with mixtures of these two heavy metals with cadmium, lead and copper regarding a possible reduction of this negative effect. The work aimed at an assessment of soil contamination with mixtures of heavy metals with zinc and nickel on two levels of pollution on the broad bean seed yield, degrees of injuries caused by broad bean beetle, and the germination energy and ability.

Material and methods

Broad bean, White Windsor c.v. was cultivated in a control soil with natural heavy metal concentrations without (Control) and with mineral treatment (Control + NPK) and in the soil contaminated with the following mixtures of heavy metals: – Cd – 2.25 mg × kg–1 soil d.m. + Zn – 350 mg × kg–1 soil d.m. (ZnII + CdII), – Cd–4mg× kg–1 soil d.m. + Zn – 1000 mg × kg–1 soil d.m. (ZnIII + CdIII), – Cu–65mg× kg–1 soil d.m. + Zn – 350 mg × kg–1 soil d.m. (ZnII + CuII), – Cu–85mg× kg–1 soil d.m. + Zn – 1000 mg × kg–1 soil d.m. (ZnIII + CuIII), – Pb – 175 mg × kg–1 soil d.m. + Zn – 350 mg × kg–1 soil d.m. (ZnII + PbII), – Pb – 530 mg × kg–1 soil d.m. + Zn – 1000 mg × kg–1 soil d.m. (ZnIII + PbIII), – Ni – 62.5 mg × kg–1 soil d.m. + Zn – 350 mg × kg–1 soil d.m. (NiII + ZnII), – Ni – 110 mg × kg–1 soil d.m. + Zn – 1000 mg × kg–1 soil d.m. (NiIII + ZnIII), – Cd – 2.25 mg × kg–1 soil d.m. + Ni – 62.5 mg × kg–1 soil d.m. (NiII + CdII), – Cd–4mg× kg–1 soil d.m. + Ni – 110 mg × kg–1 soil d.m. (NiIII + CdIII), – Cu–65mg× kg–1 soil d.m. + Ni – 62.5 mg × kg–1 soil d.m. (NiII + CuII), – Cu–85mg× kg–1 soil d.m. + Ni – 110 mg × kg–1 soil d.m. (NiIII + CuIII), – Pb – 175 mg × kg–1 soil d.m. + Ni – 62.5 mg × kg–1 soil d.m. (NiII + PbII), – Pb – 530 mg × kg–1 soil d.m. + Ni – 110 mg × kg–1 soil d.m. (NiIII + PbIII). The level of soil contamination corresponded to II and III class of pollution acc. to the classification suggested by IUNG in Pulawy [1]. The plants were cultivated in plastic pots with 9.8 kg d.m. of soil under field conditions. Detailed description of the methods of heavy metal supply into the soil was presented in another publication [5]. The experiment was conducted in 2008 on degraded chernozem developed from loess with acid reaction (pH in 1 mol × dm–3 KCl solution was 5.5 and in water 6.3) and organic carbon content 1.13 %. Harmfulness of broad bean beetle was estimated on the basis on injured seeds in relation to seed total mass. The assessment of germination energy and ability of broad bean seeds was tested in laboratory, according to generally used standards. The test was conducted in Petri dishes on filter paper as a medium. Germination energy was assessed after 4 days and germination ability after 14 days. Effect of Soil Contamination with a Mixture of Heavy Metals on Broad Bean... 1653

The significance of differences between the means were tested by means of an one-way ANOVA. Means were differentiated using the Duncan test on the significance level p < 0.05.

Results and discussion

Soil contamination with mixtures of most of the analysed heavy metals with zinc on a lower level of pollution caused a marked decline in the number of pods and seed weight per plant in comparison with the minerally fertilized control (Table 1). Only the mixture of zinc and cadmium did not significantly affect these features. The soil contamination with a mixture of zinc and lead most strongly reduced the number of developed pods and seeds. In comparison with the mixtures of zinc with other metals, the soil contamination with nickel mixtures with copper, cadmium and lead on the II pollution level limited pod and seed formation by broad bean to a smaller degree. Contamination of soil with a mixture of zinc and nickel and zinc and cadmium on a higher level of pollution weakened broad bean plant growth so that they were unable to form seeds. Contamination with the other analysed heavy metals on a higher level significantly decreased the number of formed pods and seeds in comparison with the minerally fertilized control. The effect was stronger than in soil contaminated with the mixtures on II pollution level. In the Author’s former research, as a result of soil contamination only with zinc on III level of pollution the plants did not form seeds [2, 3, 5–7].

Table 1 Characteristics of broad bean seeds from plants cultivated in natural soil and in heavy metal contaminated soil, and degree of injuries due to Bruchus rufimanus Boh.

Average number Average seed weight Weight of seeds injured of pods perplant per plant by Bruchus rufimanus Boh. Object [percent in relation to [percent in relation to [percent in relation to Control + NPK] Control + NPK] Control + NPK] ZnII + CuII 25.00 abcd* 23.05 a 33.40 a ZnII + NiII 25.00 abcd 15.84 a 40.63ab ZnII + CdII 62.50 def 50.15 abc 47.13 ab ZnII + PbII 16.67 ab 6.68 a 36.58 ab NiII + CuII 54.17 cde 33.00 ab 72.05 ab NiII + PbII 66.67 def 49.89 abc 61.13 ab NiII + CdII 58.33 de 48.24 abc 108.53 b NiIII + CdIII 15.63 abc 1.94 a 80.09 ab NiIII + PbIII 25.00 abcd 18.34 a 69.05ab ZnIII + PbIII 8.33 a 1.35 a 60.72 ab ZnIII + CuIII 37.50 abcde 17.81 a 109.73 b Control + NPK 100.00 f 100.00 c 100.00 ab Control 76.04 ef 91.07 bc 93.38 ab * Values for individual metals or control marked by different letters in columns are statistically different (p < 0.05). 1654 Janina Gospodarek

However, nickel present in soil usually caused a marked decrease in the number and weight of formed seeds [2, 3] or prevented seeds being formed by broad bean [5,6]. Despite quite considerable diversification between the analysed objects as to the degree of seed injuring by broad bean beetle, statistical analysis did not reveal any significant differences between the objects where the soil was contaminated with heavy metals and the control soil. In the former research soil contamination with nickel on III level of pollution led to a decrease in the degree of seed injuries by broad bean beetle. Decreased seed attractiveness for this pest was also observed under conditions of soils contaminated with copper and lead [2]. Bruchus beetles proved less harmful for the seeds of plants cultivated under conditions of soils contaminated with single heavy metals on I level of pollution in comparison with the minerally fertilized control soil [3]. Also in the presented investigations on most contaminated soils the percentage of seeds injured by broad bean beetle was lower than in the minerally fertilized control. On the other hand, in the research where the above-mentioned metals were applied jointly on the level of elevated content [8] no significant effect was registered on the degree of seed injuring by Bruchus larvae. Some Bruchidae during larval development absorb and accumulate zinc and copper, whereas calcium, magnesium, iron or manganese are most- ly excreted [9]. On the other hand, such metals as cadmium or mercury reveal an inhibitory effect on some enzyme activity in larvae of Acanthoscelides obtectus Say [10]. Too few seeds were collected from the objects with the soil contaminated by the mixture of lead and copper with zinc on III level of pollution to test the germination energy and ability. The other analyzed objects did not differ significantly with respect to germination energy (Fig. 1). Between 0 % and 41 % of seeds germinated 4 days after the test start, the most numerous in conditions of the soil contaminated with a mixture of nickel and lead on a higher level. On the other hand, in the object contaminated with

ab c a a a a a bc a ab a 100 90 healthy 80 shoots [%] 70 swelled and not 60 germinated 50 b ab ab ab b b ab a ab ab b seeds [%]

eeds [%] 40

S dying shoots [%] 30 20 10 0

Control

ZnII + NiII

NiII + PbII

NiII + CdII

NiII + CuII

ZnII + PbII

ZnII + CdII ZnII + CuII

NiIII + CdIII

NiIII + PbIII

Control + NPK

Fig. 1. Germination energy of broad bean seeds originating from plants growing in natural and heavy metal contaminated soil. Values for individual metals or control and for individual features, marked by different letters are statistically different (p < 0.05). Assessments were presented only if there was statistical differentiation between objects. In other cases differences were statistically insignificant Effect of Soil Contamination with a Mixture of Heavy Metals on Broad Bean... 1655

ab ab ab ab a a a b ab a ab 100 90 healthy 80 shoots [%] 70 swelled and not 60 germinated 50 ab ab ab ab ab ab b a ab ab ab seeds [%]

eeds [%] 40

S dying shoots [%] 30 20 10 0

Control

ZnII + NiII

NiII + PbII

NiII + CdII

NiII + CuII

ZnII + PbII

ZnII + CdII ZnII + CuII

NiIII + PbIII

NiIII + CdIII

Control + NPK

Fig. 2. Germination ability of broad bean seeds originating from plants growing in natural and heavy metal contaminated soil. Values for individual metals or control and for individual features, marked by different letters are statistically different (p < 0.05). Assessments were presented only if there was statistical differentiation between objects. In other cases differences were statistically insignificant. zinc and nickel on II pollution level significantly more dead seeds were found than in the conditions of the control soil fertilized minerally. Quite a few dead seeds were found also on the object with the soil contaminated with a mixture of nickel and cadmium on a higher level of pollution. On the latter object the share of dead seeds increased during the testing period (Fig. 2). Seeds from this object were characterized by the lowest germination ability. Also the number of lateral roots obtained from seedlings was low (Table 2).

Table 2 Characteristics of germinating broad bean seeds from plants cultivated in natural soil and in heavy metal contaminated soil.

Underground Number of lateral roots Objects Shoot length part length 2mm ZnII + CuII 1.700 ab* 6.000 a 8.100 ab ZnII + NiII 0.872 a 4.561 a 3.556 ab ZnII + CdII 2.194 ab 4.208 a 5.972 ab ZnII + PbII 1.333 ab 4.417 a 8.417 ab NiII + CuII 1.170 ab 6.375 a 8.183 ab NiII + PbII 1.767 ab 6.583 a 9.650 ab NiII + CdII 2.200 ab 5.733 a 11.667 bb NiIII + CdIII 4.333 b 4.333 a 6.500 ab NiIII + PbIII 1.611 ab 4.861 a 11.806 bb Control + NPK 2.458 ab 4.925 a 11.167 bb Control 0.533 a 4.533 a 6.033 ab * Values for individual metals or control marked by different letters in columns are statistically different (p < 0.05). 1656 Janina Gospodarek

Quite a high number of dead seeds was also registered under conditions of soil contaminated with mixtures of zinc and nickel and zinc with cadmium on a lower level. The number of lateral roots in seedlings obtained from the seeds on the object contaminated with zinc and nickel on II level of pollution was significantly lower than in seedlings from seeds originating from the control plants receiving mineral fertilizers. On the other hand no apparent differences were found in the germination ability and condition of the obtained seedlings between the seeds from plants cultivated in the soil contaminated with mixtures of zinc with copper and lead and nickel with copper, lead and cadmium on II level of pollution and also nickel with lead on III level of pollution, and the seeds from the control plants fertilized minerally. In the Author’s former investigations, soil contamination with cadmium, lead and copper applied separately on III level of pollution did not affect negatively the germination ability of broad bean seeds [2]. Soil contamination with single heavy metals (Cd, Pb, Cu, Zn, Ni) on I level of pollution acc. to the IUNG classification did not influence negatively germination energy or ability. Some of the tested metals (lead and cadmium) even revealed a positive effect on the number of formed seeds and their germination ability [3].

Conclusions

1. Soil contamination with mixtures of zinc and nickel with cadmium, copper and lead on III level of pollution acc. to the IUNG classification led to a significant decline in broad bean seed yield or its total loss, but the decrease in yield was lower than when the soil was contaminated by zinc or nickel used separately. 2. Soil contamination with mixtures of zinc with nickel, zinc with copper and zinc with lead and nickel with copper on II level of pollution in the IUNG classification also caused a notable decrease in broad bean seed yield. 3. Soil pollution with the tested mixtures of heavy metals did not affect the degree of seed injuries due to Bruchus rufimanus or their germination energy. 4. Soil contamination with a mixture of zinc and nickel in II class of pollution in the IUNG classification leads to a worsening of the seed quality (the percentage of dead seeds increased and the condition of obtained seedlings worsened). The above-mention- ed features were also negatively affected by the soil pollution with a mixture of nickel and cadmium on III level of pollution in the IUNG classification.

References

[1] Kabata-Pendias A. and Piotrowska M.: Ocena stopnia zanieczyszczenia gleb i roœlin metalami ciê¿kimi i siark¹. Wyd. IUNG, Pu³awy 1993., ser. P, 53 pp. [2] Gospodarek J.: Ecol. Chem. Eng. 2006, 13(6), 497–504. [3] Gospodarek J.: Ecol. Chem. Eng. 2006, 13(1–2), 47–54. [4] Cury³o T.: Zesz. Nauk. AR w Krakowie, 1999, 64, 53–65. [5] Gospodarek J.: Ecol. Chem. Eng. 2008, 15(1–2), 55–64. [6] Gospodarek J.: Ecol. Chem. Eng. 2006, 13(11), 1231–1240. [7] Gospodarek J.: Ecol. Chem. Eng. 2008, 15(1–2), 55–64. [8] Jaworska M. and Gospodarek J.: Chem. In¿. Ekol. 2003, 10(3–4), 291–295. [9] Ernst W.H.O.: J. Stor. Prod. Res. 1993, 29(1), 53–62. [10] Osuala C.J., Domer R.L. and Nielsen S.S.: Compar. Biochem. Physiol. Part B 1994, 107(2), 241–248. Effect of Soil Contamination with a Mixture of Heavy Metals on Broad Bean... 1657

ODDZIA£YWANIE SKA¯ENIA GLEBY MIESZANIN¥ METALI CIʯKICH NA JAKOŒÆ NASION BOBU (Vicia faba L.)

Wydzia³ Rolniczo-Ekonomiczny Uniwersytet Rolniczy im. Hugona Ko³³¹taja w Krakowie

Abstrakt: Celem pracy by³o okreœlenie wp³ywu ska¿enia gleby mieszaninami metali ciê¿kich (kadmu, miedzi i o³owiu) z cynkiem i niklem na dwóch poziomach zanieczyszczenia (wg II i III klasy zanieczyszczenia zgodnie z klasyfikacj¹ IUNG) na plon nasion bobu, stopieñ ich uszkodzenia przez str¹kowca bobowego oraz energiê i zdolnoœæ kie³kowania. Szkodliwoœæ str¹kowca bobowego oceniono na podstawie masy nasion uszkodzonych w stosunku do ogólnej masy nasion. Ocenê energii i zdolnoœci kie³kowania nasion bobu przeprowadzono w warunkach laboratoryjnych, zgodnie z ogólnie przyjêtymi normami. Ska¿enie gleby mieszaninami cynku i niklu z kadmem, miedzi¹ i o³owiem na poziomie III stopnia zanieczyszczenia wg klasyfikacji IUNG prowadzi do znacznego spadku plonu nasion bobu lub ca³kowitej jego utraty. Ska¿enie gleby mieszaninami cynku z niklem, cynku z miedzi¹ i cynku z o³owiem oraz niklu z miedzi¹ na poziomie II stopnia zanieczyszczenia wg klasyfikacji IUNG prowadzi tak¿e do znacznego spadku plonu nasion bobu. Ska¿enie gleby badanymi mieszaninami metali ciê¿kich nie wp³ywa na stopieñ uszkodzenia nasion przez str¹kowca bobowego oraz ich energiê kie³kowania. Ska¿enie gleby mieszanin¹ cynku i niklu na poziomie II klasy zanieczyszczenia zgodnie z klasyfikacj¹ IUNG prowadzi do pogorszenia jakoœci nasion (wzrasta odsetek nasion martwych, pogarsza siê kondycja uzyskanych siewek). Niekorzystnie na ww. cechy oddzia³uje tak¿e ska¿enie gleby mieszanin¹ niklu i kadmu na poziomie III klasy zanieczyszczenia wg klasyfikacji IUNG. S³owa kluczowe: metale ciê¿kie, zanieczyszczenie gleby, Bruchus rufimanus Boh.

ECOLOGICAL CHEMISTRY AND ENGINEERING A

Vol. 18, No. 12 2011

Pawe³ WOLSKI1*, Iwona ZAWIEJA1 and Lidia WOLNY1

IMPACT OF TEMPERATURE ON VISCOSITY OF SEWAGE SLUDGE AFTER CONDITIONING

WP£YW TEMPERATURY NA LEPKOŒÆ KONDYCJONOWANYCH OSADÓW ŒCIEKOWYCH

Abstract: The technology of wastewater treatment is always associated with utilization of sewage sludge. The process of sewage sludge utilization aims at transforming (biologically, physically or chemically) sewage sludge into a state which does not pose any threats for human health and natural environment. Conditioning – a process which has an impact on the structure and properties of sewage sludge – allows for more efficient removal of water from sewage sludge. Also, temperature is a crucial parameter in the final stage of utilization and transportation of sewage sludge. The paper presents the results of the investigations on viscosity of chemically conditioned sewage sludge exposed to selected temperatures and variable sheer velocity gradients. Municipal sewage sludge and sludge from cellulose industry were used as substrates. The impact of temperature on viscosity of fermented sewage sludge and also the impact of a polyelectrolyte dose and a velocity gradient on sewage sludge viscosity were investigated. Viscosity of sewages sludge was determined in the temperature range of 20 to 36 oC for every 2 oC at variable sheer velocity gradients (60, 100, 200 r/min). The HAAKE Viscotester 7L/R plus and the Termostat DC 10 bathtube were used in the investigations. Keywords: sewage sludge, conditioning, viscosity, temperature

The sewage treatment technology is closely associated with the disposal of sewage sludge. This process involves the transformation of sewage sludge (by biological, physical or chemical methods), which is intended to bring the sludge to a state that does not pose any hazards to the life or health of population or to the natural environment [1].

1 Institute of Environmental Engineering, Technical University of Czestochowa, ul. BrzeŸnicka 60a, 42–200 Czêstochowa, Poland, phone: +48 34 325 09 17, fax: +48 34 372 13 04, email: [email protected] * Corresponding author: email: [email protected] 1660 Pawe³ Wolski et al

Until recently, the sewage sludge treatment process went unnoticed. Unfortunately, the sewage sludge management in sewage treatment plants continues to be a topical and not completely resolved problem. Failures in this field bring about serious consequences and may be a source of risk to the natural environment of man. The construction of new sewage treatment plants and the intensification of treatment processes is accompanied also by the increase in the amount of sewage sludge which accounts for 1–2 % of the total volume of sewage flowing to the sewage treatment plant. Of the treatment plant’s overall costs, the costs of the construction and operation of sewage sludge treatment equipment can account for as much as 50 %. By treating sewage sludge as a raw material of some fertilizer or energy value, at least a partial return on the incurred outlays could be obtained [2, 3]. The neutralization of sewage sludge in treatment plants involves the removal of water that it contains, whereby a reduction of the sewage sludge volume is achieved. The process whereby this effect is achieved is conditioning, that is one of the processes which have the effect of changing the structure and properties of sewage sludge, enabling a more efficient removal of water contained in the sewage sludge. Change in the form of sewage sludge is the change of its structure, which also influences its viscosity [4, 5]. For the majority of sewage sludges, physical and chemical conditioning processes are used, which can run either under natural conditions or with the use of mechanical devices. The selection of the sewage sludge conditioning method is most often influenced by the possibility of subsequent use and utilization of the sewage sludge, as the conditioning process determines the final products formed [6, 7]. One of the conditioning methods is by changing the temperature. The temperature is one of the important parameters in the final stage of neutralizing and transporting sewage sludge (rheological parameters) [8, 9]. The purpose of the studies undertaken was to determine the viscosity of chemically conditioned sewage sludges subjected to the action of temperature and variable magnitudes of the shearing velocity gradient.

Experimental

The substrate of the tests was the sludge from municipal wastewater treatment plant after aerobic stabilization process. Its sewage treatment capacity was 1000 m3/d. The characteristic of sludge was as follow: dry matter 18.38 g/dm3, initial hydration 97.3 %, capillary suction time (CST) was 154 s, resistivity 21.4 × 1012 m/kg and viscosity was equal 132 mPa × s. Sludge was conditioned with low and high cationic polyelectrolytes. The capillary suction time was used to determine polyelectrolyte dose. Viscosity of sludge was measured in the range of temperatures from 20 to 36 oC with the step of 2 oC (20, 22, 24, 26, 28, 30, 32, 34, 36), for the variable shear velocity gradient (60, 100 and 200 rpm) and for different polyelectrolytes doses. The viscosity was measured with HAAKE Viscotester 7L/R plus – rotational viscosimeter designed to perform the fast determination of viscosity in accordance with the ISO 2555 standards (Fig. 1). To maintain the required temperature thermostat DC10 was used. Impact of Temperature on Viscosity of Sewage Sludge After Conditioning 1661

Fig. 1. Viscosity measurement set

Analysis of investigation results

The effect of temperature on the change in the viscosity of sewage sludges examined

By analyzing the curves of the viscosity of sewage sludges conditioned with polyelectrolytes versus temperature it was found that the sewage sludge viscosity decreased with increasing temperature. Already at a slight temperature increase by 2 oC, a drop in viscosity was noted. Variations in viscosity with increasing sewage sludge temperature at different shearing velocities (60, 100, 200 rpm) are illustrated in Figures 2 to 4. Similar relationships can be observed for sewage sludges conditioned with a weak-cationic polyelectrolyte (Praestol 650BC).

140 raw sludge

120 sludge + polyelectrolyte (1 mg/g d.m.) 100 sludge + × polyelectrolyte 80 (2 mg/g d.m.)

sludge + 60 polyelectrolyte (3 mg/g d.m.)

Viscosity [mPa s] 40 sludge + polyelectrolyte (4 mg/g d.m.) 20

0 20 22 24 26 28 30 32 34 36 Temperature [oC]

Fig. 2. Influence of temperature on the viscosity of the sludge conditioned with different doses of polyelectrolyte Praestol 658BC-S (rotational speed 60 rpm) 1662 Pawe³ Wolski et al

120 raw sludge

sludge + 100 polyelectrolyte (1 mg/g d.m.) 80 sludge + × polyelectrolyte (2 mg/g d.m.) 60 sludge + polyelectrolyte (3 mg/g d.m.)

Viscosity [mPa s] 40 sludge + polyelectrolyte 20 (4 mg/g d.m.)

0 20 22 24 26 28 30 32 34 36 Temperature [oC]

Fig. 3. Influence of temperature on the viscosity of the sludge conditioned with different doses of polyelectrolyte Praestol 658BC-S (rotational speed 100 rpm)

70 raw sludge

60 sludge + polyelectrolyte (1 mg/g d.m.) 50 sludge + × polyelectrolyte 40 (2 mg/g d.m.)

sludge + 30 polyelectrolyte (3 mg/g d.m.)

Viscosity [mPa s] 20 sludge + polyelectrolyte (4 mg/g d.m.) 10

0 20 22 24 26 28 30 32 34 36 Temperature [oC]

Fig. 4. Influence of temperature on the viscosity of the sludge conditioned with different doses of polyelectrolyte Praestol 658BC-S (rotational speed 200 rpm)

The effect of the dose of polyelectrolyte on the change in the viscosity of sewage sludges examined

From the analysis of the graphs showing the effect of the polyelectrolyte dose on sewage sludge viscosity it was found that the increase in the polyelectrolyte dose caused a decrease in sewage sludge viscosity (Figs. 5–7). With increasing polyelectrolyte dose, the sewage sludge viscosity decreased, reaching minimum values at the highest doses applied. Impact of Temperature on Viscosity of Sewage Sludge After Conditioning 1663

140

120 sludge + PRAESTOL 650BC sludge + PRAESTOL 658BC-S 100

×

80

60

Viscosity [mPa s] 40

20

0 0 0.5 1 1.5 2.0 2.5 3 3.5 4 Polyelectrolyte dose [mg/g d.m.]

Fig. 5. Influence of polyelectrolytes dose on the viscosity of tested sludge (rotational speed 60 rpm; temperature 20 oC)

140 sludge + PRAESTOL 650BC 120 sludge + PRAESTOL 658BC-S 100

×

80

60

Viscosity [mPa s] 40

20

0 0 0.5 1 1.5 2.0 2.5 3 3.5 4 Polyelectrolyte dose [mg/g d.m.]

Fig. 6. Influence of polyelectrolytes dose on the viscosity of tested sludge (rotational speed 60 rpm; temperature 28 oC)

100

90 sludge + PRAESTOL 650BC 80 sludge + PRAESTOL 658BC-S 70

× 60 50 40

Viscosity [mPa s] 30 20 10 0 0 0.5 1 1.5 2.0 2.5 3 3.5 4 Polyelectrolyte dose [mg/g d.m.]

Fig. 7. Influence of polyelectrolytes dose on the viscosity of tested sludge (rotational speed 60 rpm; temperature 36 oC) 1664 Pawe³ Wolski et al

The effect of the velocity gradient on the change in the viscosity of sewage sludges examined

By analyzing the effect of the gradient of velocity on the change in the viscosity of sewage sludge it was found that the increase in shearing velocity was accompanied by a decrease in viscosity (Fig. 8). The viscosity curves show a drop in viscosity against the increase in shearing velocity. The decrease in viscosity with the increase in shearing velocity was noted for the entire of temperature examined (20–36 oC).

140

120

100

× 80

60

Viscosity [mPa s] 40 raw sludge 20 sludge + PRAESTOL 650BC (2 mg/g d.m.) sludge + PRAESTOL 658BC-S (2 mg/g d.m.)

0 60 100 200 Shear velocity [rpm]

Fig. 8. Influence of shear velocity gradient on the viscosity of tested sludge in the temperature 20 oC

Conclusions

From the investigation carried out it was found that the viscosity of sewage sludge decreased with increasing temperature. Already at a slight temperature increase by 2 oC, a drop in the viscosity of sludges examined was noted. By analyzing the effect of polyelectrolyte doses on the sewage sludge viscosity it was determined that the increase in the polyelectrolyte dose results in a decrease in sewage sludge viscosity. With the increase in the dose of polyelectrolytes used, the viscosity decreased, reaching minimum values at the highest doses applied. In most cases, introducing doses of a strong-cationic polyelectrolyte, Praestol 658BC-S, caused greater viscosity decreases compared with the weak-cationic polyelectrolyte Praestol 650BC with the identical doses. The viscosity of sludge treated with the strong-cationic polyelectrolyte Praestol 658BC-S at a temperature of 20 oC and a rotational speed of 100 rpm was 98 mPa × s, while with the highest doses of this polyelectrolyte and under the identical temperature and rotational speed conditions it was 45 mPa × s. The investigation carried out indicates that the increase in shear velocity is accompanied by a decrease in the viscosity of sewage sludges examined. This applies to Impact of Temperature on Viscosity of Sewage Sludge After Conditioning 1665 untreated sludge, as well as to sludge conditioned with different doses of poly- electrolyte. The values on the viscosity curves decrease from the first examined velocity of 60 rpm, then consecutively increased to 100 rpm and 200 rpm, against the increase in shear velocity.

Acknowledgements

The research was founded by the BW-401/202/07 resources.

References

[1] Bieñ J., Stêpniak L. and Wolny L.: UltradŸwiêki w dezynfekcji wody i preparowaniu osadów œciekowych przed ich odwadnianiem, Wyd. Polit. Czêstochowskiej, Czêstochowa 1995. [2] Bieñ J.B.: Osady œciekowe. Teoria i praktyka, Wyd. Polit. Czêstochowskiej, Czêstochowa 2002. [3] Leschber R. and Spinosa L.: Developments in sludge characterization in Europe, Water Sci. Technol. 1998, 38(2), 1–7. [4] Szwajczak E.: Wp³yw mikrostruktury p³ynu na lepkoœæ, Mechanika w Medycynie, Rzeszów 2002, 6, 167–176. [5] Lee C.H. and Liu J.C.: Sludge dewaterability and floc structure in dual polymer conditioning, Adv. Environ. Res. 2001, 5, 129–136. [6] Bieñ J.B., Bieñ J.D. and Matysiak B.: Gospodarka odpadami w oczyszczalniach œcieków, Wyd. Polit. Czêstochowskiej, Czêstochowa 1999. [7] Oleszkiewicz J.A.: Gospodarka osadami œciekowymi. Poradnik decydenta, Lem s.c., Kraków 1998. [8] Szlezyngier W.: Podstawy reologii polimerów, Wyd. Polit. Rzeszowskiej, Rzeszów 1994. [9] Slatter P.T.: The rheological characterization of sludge, Water Sci. Technol. 1997, 36(11), 9–18. [10] Wojtala £.: Wp³yw temperatury na lepkoœæ kondycjonowanych osadów œciekowych, praca dyplomowa, Politechnika Czêstochowska, Czêstochowa 2009.

WP£YW TEMPERATURY NA LEPKOŒÆ KONDYCJONOWANYCH OSADÓW ŒCIEKOWYCH

Instytut In¿ynierii Œrodowiska, Wydzia³ In¿ynierii i Ochrony Œrodowiska, Politechnika Czêstochowska

Abstrakt: Oczyszczanie œcieków jest œciœle zwi¹zane z problemem unieszkodliwiania osadów œciekowych. Proces unieszkodliwiania jest przekszta³ceniem, które ma doprowadziæ osady do stanu, który nie stwarza zagro¿eñ dla ¿ycia lub zdrowia ludnoœci oraz dla œrodowiska. Kondycjonowanie, czyli jeden z procesów maj¹cy wp³yw na zmianê struktury i w³aœciwoœci osadów, pozwala na bardziej skuteczne usuwanie zawartej w osadach wody. Równie¿ temperatura jest wa¿nym parametrem w koñcowym etapie unieszkodliwiania i transportu osadów œciekowych. W artykule przedstawiono wyniki badañ dotycz¹ce wyznaczenia lepkoœci kondycjonowanych chemicznie osadów œciekowych, poddanych dzia³aniu temperatury oraz zmiennym wartoœciom gradientu prêdkoœci œcinania. Przeprowadzono badania komunalnych osadów œciekowych oraz osadów z przemys³u celulozowego. Ocenie poddano zarówno wp³yw temperatury na lepkoœæ przefermen- towanych osadów œciekowych, jak i wp³yw dawki polielektrolitów oraz gradientu prêdkoœci na lepkoœæ badanych osadów œciekowych. Lepkoœæ osadów zosta³a wyznaczona w zakresie temperatur od 20 do 36 oC, co 2 oC, przy zmiennym gradiencie prêdkoœci œcinania (60, 100, 200 obr/min). W badaniach wykorzystano wiskozymetr HAAKE Viscotester 7L/R plus oraz wannê Termostat DC10. S³owa kluczowe: osady œciekowe, kondycjonowanie, lepkoœæ, temperatura

ECOLOGICAL CHEMISTRY AND ENGINEERING A

Vol. 18, No. 12 2011

Teresa RAUCKYTE-¯AK1 and S³awomir ¯AK1

CHANGEABILITY OF THE SPECIATION FORMS OF HEAVY METALS IN SOIL SUBJECT TO MANY YEARS OF FERTILIZATION WITH WASTEWATERS FROM VEGETABLE FAT PRODUCTION

ZMIENNOŒÆ FORM SPECJACYJNYCH METALI CIʯKICH W GLEBIE PODDANEJ WIELOLETNIEMU NAWO¯ENIU ŒCIEKAMI Z PRODUKCJI T£USZCZÓW ROŒLINNYCH

Abstract: Agro-utilization of pretreated industrial wastewaters from the vegetable fat production for irrigation and as fertilizers is, among others, limited by heavy metal concentrations in wastewaters and the soil where wastewaters are directed. This paper presents the results of 4-year research on the influence of the irrigation process and fertilization with wastewaters on the concentration and changeability of heavy metal (Cd, Cr, Cu, Hg, Ni, Pb, Zn) speciation forms in the superficial soil layer. The fractional composition of metals was determined in accordance with Tessier’s methodology. The obtained results prove a significant stability of speciation loads of metals determined in the examined soil within the experimental period. Analysis of the fractional composition showed an insignificant decreasing tendency of the leached fraction percentage and an increase in the carbonate fraction share in the case of the majority of the analyzed heavy metals. Keywords: heavy metals, metals speciation forms, agro-utilization of wastewaters from vegetable fat production

After physicochemical pretreatment, industrial wastewaters from vegetable fat production are free from heavy metals, and in the pollutant load there are mainly soluble low molecular substances of vegetable origin (ie lecithins, glycerols, proteins, aldehydes, ketones, lactones, sterols, tocopherols) and orthophosphates from refining processes [1, 2]. Composition qualifies wastewaters of this type to the group of biodegradable sectors, enabling them to be agriculturally used for irrigation and as fertilizers [3]. The fundamental limitation of agro-utilization is the limitation with nitrogen and heavy metal threshold concentrations in soils, where wastewaters are directed [3, 4]. The share of metal speciation forms in soil – the multi-phase polydispersion system – depends on

1 Department of Chemical Technology and Engineering, University of Technology and Life Sciences, ul. Seminaryjna 3, 85–326 Bydgoszcz, Poland, phone: +48 52 374 90 63, fax: +48 52 374 90 05, email: [email protected] 1668 Teresa Rauckyte-¯ak and S³awomir ¯ak a number of factors: minerals forming the solid phase, organic matter content and composition, content, type and water transfer, local climate, variety of the world of microorganisms, the level of energetic processes, etc. [5, 6]. With many various factors deciding about the quality of a given soil, the introduction of the outer substance load must result in significant changes in the past mechanisms of chemical, biochemical and biological transformations [7, 8]. It is like that also in the case of introducing pretreated wastewaters from the production of fats of vegetable origin [9–11]. Wastewaters, especially the pretreated ones additionally contain mineral substances – orthophosphates and the calcium ion (ie in the form of Ca2+ × aq, CaOH+ or 2+ × 2– 2+ × – ion pairs Ca CO3 and Ca HCO3 ), which show definite reactivity in relation to soil components [2, 12]. Only the toxic chemical components which can occur in wastewaters are the subject to a limit, while the quantity of water does not include any limitations, and this factor significantly influences the dynamics of transformations and transport in soil [13]. Introducing a new load of organic and mineral substances with a significant amount of water can lead to significant transformations in soils and at the same time change the metal speciation form composition, especially in the long-term period of time when the wastewaters are used for agriculture [8, 18].

Materials and methods

Soil material (from the depth of surface layer 0.0–20.0 cm) for the analyses of the individual heavy metal fractions was collected for 4 years at regular intervals – one series every half a year (one in April, the second one in October) from the experimental field (of the area of 15.0 ares) irrigated and fertilized with pretreated wastewaters from the production of fats of vegetable origin. Analyses were made in accordance with Tessier’s methodology, in the detailed form presented in Table 1 [14–18]. There were the following parameters [mg/dm3] of the pretreated wastewaters during the time of running experiments: total nitrogen (TN) 10.9–53.9, total phosphorus (TP) 7.4–39.3, etheric extract (EE) 2.7–28.5, total suspensions (TSS) 38.9–105.2, chemical oxygen demand (COD) 1307.9–3095.7, biochemical oxygen demand (BOD5) 448.9–2055.8 and reaction (pH) 7.8–8.8. The wastewater pretreatment process was carried with the use of coagulation with lime milk (at a dose of 0.75–1.0 kg CaO/m3 of wastewaters), aided with flocculation (Praestol 2340 BC Stockhausen) at a dose of 15.0 g/m3 of wastewaters and pressure flotation (saturation time 10.0 min, saturation pressure 500 kPa, level of pretreated waste recirculation 20 %), aided with hydrogen peroxide oxidation (at a dose 3 of 150.0 gH2O2/m of wastewaters) [4].

Discussion

The aim of the research was to check the influence of four-year irrigation and fertilization with pretreated wastewaters from the vegetable fat production on the fractional composition and changeability of heavy metal fractional loads contained in Changeability of the Speciation Forms of Heavy Metals in Soil... 1669 2 O 2 was 2 O Table 1 (1.0 M) 2 3 30%H 3 30%H 3 C. Analyses were made after o (v/v) was added and shaken for a 3 C. Next, 200.0 cm o in 20 % HNO b 4 COONH 3 twice until releasing white ashes. Next, 400.0 cm 2 O 2 were added to the solid sample obtained from the procedure run for 2 and after intensive mixing (at pH 7.0) shaken for 1.0 hour at room O 2 2 and H (3.0 M) CH 3 3 COOH was added to the dried sample of the solid phase after applying the 3 were subject to AAS analyses (apparatuses: BUCK Scientific 210 VGP and Spectr (1.0 M) CaCl 3 sI–V 30 % aqueous solution of H 3 COOH to pH 7.0. After mixing, the mixture was shaken for 5.0 hours at room temperature. During 3 HCl solution in 25 % CH × OH 2 and 200.0 cm 3 and acidifying 80 % CH 4 (0.04 M) of NH filtrate for determination of fraction (0.02 M) HNO b 3 3 where agro-utilization of pretreated wastewaters from vegetable fats production was carried out COONH 3 Description of Tessier’s methodology used to determine the heavy metal fraction in the examined soil, fraction III, then the mixture was mixed (pH 2.0) and shaken for 135.0 min at the temperature of 85 ± 2 procedure to fraction II and mixed. Samples (of pH 2.0) were shaken for 5.0 hours at the temperature of 96 ± 3 temperature. Analyses were made after separating solid components with the use of filtration. separating the solid components. added and shaken for 135.0 minutes. After the time, 400.0 cm 30.0 min. at room temperature (pH 2.0). CH was added and shaken for 30.0 min at boiling point. Analyses were made after cooling. this operation the reaction was controlled: when pH > 7.0 – it was acidified. The analyses were made after separating solid components. Fraction Details concerning preparation procedure I exchangeable Soil samples (100.0 g) were treated with 400.0 cm II carbonate Dried soil samples which remained after carrying out the procedure for fraction I were subject to extraction, adding 800.0 cm V residual Shaken for 1.0 hour at boiling point, at the same time adding HNO III Fe/Mn oxides 800.0 cm IV organic 200.0 cm Based on the publications [14–18]; number Fraction AA Varian 220 SS). a 1670 Teresa Rauckyte-¯ak and S³awomir ¯ak the soil where wastewaters were directed. The problem of metal speciation in soil material is strictly related to the particular specificity of a given soil [5, 6]. On the basis of the analyses made, the insignificant periodical fluctuations of total concentrations for individual metals in the analyzed layer were found. The problem was presented and interpreted in detail in publication [4], and the present paper is just the extension of the problems discussed in the publication. Despite applying wastewaters of reaction pH 7.8–8.8 (after coagulation with lime milk) within 4 years of experiments, no change was found in soils from experimental field because the level of this parameter was within the range of pH » 6.7 ± 0.2. Therefore, it is the reaction level at which the ionic metal forms (M) such as: chromium, zinc, copper, lead or mercury show particular preference to form difficult to solve hydroxide forms of type M(OH)n(s) [19, 20]. Also, in the case of these metal carbonates, the values of their solubility products are at a very low level (below 1.0 × 10–10), which shows that these forms are favorable to form difficult to solve MCO3(s) carbonates in conditions of the subject soil reaction [21]. At the significant participation of chemical and biochemical reactions which lead to mineralization of some organic contaminations directed to soil, the stability of soil reaction can also result from forming a strong carbonate–hydrocarbonate buffer system. On the basis of the present research within four years of experiment, it was found that the total average metal concentrations during this period were not subject to significant changes [4]. In the case of chromium and nickel, an insignificant growth tendency was registered for total concentration in relation to initial concentrations (C0) [4]. Analyzing the fractional composition of individual metals in the subject soil during the experimental period, the limited phenomenon of fractional load percentage fluctuation was found: exchangeable (fraction I) and carbonate (fraction II) for most of the analyzed metals (Table 2). However, the registered changes for exchangeable fraction showed a long-term decreasing tendency, and for carbonate fraction the increasing percentage tendency. For instance, there were the following exchangeable fraction (fraction I) percentages for the examined elements before the experiment [%]: Cd – 4.8, Cr – 3.4, Cu – 3.9, Hg – 4.9, Ni – 3.1, Pb – 4.5, Zn – 1.7, and after four years of wastewaters agro-utilization they were respectively [%]: Cd – 0.6, Cr – 0.2, Cu – 0.5, Hg – 0.1, Ni – 0.2, Pb – 0.1, Zn – 0.2. In the case of carbonate fraction these changes were respectively: before the experiment [%]: Cd – 2.6, Cr – 1.7, Cu – 0.5, Hg – 1.7, Ni – 1.7, Pb – 2.7, Zn – 0.9, and after the experiment [%]: Cd – 7.3, Cr – 6.6, Cu – 2.5, Hg – 6.6, Ni – 4.9, Pb – 6.6, Zn – 4.9 (Table 2). It should be emphasized that the registered changes in the percentages of these fractional loads in both cases are connected with the values not exceeding 5 %. Analyzing the residual fractional composition, definitely the highest percentage was registered in the case of so called residual fraction, exceeding 70 % for each metal (fraction V, Table 2). In the case of a fraction related to hydrated Fe/Mn oxides, a stabile tendency to maintain a similar level of this fraction content for all examined metals in the experimental period was observed. The organic fraction (fraction IV) was characterized by periodical changes of its percentage with time. The obtained results of the analyses showed a significant speciation load stability in the case of metals which were in the examined soil within four years of agro-utilization of wastewaters from vegetable fat production. Changeability of the Speciation Forms of Heavy Metals in Soil... 1671

Table 2

Changeability of heavy metal fractional load percentages determined during the four-year experiment for soil where wastewaters from vegetable fats production were agro-utilized

% participation of fraction in subsequent measurements Heavy No. Fractions metal P0 P1 P2 P3 P4 P5 P6 P7 P8

1 Chromium I 3.4 2.0 2.8 1.4 1.8 1.4 1.1 0.3 0.2 (Cr) II 1.7 2.7 2.4 2.9 3.3 3.7 4.6 4.1 6.6 III 7.4 8.7 5.1 6.9 8.1 10.0 10.1 10.0 8.2 IV 12.0 12.1 14.0 14.0 10.1 13.2 11.2 11.5 12.3 V 75.5 74.5 75.7 72.8 76.7 71.7 73.0 74.1 72.7

2 Cadmium I 4.8 4.2 4.3 3.6 3.3 3.5 2.7 1.3 0.6 (Cd) II 2.6 3.2 4.4 4.7 4.1 4.7 5.7 6.9 7.3 III 6.7 9.6 9.5 8.2 11.9 10.6 9.1 8.4 8.9 IV 13.0 11.5 10.4 13.1 10.6 11.1 10.6 13.7 12.4 V 72.9 71.5 71.4 70.4 70.1 70.1 71.9 69.7 70.8

3 Nickel I 3.1 3.4 2.8 1.8 1.8 1.5 1.4 0.4 0.2 (Ni) II 1.7 2.2 2.7 3.4 3.3 3.7 4.4 4.1 4.9 III 11.0 9.2 9.1 10.8 12.9 10.5 9.8 11.9 13.2 IV 12.7 9.7 10.7 13.2 10.3 14.6 14.3 13.9 13.5 V 71.5 75.5 74.7 70.8 71.7 69.7 70.1 69.7 68.2

4 Lead I 4.5 4.5 3.5 3.6 2.6 1.6 1.7 0.9 0.1 (Pb) II 2.7 3.6 3.4 3.7 4.6 5.0 5.4 6.6 6.6 III 10.9 10.0 11.5 10.0 10.8 10.5 11.5 10.4 11.8 IV 8.0 8.1 8.8 10.8 10.2 11.7 10.8 11.4 11.0 V 73.9 73.8 72.8 71.9 71.8 71.2 70.6 70.7 70.5

5 Copper I 3.9 3.1 1.9 1.0 1.1 0.9 0.6 0.7 0.5 (Cu) II 0.5 0.8 0.7 0.7 0.9 0.9 1.5 1.5 2.5 III 8.9 10.7 15.8 15.9 13.5 17.2 13.5 13.5 12.9 IV 14.6 15.2 11.7 12.3 14.5 11.2 13.5 14.1 12.5 V 72.1 70.2 69.9 70.1 70.0 69.8 70.9 70.2 71.6

6 Mercury I 4.5 4.5 3.5 2.0 2.6 1.7 1.6 0.9 0.1 (Hg) II 1.7 3.4 3.7 3.9 4.7 5.4 5.0 6.6 6.6 III 9.9 10.0 9.5 10.0 9.8 9.5 8.7 10.4 9.8 IV 9.0 8.3 9.5 11.2 9.1 8.2 8.4 9.4 9.0 V 74.9 73.8 73.8 72.9 73.8 75.2 76.3 72.7 74.5

7 Zinc I 1.7 1.9 1.7 1.3 1.0 1.1 0.6 0.7 0.2 (Zn) II 0.9 2.2 2.3 2.9 2.9 3.2 3.3 3.2 4.9 III 12.4 14.3 14.2 14.6 13.9 12.6 9.8 12.3 10.5 IV 11.2 11.6 10.6 11.3 11.7 11.2 14.0 13.2 13.2 V 73.8 70.0 71.2 69.9 70.5 71.9 72.3 70.6 71.2

Where: P0 – zero sample (before agro-utilization of pretreated wastewaters) – total initial concentrations (C0) were respectively [mg/kg d.m.]: Cd – 0.19, Hg – 0.16, Cr – 5.29, Ni – 5.02, Cu – 8.11, Pb – 9.28 and Zn – 20.35; P1–P8 measurement series carried every six months within four years of analyses. 1672 Teresa Rauckyte-¯ak and S³awomir ¯ak

Conclusions

The obtained results of the analyses show a significant stability of metals speciation loads present in the subject soil within four years of carrying out agro-utilization of wastewaters from vegetable fat production. Therefore, it can be expected that in the case of further many-year irrigation and fertilization of the examined soil in the same process pretreatment conditions and agro-technical ones, no significant changes in individual shares of heavy metal speciation loads will be found.

References

[1] Rüffer H. and Rosenwinkel K.H.: Taschenbuch der industriebwasserreinigung. Oldenbourg Verlag, München–Wien 1991, pp. 115–134 (in German). [2] Rack M.: Ölsaatenaufbereitung, Speisefett- und Speiseölraffination – Hinweise und Erläuterungen zu Anhang 4 der Abwasserverordnung. BAnz. Beilage Nr. 3a/2007. Frankfurt 2007 (in German). [3] Regulation of the Minister of the Enviroment of 24 July 2006 on required quality standards for introducing sewage to water or soil and on extremely hazardous substances for the water environment (Polish Journal of Laws 2006, No. 137, item 984). [4] Rauckyte T., ¯ak S. and Pawlak Z.: Research on heavy metal contents in soils used for utilization of wastewaters from vegetable fats production. Ecol. Chem. Eng. A 2008, 15(4–5), 405–413. [5] Manz M., Weissflog L., Kühne R. and Schürmann G.: Ecotoxicological hazard and risk assessment of heavy metal contents in agricultural of central Germany. Ecotoxicol. Environ. Safety 1999, 42(2), 191–201. [6] Wac³awek W. and Moæko A.: Relationships between soil properties and speciation forms of heavy metals. Garden allotments in Opole and Strzelce Opolskie. Chem. In¿. Ekol. 2001, 8(2–3), 253–268. [7] Aleem A., Isar J. and Malik A.: Impact of long-term application of industrial wastewater on the emergence of resistance traits in Azotobacter chroococcum isolated from rhizospheric soil. Biores. Technol. 2003, 86(1), 7–13. [8] Rattan R.K., Datta S.P., Chhonkar P.K., Suribabu K. and Singh A.K.: Long-term impact of irrigation with sewage effluents on heavy metal content in soils, crops and groundwater – a case study. Agr. Ecosys. Environ. 2005, 109(3–4), 310–322. [9] Paredes C., Ceggara J., Roing A., Sánchez-Monedero M.A. and Bernal M.P.: Effect of waste waters from olive oil extraction plants on the bacterial population of soil. Chemosphere 1986, 15(5), 659–664. [10] Paredes M.J., Moreno E., Ramos-Cormenzana A. and Martinez J.: Characteristics of soil after pollution with waste waters from olive oil extraction plants. Chemosphere 1987, 16(7), 1557–1564. [11] Tardioli S., Bànnè E.T.G. and Santori F.: Species-specific selection on soil fungal population after olive mill waste-water treatment. Chemosphere 1997, 34(11), 2329–2336. [12] Aktas E.S., Imre S. and Ersoy L.: Characterization and lime treatment of olive mill wastewater. Water Res. 2001, 35(9), 2336–2340. [13] Alvarez-Bernal D., Contreras-Ramos S.M., Trujillo-Tapia N., Olalde-Portugal V., Frías-Hernández J.T. and Dendooven L.: Effects of tanneries wastewater on chemical and biological soil characteristics. Appl. Soil Ecol. 2006, 33(3), 269–277. [14] Tessier A., Campbell P.G.C. and Bisson M.: Sequential extraction procedure for the speciation of particulate trace metals. Anal. Chem. 1979, 51(7), 844–851. [15] Zimmerman A.J. and Weindorf D.C.: Heavy metal and trace metal analysis in soil by sequential extraction: a review of procedures. Int. J. Anal. Chem. vol. 2010, Article ID 387803, 7 pages, 2010. doi:10.1155/2010/387803. [16] Xiaoquan S. and Bin C.: Evaluation of sequential extraction for speciation of trace metals in model soil containing natural minerals and humic acid. Anal. Chem. 1993, 65(6), 802–807. [17] Hirner A.V.: Trace element speciation in soils and sediments using sequential chemical extraction methods. Int. J. Environ. Anal. Chem. 1992, 46(1–3), 77–85. [18] Ahumada I., Mendoza J., Navarrete E. and Ascar L.: Sequential extraction of heavy metals in soils irrigated with wastewater. Comm. Soil Sci. Plant Anal. 1999, 30(9–10), 1507–1519. Changeability of the Speciation Forms of Heavy Metals in Soil... 1673

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ZMIENNOŒÆ FORM SPECJACYJNYCH METALI CIʯKICH W GLEBIE PODDANEJ WIELOLETNIEMU NAWO¯ENIU ŒCIEKAMI Z PRODUKCJI T£USZCZÓW ROŒLINNYCH

Wydzia³ Technologii i In¿ynierii Chemicznej Uniwersytet Technologiczno-Przyrodniczy w Bydgoszczy

Abstrakt: Agroutylizacja podczyszczonych œcieków technologicznych z produkcji t³uszczów roœlinnych na cele nawadniaj¹ce oraz nawozowe jest m.in. limitowana stê¿eniem metali ciê¿kich w œciekach oraz glebie, na któr¹ kieruje siê odpady. W pracy przedstawiono wyniki czteroletnich badañ nad wp³ywem procesu nawadniania oraz nawo¿enia œciekami na stê¿enie oraz zmiennoœæ form specjacyjnych metali ciê¿kich (Cd, Cr, Cu, Hg, Ni, Pb, Zn) w przypowierzchniowej warstwie gleby. Sk³ad frakcyjny metali by³ oznaczany zgodnie z metodyk¹ Tessiera. Uzyskane wyniki wskazuj¹ na wystêpowanie istotnej stabilnoœci pól specjacyjnych metali oznaczanych w badanej glebie w okresie doœwiadczalnym. Analizuj¹c sk³ad frakcyjny, stwierdzono wystêpowanie nieznacznego trendu zmniejszania udzia³u procentowego frakcji wymywanej oraz wzrostu udzia³u frakcji wêglanowej w przypadku wiêkszoœci badanych metali ciê¿kich. S³owa kluczowe: metale ciê¿kie, formy specjacyjne metali, agroutylizacja œcieków z produkcji t³uszczów roœlinnych

ECOLOGICAL CHEMISTRY AND ENGINEERING A

Vol. 18, No. 12 2011

Beata GRYGIERZEC1

EFFECT OF NITROGEN FERTILIZATION ON SEED PRODUCTION OF Lolium perenne L. TURFGRASS CULTIVARS

NAWO¯ENIE AZOTEM W PRODUKCJI NASIENNEJ GAZONOWYCH ODMIAN Lolium perenne L.

Abstract: The paper contains the results of field and laboratory experiments on three Lolium perenne L. turfgrass cultivars (Gazon, Nira and Sandra) at Malopolska Plant Breeding Station (HBP) in Skrzeszowice near Krakow (220 m a.s.l.) conducted in the years 2006–2008. The analyses assessed the effect of doses of mineral fertilization with nitrogen (60, 90 and 120 kg N × ha–1) applied at early spring, the start of the earing stage and autumn on the seed yield. The highest yield was monitored by the inflorescence shoot density of cv. Nira (1895 shoots/m2), cv. Gazon (1976 shoots/m2) and cv. Sandra (2098 shoots/m2). Fertilization dose of 90 kg N × ha–1 applied three times effected the most considerably the number of spikelets per ear and the efficiency of seed settling on the ear: from 73 % (cv. Nira) to 76 % (cv. Sandra). The highest seed yield was obtained as a result of the 120 kg N × ha–1 fertilizations applied three times. Keywords: Lolium perenne, cultivars, fertilization, seed yield

Perennial ryegrass (Lolium perenne L.) belongs to the species which still raise interest of researchers, as evidenced by the most numerous collection of cultivars in Poland. It is also apparent as the ever growing market demand for this species. It has been estimated that the European Union’s demand for perennial ryegrass seeds is the greatest among grasses and on average reaches 60 000 Mg a year [1]. In Poland it occupies a prominent position in the seed production, accounting for ca 35–45 % of the whole seed crop acreage. According to Martyniak [2] and Svensson and Boelt [3] seed plantations of perennial ryegrasses may provide an alternative to cereal cultivation, the overproduction of which will make farmers seek new sources of income. It is commonly known that in order to improve the profitability of seed crop production it is necessary to increase seed yield and the main element determining the productivity the most is nitrogen fertilization. It turns out that optimisation of nitrogen fertilization of perennial ryegrass plantations involves not only determining the dose in the production

1 Department of Grassland, University of Agriculture in Krakow, al. A. Mickiewicza 21, 31–120 Kraków, Poland, phone: +48 12 662 43 61, fax: 48 12 663 44 43, email: [email protected] 1676 Beata Grygierzec cycle but also its proper division into parts and setting proper dates of application [4, 5]. Therefore, the aim of the Author’s investigations was an attempt at assessment of the nitrogen fertilization dose and its application date on the amount of the seed yield of three lawn cultivars of perennial ryegrass.

Materials and methods

The research was conducted in 2006–2008 at the Malopolska Plant Breeding Experimental Station – HPB in Skrzeszowice (220 a.s.l.) near Krakow, on degraded chernozem developed from loess.

The soil revealed the following chemical properties: pHKCl – 6.5; available P – 56, K – 132 and Mg – 39 g × kg–1; organic N – 1.5 and total carbon – 16.1 g × kg–1 of soil. The experiment was set up in the autumn 2005 using the randomised block method in four replications. The area of each plot was 10 m2 (1 × 10). The object of the investigation was a seed plantation of three fodder cultivars of perennial ryegrass (Lolium perenne): Gazon, Nira and Sandra. × –1 Phosphorus, dosed 30 kg P ha as triple superphosphate (46 % P2O4), was used for fertilization conducted once in spring. Potassium, in the amount of 66 kg K × ha–1 as high grade potassium salt (60 % K2O), was applied once, also in spring. Nitrogen fertilization in the doses of 60, 90 and 120 kg N × ha–1 was applied on the following dates: – once in the early spring; – twice in two equal portions (in the early spring and at the beginning of the earing stage); – three times in three equal parts (in the early spring, at the beginning of earing and in autumn). Chemical weed control on the seed plantation, with a dose of 1 dm3 × ha–1 of Aminopielik Gold, was conducted each year at the beginning of April (when the vegetation started) and in September. Before the seed harvesting single weeds were hand removed. In each year of the investigations, field observations and biometric plant measure- ments were conducted at the beginning of August. The number of plants was counted on individual plots on the area of 1 m2, as well as the number of spikelets per spike, and seed setting efficiency in spikes was assessed. The harvesting of the seed plantations was carried out in one stage by means of Wintersteiger plot combined harvester. The harvested seed material was dried in a storeroom and subsequently cleaned on the winnower. The cleaned grains were weighed, the obtained yield was calculated per 1 ha and a thousand grain weight was determined. The obtained results were verified statistically by means of the analysis of variance. The differences between means were estimated using the Student test at the significance level p = 0.05 and the correlation coefficient was calculated for selected features. The annual precipitation total for the period of investigations ranged from 463.8 to 683.4 mm, whereas the precipitation total for the six months (April–September) from Effect of Nitrogen Fertilization on Seed Production of Lolium perenne L. ... 1677

345.2–561.7 mm. Average annual temperature reached from 6.4 to 6.9 oC and between 11.9 and 12.8 oC during the vegetation season. The work presents the mean results for the 3 years of the investigations.

Results and discussion

The level of nitrogen fertilization is the factor affecting generative shoot formation on plantations of perennial ryegrass [6]. This thesis has been confirmed also by the results of the Author’s own investigations, where nitrogen fertilization significantly modified the number of generative shoots per area unit (Table 1). The highest shoot density in Gazon cv. – 2018 and Nira cv. – 1953 shoots/m2 was registered after a single application of nitrogen (in early spring) in the dose of 120 kg N × ha–1. On the other hand in Sandra cv. the largest number of generative shoots (2098 shoots/m2) was calculated on the object fertilized with 120 kg N × ha–1 applied on three dates: early in spring, at the beginning of the earing stage and in autumn.

Table 1

The mean number of generative stems of three Lolium perenne cultivars [units × m–2]

Specification cv. Gazon cv. Nira cv. Sandra Control 1 354 1 119 1 543

N60 – in one dose + P30K66 1 741 1 527 1 792

N60 – in two doses + P30K66 1 527 1 485 1 761

N90 – in one dose + P30K66 1 963 1 751 1 957

N90 – in two doses + P30K66 1 934 1 728 1 934

N90 – in three doses + P30K66 1 972 1 895 2 015

N120 – in one dose + P30K66 2 018 1 953 2 086

N120 – in two doses + P30K66 1 853 1 796 2 041

N120 – in three doses + P30K66 1 976 1 895 2 098 LSD (p = 0.05) 101.7 127.5 148.9

Perennial ryegrass is a nitrophilous species, visibly responding to nitrogen supplied with fertilizers by showing the changes of eg morphological properties determining the amount of seed yield. In the opinion of Young et al [7] and Golinski [6], nitrogen fertilization influences the number of kernels formed per spikelet in this species. Proper plant nutrition with nitrogen determines the effectiveness of their setting in the ear [8–11]. On the basis of the Author’s own research, a favourable effect of nitrogen fertilization on this feature value was observed only to the level of 90 kg N × ha–1 (Table 2). In conditions of nitrogen application in the dose of 120 kg N × ha–1 a worsening of kernel setting in ear efficiency was registered in each of the three tested cultivars. According to Marshall [12], the reason for a decline in the efficiency of kernel setting in the ear when larger nitrogen doses were applied was the competition for assimilates between the vegetative and generative shoots which follows the flowering stage and may affect a dieback of embryos and developing seeds. 1678 Beata Grygierzec Table 2 number of spikelets per spike cultivars effectiveness of embedding Lolium perenne of seeds in spikes number of spikelets per spike effectiveness of embedding of seeds in spikes number of spikelets per spike cv. Gazon cv. Nira cv. Sandra 527518764752781 2.32936 2.24923 2.45948 2.40 2.28 752 2.48 746 2.39 893 2.40 875 898 2.37 974 2.34 961 2.51 985 2.31 2.36 759 2.52 741 2.46 885 2.49 867 894 2.79 1 026 2.85 997 2.83 1 031 2.78 2.80 2.94 2.79 2.85 effectiveness of embedding of seeds in spikes 66 66 66 K 66 66 66 K K 66 66 30 K K K 30 30 K K 30 30 30 30 30 Effectiveness of embedding of seeds in spikes [%] and mean number of spikelets per spike of three Specification – in one dose + P – in two doses + P – in three doses + P – in one dose + P – in two doses + P – in one dose + P – in two doses + P – in three doses + P 60 60 90 90 90 120 120 120 ControlN 379 2.15 306 2.28 463 2.71 N N N N N N N LSD (p = 0.05) 23.9 0.16 21.8 0.17 27.9 0.17 Effect of Nitrogen Fertilization on Seed Production of Lolium perenne L. ... 1679 Table 3 mass of a thousand seeds yield cultivars Lolium perenne mass of a thousand seeds yield mass ] and mass of a thousand seeds [g] of three –1 ha of a thousand seeds × cv. Gazon cv. Nira cv. Sandra 527518764752781 2.32936 2.24923 2.45948 2.40 2.28 752 2.48 746 2.39 893 2.40 875 898 2.37 974 2.34 961 2.51 985 2.31 2.36 759 2.52 741 2.46 885 2.49 867 894 2.79 1 026 2.85 997 2.83 1 031 2.78 2.80 2.94 2.79 2.85 yield The mean yields [kg 66 66 66 K 66 66 66 K K 66 66 30 K K K 30 30 K K 30 30 30 30 30 Specification – in one dose + P – in two doses + P – in three doses + P – in one dose + P – in two doses + P – in one dose + P – in two doses + P – in three doses + P 60 60 90 90 90 120 120 120 ControlN 379 2.15 306 2.28 463 2.71 N N N N N N N LSD (p = 0.05) 23.9 0.16 21.8 0.17 27.9 0.17 1680 Beata Grygierzec

Young et al (1996) state that there is an optimal nitrogen dose for each cultivar which allows for obtaining a maximum seed yield; these doses are diversified and may range from 60 to 180 kg N × ha–1. In the conducted experiments, fertilization with each dose of nitrogen significantly diversified seed yield (Table 3). The highest seed yield from each cultivar was obtained as a result of 120 kg N × ha–1 applied in the early spring term, at the beginning of earing and in autumn. About thrice higher yields, ranging from 985 (Nira cv.) to 1031 kg × ha–1 (Sandra cv.) were produced on the unfertilized objects. An increase in the total nitrogen dose was usually accompanied by an increase in one-thousand-seed weight. However, these tendencies were not stabilized. A crucial importance of early spring fertilization was noted in the assessment of the seeding value of the seeds by means of one-thousand-seed weight. A single nitrogen dose applied in early spring increased one-thousand-seed weight in comparison with the control: in Gazon cv. from 7 % (60 kg N × ha–1) to 15 % (120 kg N × ha–1), in Nira cv. from 3 % (60 kg N × ha–1) to 10 % (90 and 120 kg N × ha–1) and in Sandra cv. between 2 % (60 kg N × ha–1) and 8 % (120 kg N × ha–1).

Conclusions

1. The highest seed yield of L. perenne cvs per 1 m2 was obtained at the following flower shoot densities: 1895 in Nira cv., 1976 in Gazon cv. and 2098 in Sandra cv. at dose of the 120 kg N × ha–1 fertilizations applied three times. 2. Nitrogen fertilization dose of 90 kg N × ha–1, applied three times (at the early spring, the beginning of earing and in autumn) increased the number of spikelets per ear and seed setting efficiency in ear to 73 % (Nira cv.) and to 76 % (Sandra cv.). 3. The increment of seed setting efficiency as a result of nitrogen fertilization at the use of 90 kg N × ha–1, which decrease after treatment. 4. The highest seed yield was obtained under the influence of the 120 kg × ha–1 nitrogen dose. 5. The crucial importance of a single fertilization dose applied in early spring was registered in the assessment of the sowing value of seeds by means of one-thousand- -seed weight.

References

[1] Kley G.: Seed production in grass and clover species in Europe: Proc. 3rd Int. Herb. Seed Conf., Halle 1995, 12–22. [2] Martyniak J.: Konkurencyjnoœæ upraw traw na nasiona przy aktualnie stosowanej technologii, Mat. Konf. Nauk. „Ekonomika technologii produkcji roœlinnej”, Bonin 1998, 103–110. [3] Svensson K. and Boelt B.: Lolium perenne L. (Perennial Ryegrass) in Denmark, [in:] Frage seed production. vol. 1: Temperate species, D.T. Fairey and J.G. Hampton (eds.), CABI, Wallingford 1997, 321–328. [4] Goliñski P.: Mo¿liwoœci zwiêkszenia wydajnoœci plantacji nasiennych Lolium perenne. £¹karstwo w Polsce 2002, 5, 65–74. [5] Rowarth J.S.: Nutrients and moisture inputs for grass seed field. J. Appl. Seed Prod. 1997, 15, 103–110. [6] Goliñski P.: Efektywnoœæ nawo¿enia azotem w produkcji nasion Lolium perenne L. Rocz. Akad. Roln. Poznañ 2001, 321, 15–89. Effect of Nitrogen Fertilization on Seed Production of Lolium perenne L. ... 1681

[7] Young W.C.III, Youngberg H.W. and Chilcote D.O.: Spring nitrogen rate and timing influence on seed yield components of perennial ryegrass. Agron. J. 1996, 88, 947–951. [8] Hebblethwaite P.D., Wright D. and Noble A.: Some physiological aspects of seed yield in Lolium perenne L., [in:] Seed production, (ed.) P.D. Hebblethwaite, Butterwotths, London 1980, 71–90.. [9] Szczepanek M., Skinder Z., Wilczewski E. and Borys W.: Ksztaltowanie i wspólzaleznosc cech odmiany trawnikowej zycicy trwalej w warunkach zróznicowanego poziomu nawozenia azotem w czteroletnim okresie uzytkowania na nasiona. Acta Sci. Polon., Agricultura 2007, 6(4), 65–72. [10] Szczepanek M.: Trwa³oœæ Lolium perenne L. uprawianej na nasiona w zale¿noœci od sposobu siewu i rozstawy rzêdów. Acta Sci. Polon, Agricultura, 2005, 4(2), 101–112. [11] Szczepanek M.: Stability of perennial ryegrass (Lolium perenne L.) plants cultivated for seeds at varied levels of nitrogen fertilization. EJPAU, 2006, 9(4), #56, www.ejpau.media.pl/volume9/issue4/art-56.html. [12] Marshall C.: Developmental and physiological aspects of seed production in herbage grasses. J. Appl. Seed Prod. 1985, 3, 43–49.

NAWO¯ENIE AZOTEM W PRODUKCJI NASIENNEJ GAZONOWYCH ODMIAN Lolium perenne L.

Zak³ad £¹karstwa Uniwersytet Rolniczy im. Hugona Ko³³¹taja w Krakowie

Abstrakt: Praca zawiera zestawienie wyników badañ polowych i laboratoryjnych przeprowadzonych w Stacji Ma³opolskiej Hodowli Roœlin – HBP w Skrzeszowicach ko³o Krakowa (220 m n.p.m.) w latach 2006–2008 z trzema gazonowymi odmianami ¿ycicy trwa³ej (Gazon, Nira, Sandra). W badaniach poddano ocenie wielkoœæ dawki nawo¿enia mineralnego azotem (60, 90, 120 kg N × ha–1) oraz termin jego stosowania (wczesnowiosenny, pocz¹tek k³oszenia i jesienny), na wielkoœæ plonu nasion. W przeliczeniu na 1 m2 uzyskanie najwiêkszego plonu nasion zape³nia³a obsada pêdów kwiatowych: 1895 u odmiany Nira, 1976 u odmiany Gazon oraz 2098 u odmiany Sandra. Nawo¿enie azotem w dawce 90 kg N × ha–1 zastosowanej w trzech terminach: wczesnowiosennym, na pocz¹tku k³oszenia i jesiennym najbardziej zwiêksza³o liczbê k³osków w k³osie oraz efektywnoœæ osadzania nasion w k³osie od 73 % u odmiany Nira do 76 % u odmiany Sandra. Najwiêkszy plon nasion uzyskano pod wp³ywem dawki 120 kg × ha–1, zastosowanej w trzech terminach: wczesnowiosennym, na pocz¹tku k³oszenia i jesiennym. S³owa kluczowe: Lolium perenne, odmiany, nawo¿enie, plon nasion

ECOLOGICAL CHEMISTRY AND ENGINEERING A

Vol. 18, No. 12 2011

Joanna KOSTECKA1 and Grzegorz P¥CZKA

KITCHEN WASTES AS A SOURCE OF NITROGEN AND OTHER MACROELEMENTS ACCORDING TO TECHNOLOGY OF VERMICULTURE

ODPADY KUCHENNE JAKO RÓD£O AZOTU I INNYCH MAKROPIERWIASTKÓW ZALE¯NIE OD TECHNOLOGII PROWADZENIA WERMIKULTURY

Abstract: The proper utilization of organic wastes has become an important environmental problem (in relation to economic and social aspect of sustainable development). In case of selected organic kitchen wastes, the process of vermicomposting on site where they are produced (in innovative “earthworm ecological box”) could be a successful method of their neutralization. The aim of this work was to specify the fertilizing qualities of vermicompost from kitchen wastes depending on the technology of its production (vermicomposting took place in “the earthworm ecological boxes” by means of a condensed substrate and a population of earthworms technology (Z) or their dividing every 4 weeks (R)). The qualities of the vermicomposts produced this way were determined in a dynamic system (three times). pH was specified potentiometrically, the concentration of salt – by means of the conductometric method, the N-NO3 content in the extract of 0.03 moles of acetic acid – potentiometrically, selected macroelements: assimilable P, K, Mg and calcium Ca were determined in the extract of 0.03 moles of acetic acid CH3COOH. It has been proven that the content of obtained products depended on the way of treatment of the boxes. A significantly bigger growth of nitrate nitrogen content N-NO3, assimilable phosphorus, potassium and magnesium was the result of the application of the (Z) technology – the intense vermicomposting without dividing the substrates and earthworm population. Vermicomposting organic kitchen wastes by a more frequent dividing of the substrates and earthworm population – had a more favourable influence on the reduction of the average concentration of salt in the vermicompost. Keywords: organic kitchen waste, vermicompost, macroelements, pH, salinity

The organic wastes produced by a vast array of business sectors related to agriculture have become an increasingly significant problem. Their proper utilization has become of utmost importance not only from the environmental point of view (the replenishment

1 Chair of Natural Theories of Agriculture and Environmental Education, Faculty of Agriculture and Biology, University of Rzeszow, ul. M. Æwikliñskiej 2, 35–601 Rzeszów, Poland, phone: +48 17 872 17 33, fax: +48 17 872 17 96, email: [email protected] 1684 Joanna Kostecka and Grzegorz P¹czka of the organic substances in soil, the threat of air, soil and water degradation) [1], but also in relation to the continuously growing prices of land located next to urban and rural areas [2]. Depending on the features of such wastes, the method of their neutralization could be the process of fermentation or composting and vermi- composting. In case of composting, the main problem is the relative length of this process and the necessity to aerate the material, while the advantage could be the high temperature of the process (about 70 oC), which causes the death of all the pathogens in the composted material [3, 4]. Because of the above-mentioned factors, a considerable interest has been aroused by the integration of the process of composting and vermicomposting, which, on the one hand, allows to dispense with heavy equipment used to aerate heaps [2], and, on the other, prevents the created products from the lack of hygienizing temperature. The technology of vermicomposting facilitates a more rapid transformation especially of nitrogen, potassium, phosphorus and calcium compounds into forms, which are more easily dissolved and more available for plants. This is connected with the fact that each day an earthworm absorbs a large amount of organic material from soil, yet only 5–10 % of the digested material is assimilated by its organism, the remainder is excreted in the form of coprolites rich in nitrogen, phosphorus, potassium and microorganisms which are beneficial for soil [5–8]. Nitrogen accounts for only 0.5–4 % of dry mass in plants, yet it plays a critical part in their growth as a constituent of proteins as well as other important compounds such as nucleic acids, phytohormones, and nucleotides. Mineral nitrogen in the form of N-NH4 and N-NO3 is most readily available for plants of higher order, ie those not in symbiotic relationships with microorganisms binding elemental nitrogen [9]. Nitrogen is one of the chemical elements controlling the growth of organisms, both plants and animals, in accordance with Liebig’s Law of the Minimum. Shortage of this element severely hinders the growth and normal development in plants. The most significant symptoms of nitrogen deficiency include: leaves falling off early, deteriorating over and underground organs, and irregular shape of fruit [10, 11]. Of great importance is the interaction between nitrogen and potassium. Even with the most favourable contents of nitrogen in forms which are readily available to plants, a deficiency of potassium in soil is a disturbing factor for the transformation of mineral nitrogen into proteins. N/K imbalance leads to increased content of nitrates in plants which adversely impacts their nutritional value. Adequate content of potassium in plants improves their response to nitrogen fertilizers, and similarly potassium fertilization is more effective in plants adequately supplied with nitrogen. On the other hand, overfertilization with potassium causes magnesium deficiency because K is antagonistic in relation to Mg and hinders both its assimilation and its movement in the plant [10]. As a result of the long-lasting exploitation, the soils of Europe contain at present from 2 to 3.5 % of organic substances [12]. As a consequence, the European Strategy for Soil Protection [13] contains the conclusion that the exploitation of land in regions, where the content of organic substances in soil is lower than 2 %, should be necessarily connected with actions aimed at soil stabilization or absolutely indispensably connected with the gradual increase of nitrogen-rich organic substances in soil. The search for nitrogen sources in wastes of various origins is well justified. Kitchen Wastes as a Source of Nitrogen and Other Macroelements... 1685

A citizen with a proper awareness of this issue can segregate organic kitchen wastes on site where they are produced in order to obtain fertilizing material of high quality, with no pollution. The aim of this work was to specify the fertilizing qualities of vermicompost from kitchen wastes depending on the technology of its production in the ecological boxes.

Materials and methods

The research was conducted in the laboratory conditions (temp. 20 ± 5 oC) of Department of Natural Theories of Agriculture and Environmental Education at the Faculty of Agriculture and Biology of the University of Rzeszow, over the period of 4 months. In order to improve the conditions of vermicomposting, kitchen wastes were mixed with cellulose in the ratio of 4:1 [14] and the vermicomposting took place in “the earthworm ecological boxes” by means of the technology of a condensed substrate and a population of earthworms (Z) and their dividing every 4 weeks (R) (each type in 3 repetitions) (Table 1).

Table 1

Arrangement of the experiment

Containers 1–3 (type (Z)): the earthworms stay there over the period of 4 months without dividing their population Containers Containers 4–6 (type (R)): the substrates and the earthworms are divided every 4 weeks and put into other containers, which simulates the frequent division of breeding beds In each case 1 dm3 of gardening soil with a specific chemical composi- Initial substrate tion* In each case 50 specimens of mature earthworms E. fetida of known Earthworms biomass (with the density of 25 specimens × dm–3 and 0.541 ± 0.006 g × dm–3 ) Bread and pasta leftovers, apple and potato peelings + cellulose (in the Wastes ratio of 4:1) In each case 0.2 dm3 of the above-mentioned wastes, each time in the Feeding / utilization in the process same way, into each container, during the analysis of the conditions of of vermicomposting the earthworm population performed every four weeks (in the same way in all the boxes) Watering In each case the same amount of tap water** –3 –3 –3 –3 *pHinH2O – 6.2, salinity NaCl – 0.5 g × dm , N-NO3 – 180 mg × dm ,P–63mg× dm , K – 186 mg × dm , Ca – 1027 mg × dm–3, Mg – 141 mg × dm–3; ** pH – 7.6, conductivity – 542 mS × cm–1, nitrates(V) – –3 –3 –3 8.9 mg × dm , Mg – 15.7 mg × dm , hardness – 257 mg CaCO3 × dm .

The qualities of the vermicomposts produced this way were determined in a dynamic system (three times), while: pH in the water was specified by means of the potentiometric method, the concentration of salt – by means of the conductometric method, the N-NO3 content in the extract of 0.03 moles of acetic acid – potentio- metrically, selected macroelements: assimilable P, K, Mg and calcium Ca were 1686 Joanna Kostecka and Grzegorz P¹czka

determined in the extract of 0.03 moles of acetic acid CH3COOH using the Spurway method, as modified by Nowosielski [15]. The results were analysed in the Statistica PL program, with the application of two-factor analysis of variance (the time and technology of vermicomposting).

Results and discussion

In the conducted experiment it has been proven that kitchen wastes are a valuable source of nutrients for plants. The levels of macroelements in the form available for plants in the vermicomposting products under research exceeded the average nutritional requirements of most garden plants [16]. The composition of the obtained vermicomposts depended on the technology applied to the compost box (Table 2). The statistical analysis showed differences in the average content of macroelements (except for calcium Ca) present in the substrates created by vermicomposting (over the period of 16 weeks) the 1 dm3 of the initial substrate and 4 dm3 of organic wastes. The content of nitrate nitrogen (N-NO3) depended on the way of treating the boxes. In the containers (Z) (where earthworms with the initial density of 25 specimens × dm–3 and the mass 0.541 ± 0.006 g × dm–3 stayed without dividing their population over the period of 4 months) the level of the element was determined to be over ten times higher than the nutritional requirements of plants. In the containers (R) (where similar substrates and earthworm populations were divided every 4 weeks and put into other containers, which simulated the frequent separation of breeding beds) the content exceeded the requirements only over five times (p < 0.001). The changes in the content of assimilable phosphorus P, potassium K and magnesium Mg were similar. While in the non-separated containers (Z) the potassium content should be diluted depending on the requirements of the garden plants, from eight to four times (in case of phosphorus from ten to five times and in case of magnesium from six to three times, respectively), the content of all these elements in the divided containers (R) was significantly lower; however, in this case it was also too high (Table 2) (p < 0.001). The content of calcium Ca was different from the elements described so far. The calcium content was close to the nutritional requirements of garden plants and remained unchanged regardless of the different technologies applied to the boxes.

After four months’ period of utilizing kitchen wastes, pHH2O of the produced vermicomposts decreased and was included in the range from 4.90 to 5.58. It was slightly lower than the one obtained by Kostecka [14] (6.0–8.6) and Garczynska [17] (5.9–6.1) while vermicomposting the same wastes. Under the influence of vermicomposting in the technology of dividing (R) the obtained salinity of vermicompost was significantly lower – 3.79 ± 0.63 g NaCl × dm–3. In case of the divided containers this parameter fluctuated more slowly during the first 16 weeks of the research. Vermicomposting organic kitchen waste in the containers with a condensed earthworm population triggered an immediate increase in salinity to 5.5 ± 0.21 g NaCl × dm–3 (by 45 %) (p < 0.001) (Table 2). The issue of the significantly Kitchen Wastes as a Source of Nitrogen and Other Macroelements... 1687 8 ± b Table 2 2 ± 3 179 ± assimilable a 190 349 125 187 ± 7 177 ± 9 195 207 ± 11 181 ± 8 ± ± ± a 18 141 ± Ca Mg 91 1990 ± 60 1932 ± a 190 2098 ; 2 – vermicomposting over the period of 12 weeks; 3 – ± ] b –3 dm × K [mg 106 605 ± 41 1674 ± 56 2061 162 682 ± 206 1796 ± 8.5 649 assimilable ± ± a 7 731 34 1359 17 877 ± ± ± b 2 186 1027 ± P ; Z – research containers – technology without dividing the substrate; R – research containers 12 120 21 159 assimilable ± ± a 90 170 ± 10 145 142 315 107 202 ± ± ± b : 1 – measurement after 8 weeks of vermicomposting 3 163 ± N-NO 58 659 50 573 117 757 ± ± a ± 0.63 1217 0.27 688 0.55 962 ± ± ± b ] –3 dm × [g 0.12 3.48 0.62 4.32 of salt NaCl 0.21 3.79 ± ± ± The concentration a 5.5 4.09 5.34 min min min max max max O 2 5.19 5.05 4.90 5.48 5.31 5.58 H pH min min min max max max 6.0–7.5 about 1.0 50–120 40–80 125–250 1000–2000 60–120 6.16–6.19 0.5 18 RRZRZRZRZ R ZR ZRZR The qualities of vermicompost obtained as a result of different technologies of the vermicomposting process in a dynamic system 5.11 4.96 5.44 5.32 5.27 5.47 1 2 3 ment Measure- Initial substrate The optimal level for plants* * According to Konczak-Konarkowska [16]; a, b – statistically significant differences (p < 0.05) – technology of dividingvermicomposting the over substrate the and period the of population 16 of weeks. earthworms 1688 Joanna Kostecka and Grzegorz P¹czka increased salinity of the vermicomposts from household waste has already been highlighted in previous publications [18,19], pointing out the limitations concerning their use. The discussed problem has to be considered important because at present non- -segregated organic wastes fill about 30–50 % of the volume of our dumps. However, they could become a material to produce composts and vermicomposts – important from the point of view of replenishing organic substances in the soils. Vermicomposts produced in both technologies were not suitable to be used directly as substrates for plant growth because most of the researched qualities highlighted the necessity to dilute them. The positive influence of dilution can also solve the problem of excessive concentration of salt in the analysed substrate.

Conclusions

1. Organic kitchen wastes can be vermicomposted in ecological boxes on site where they were produced. 2. It has been proven that the content of obtained products depended on the way of treatment of the boxes. From the point of view of the content of the basic nutrients for plants, a significantly bigger growth of nitrate nitrogen content N-NO3, assimilable phosphorus, potassium and magnesium was the result of the application of the (Z) technology – the intense vermicomposting without dividing the substrates and earth- worm population. 3. Technology (R) – vermicomposting organic kitchen wastes by a more frequent dividing of the substrates and earthworm population – had a more favourable influence on the reduction of the average concentration of salt in the vermicompost. 4. When analysing the obtained results from the point of view of the growing need for natural fertilizers and the necessity for pro-environmental utilization of huge amounts of organic wastes without placing them in dumping sites, another advantage of (R) technology (with the more frequent dividing of the substrates and the population of earthworms) is the fact that over the period of 16 weeks three times more organic kitchen wastes were neutralised using this method.

References

[1] DeLuca T.H. and DeLuca D.K.: J. Prod. Agric. 1997, 10, 235–241. [2] Ndegwa P.M. and Thompson S.A.: Bioresour. Technol. 2001, 76, 107–112. [3] Hay C.J.: BioCycle 1996, 6, 67–76. [4] Papadimitriou E.K. and Balis C.: Compost. Sci. Utilization 1996, 4, 52–61. [5] Winding A., Ronn R. and Hendriksen N.B.: Biol. Fertility Soils 1997, 24, 133–140. [6] Atiyeh R.M., Lee Edward C.A., Arancon N.Q. and Metzger J.D.: Bioresour. Technol. 2002, 84, 7–14. [7] Chaoui H.I., Zibilske L.M. and Ohno T.: Soil Biol. Biochem. 2003, 35, 295–302. [8] Capowiez Y., Cadoux S., Bouchand P., Roger-Estrade J., Richard G. and Boizard H.: Soil Biol. Biochem. 2009, 41, 711–717. [9] Bloom A.J., Meyerhoff P.A., Taylor A.R. and Rost T.L.: J. Plant Growth Regul. 2002, 21, 416–431. [10] Kopcewicz J. and Lewak S.: Podstawy fizjologii roœlin, PWN, Warszawa 2002. [11] Kowalska I.: Dzia³kowiec 2002, 7, 50–51. [12] Goœciñski J.: Przegl¹d Komunalny 2007, 1, 58–60. Kitchen Wastes as a Source of Nitrogen and Other Macroelements... 1689

[13] European Strategy for Soil Protection IP/06/1241 22.09.2006r (C282E/139) COM(2006)231. [14] Kostecka J.: Zesz. Nauk. AR w Krakowie, Ser. Rozprawy, 2000, pp. 88. [15] Nowosielski O.: Zasady opracowywania zaleceñ nawozowych w ogrodnictwie. PWRiL, Warszawa 1988. [16] Koñczak-Konarkowska B.: Podstawy zaleceñ nawozowych w ogrodnictwie, Podrêcznik dla pracowni ogrodniczych stacji chemiczno-rolniczych. KSCHM w Warszawie, OSCHR w Gorzowie Wielko- polskim, Gorzów Wielkopolski 2009. [17] Garczyñska M.: Wp³yw wybranych preparatów na populacje d¿d¿ownic Eisenia fetida fetida Sav. w skrzynkach ekologicznych, PhD thesis, University of Rzeszow, Rzeszów 2010. [18] Kiepas-Kokot A. and Szczech M.: Roczn. Akad. Roln. w Poznaniu 1998, 27, 137–143. [19] Kostecka J., Kaniuczak J. and Nowak M.: Folia Univ. Agric. Stet. 1999, 200(77), 173–178.

ODPADY KUCHENNE JAKO RÓD£O AZOTU I INNYCH MAKROPIERWIASTKÓW ZALE¯NIE OD TECHNOLOGII PROWADZENIA WERMIKULTURY

Zak³ad Biologicznych Podstaw Rolnictwa i Edukacji Œrodowiskowej Uniwersytet Rzeszowski

Abstrakt: Prawid³owe unieszkodliwianie odpadów organicznych sta³o siê wa¿nym problemem ekologicznym (w powi¹zaniu z gospodarczym i spo³ecznym aspektem zrównowa¿onego rozwoju). W odniesieniu do selekcjonowanych organicznych odpadów kuchennych mo¿liwe jest ich wermikompostowanie na miejscu powstawania (w innowacyjnych „d¿d¿ownicowych skrzynkach ekologicznych”). Celem prezentowanej pracy by³o okreœlenie w³aœciwoœci nawozowych wermikompostu z odpadów kuchennych zale¿nie od technologii jego produkcji (wermikompostowanie prowadzono w „d¿d¿ownicowych skrzynkach ekologicznych” w technologii zagêszczonego pod³o¿a i populacji d¿d¿ownic (Z) albo ich rozdzielania co 4 tygodnie (R)). Cechy tak produkowanych wermikompostów okreœlono w uk³adzie dynamicznym przy czym: pH okreœlano metod¹ potencjometryczn¹, stê¿enie soli metod¹ konduktometryczn¹, zawartoœæ N-NO3 w wyci¹gu 0,03 mol kwasu octowego potencjometrycznie, wybrane makroelementy – przyswajalny P, K, Mg i Ca, oznaczano w wyci¹gu 0,03 mol kwasu octowego CH3COOH. W prowadzonym doœwiadczeniu uzyskano wermikomposty o sk³adzie zale¿nym od technologii wermi- kompostowania. Znacznie wiêkszy wzrost zawartoœci azotu azotanowego N-NO3, przyswajalnego fosforu, potasu, magnezu oraz wapnia zapewnia³a technologia intensywnego wermikompostowania bez podzia³u pod³o¿a i populacji d¿d¿ownic. Technologia wermikompostowania kuchennych odpadów organicznych przez czêste rozdzielanie pod³o¿a i populacji d¿d¿ownic by³a korzystniejsza dla obni¿enia œredniego zasolenia wermikompostu. S³owa kluczowe: organiczne odpady kuchenne, wermikompost, makropierwiastki, pH, zasolenie

ECOLOGICAL CHEMISTRY AND ENGINEERING A

Vol. 18, No. 12 2011

Teresa RAUCKYTE-¯AK1 and Bo¿ena SZEJNIUK2

INFLUENCE OF 1,3,4-THIADIAZOLE DERIVATIVES ON THE BIOLOGICAL ACTIVITY OF THE SELECTED ENVIRONMENTAL BACTERIA

WP£YW POCHODNYCH 1,3,4-TIADIAZOLI NA AKTYWNOŒÆ WYBRANYCH BAKTERII ŒRODOWISKOWYCH

Abstract: The paper presents the results of microbiological research carried out with the living micro- organisms participation and in the domestic sewage and sludge from municipal sewage treatment plants environment. The influence of the selected heterocyclic compounds – the derivatives of 1,3,4-thiadiazole on susceptibility of aerobic Gram-positive and Gram-negative bacteria was investigated. For the research of biological activity the following environmental bacterial strains were selected: Escherichia coli, Staphylo- coccus aureus, Bacillus subtilis, Bacillus cereus, Proteus mirabilis, Enterococcus faecium, Salmonella senftenberg, and Salmonella typhimurium. For the group of the tested bacteria, 2-amino-1,3,4-thiadiazole turned out to be the most active chemical compound. It was found that evenly matched concentrations of this compound allow for the selective elimination of such microorganisms as: Salmonella senftenberg, Bacillus cereus, Staphylococcus aureus and Enterococcus faecium. In the case of Escherichia coli, the minimum inhibitory concentration (MIC) was detected at the level of 12.5 mg/cm3 (determined in relation to 2-acetylamino-1,3,4-thiadiazole-5-sulfonamide, bis(2-acetylamino-1,3,4-thiadiazole)-5,5’-disulfonamide), and for 2-amino-1,3,4-thiadiazole: 0.5 mg/cm3. Similar results were observed in case of Enterococcus faecium bacterium: MIC for 2-amino-1,3,4-thiadiazole was less than 0.25 mg/cm3, and MIC for bis(2-acetylamino- -1,3,4-thiadiazole)-5,5’-disulfonamide this level was equal to 25.0 mg/cm3. Keywords: 1,3,4-thiadiazole derivatives, microbiological research, sanitary status indicator

The interest in derivatives of 1,3,4-thiadiazole has increased since phenylhydrazine and hydrazine were found. Thus, the research widened our knowledge concerning the preparation and chemical properties of 1,3,4-thiadiazoles. Derivatives of this compound showing biological activity are known as medical preparations [1]. A lot of research is

1 Faculty of Chemical Technology and Engineering, University of Technology and Life Sciences, ul. Seminaryjna 3, 85–326 Bydgoszcz, Poland, phone: +48 52 374 90 63, fax: +48 52 374 90 05, email: [email protected]

2 Faculty of Animal Breeding and Biology, University of Technology and Life Sciences, ul. Mazowiecka 28, 85–085 Bydgoszcz, Poland, phone: +48 52 374 97 95, fax: +48 52 322 81 58, email: [email protected] 1692 Teresa Rauckyte-¯ak and Bo¿ena Szejniuk being carried out concerning the issue of the application of heterocyclic derivatives in viral and bacterial diseases treatment [2–4]. It is known that 1,3,4-thiadiazole derivat- ives possess such biological activity as: antibacterial [5, 6] antimycobacterial [7, 8], antidepressive [9] and cardiotonic [10]. Resent investigation has also detected the analgesic [11] and anti-inflammatory [12–14] activity for these heterocyclic com- pounds. The compositions containing this heterocyclic ring form reactive centres in many enzymes and coenzymes, and are included in the composition of structures such as nucleic acids, chlorophyll, haemin – red blood pigment, pesticides, medicines (such as: 4-dimethylamino-2,3-dimethyl-1-phenyl-3-pyrazolin-5-one, penicillin G, diazepam, resirpin, acridine), hormones (serotonin, melatonin), amino acids (eg: tryptophan, histidine) and many others. Other 1,3,4-thiadiazole derivatives are practically obtained by the application of different methods and their preparation technologies are strictly related to water and the pollution of its derivative compounds as well as to sewage and solid waste formation. In the pharmaceutic industry, diuramid is mostly prepared from 2-acetylamine-5-mercapto-1,3,4-thiadiazole by the process of chlorination involving gaseous chlorine and the ammonolysis of the semi-product [15]. The compounds such as: 2-acetylamino-1,3,4-thiadiazole-5-sulfonamide (known under the trade name of diuramid, acetazolamide, diamox, diacarb, diluran), 2-amino-1,3,4-thiadiazole and bis(2-acetylamino-1,3,4-thiadiazole)-5,5’-disulfonamide are found in the wastes and wastewaters from different diuramid production technologies [16]. These compounds also occur in nature and show high biological activity [17, 18]. The aim of the work is investigating the influence of the 1,3,4-thiadiazole derivatives on the activity of the selected environmental bacterial strains: Escherichia coli, Staphylococcus aureus, Bacillus subtilis, Bacillus cereus, Proteus mirabilis, Entero- coccus faecium, Salmonella senftenberg, and Salmonella typhimurium.

Experimental part

The following heterocyclic compounds were applied in the biological research: 2-acetylamino-1,3,4-thiadiazole-5-sulfonamide (ATS), 2-amino-1,3,4-thiadiazole (AT) and bis(2-acetylamino-1,3,4-thiadiazole)-5,5’-disulfonamide (BATD). They were prepar- ed with the use of the procedures described below. Reagents of the analytically pure class produced by Sigma-Aldrich and POCh Gliwice were used for syntheses.

Preparing 2-acetylamino-1,3,4-thiadiazole-5-sulfonamide (ATS)

For the purpose of the biological study, concerning the subject of our research, we prepared acetazolamide with the use of hydrogen peroxide – hydrochloric acid. The synthesis was carried out as follows: First, 50.0 cm3 (1.62 mole) of concentrated 36.0 % hydrochloric acid (d = 1.18 g/cm3) was introduced into the reactor equipped with the reflux condenser, the mixer and the thermometer. Then, holding temperature within the range of: 5 to 10 oC, 80 cm3 (2.64 mole) of 30.0 % hydrogen peroxide (d = 1.11 g/cm3) was added in small doses. After introducing the whole amount of the oxidizer, 5.0 g (0.029 mole) of 2-acetylamino-5-mercapto-1,3,4-thiadiazole was added in doses. The Influence of 1,3,4-Thiadiazole Derivatives on the Biological Activity... 1693 last stage included the introduction of 99.5 % (d = 1.05 g/cm3) acetic acid in the total volume of 50.0 cm3 (0.88 mole). All the components were stirred for 120 min in temperature ranging from 5 to 10 oC. The obtained precipitate of 2-acetylamino-5-sulfo- chloro-1,3,4-thiadiazole (7.7 g in the form of the moist substance) was filtered and washed with ice-cold distilled water and submitted to ammonolysis by applying 9-time surplus of 25 % aqueous ammonia (d = 0.90 g/cm3). Later, 2-acetylamino-5-sulfo- chloro-1,3,4-thiadiazole was introduced into 68.6 cm3 of ammonia with the temperature not exceeding 20 oC. Then, the reaction mixture was heated for 120 minutes at 50 oCin order to remove the excess of ammonia. After that, charcoal was added and heated for 10 minutes. The hot mixture was filtered and cooled down to 2–3 oC. Later it underwent acidification with 36.0 % hydrochloric acid up to pH = 1. The obtained acetazolamide was separated and recrystallised from hot, distilled water. 2.05 g (yield: 32.0 %) of 2-acetylamino-5-sulfonamido-1,3,4-thiadiazole at melting point (m.p.) 257 oC (lit. m.p. 258–259 oC [15]) was prepared.

Preparing 2-amino-1,3,4-thiadiazole (AT)

The next compound: 2-amino-1,3,4-thiadiazole (AT) was prepared from thiosemi- carbazide and ethyl orthoformate using the method described in [19]. A mixture containing 4.5 g (0.05 mole) of thiosemicarbazide and 8.0 cm3 (0.05 mole) of ethyl orthoformate was introduced into the reactor equipped with a reflux condenser, a mixer and a thermometer. Then the mixture was heated on a water bath for 6 hours. Then, 250 cm3 (4.3 mole) of boiling ethyl alcohol was added. The hot mixture was filtered in order to remove N,N’-bis-(1,3,4-thiadiazole-2) formamide. The filtrate was concentrated in a vacuum evaporator until approx. 50 cm3 and then cooled. As much as 2.31 g (yield: 32.6 %) of 2-amino-1,3,4-thiadiazole was obtained with the melting point ranging from 190 to 191 oC (lit. m.p. 191 oC [19]).

Preparing bis(2-acetylamino-1,3,4-thiadiazole)-5,5’-disulfonamide (BATD)

Bis(2-acetylamino-1,3,4-thiadiazole)-5,5’-disulfonamide (BATD) (dimer) was obtain- ed according to the method of Sarbu and co-workers described in item [20]. First, 0.40 g (0.01 mole) of sodium hydroxide was diluted in 100 cm3 of water. Then, 2.2 g (0.01 mole) of technical acetazolamide was introduced at temperature 5–10 oC. Within the time of 1 hour, wet 2-acetylamino-5-sulfochloro-1,3,4-thiadiazole was added. After introducing sulfochloride, the mixture was kept for 90 minutes at the temperature ranging from 5 to 10 oC. Residual, unreacted acetazolamide and sulfochloride were filtered out. Afterwards, the filtrate was treated with the 40 % aqueous solution of sodium hydroxide until it reached pH = 12. After 24 hours, a white precipitate was found in the filtrate, then it was separated and dissolved in the minimum amount of water, heated and acidified with 36.0 % hydrochloric acid (d = 1.18 g/cm3)upto1to3 pH. After cooling, white crystals were separated from the solution, which were filtered and dried in the ambient temperature to the constant weight. 0.645 g of the product was obtained (yield: 15.3 %); melting point: 307–311 oC (lit.: m.p. 318 oC [20]). 1694 Teresa Rauckyte-¯ak and Bo¿ena Szejniuk

The solutions of the synthesised compounds of concentration from 0.5 to 100.0 mg/cm3, presented above, were prepared with the use of the dimethylsulfooxide (DMSO) solvent. A control sample with bactericidal properties was made for every strain of bacteria. The effect of tested heterocyclic compounds concerning their sensitivity to Gram-positive and Gram-negative aerobic bacteria, such as: Escherichia coli, Staphylococcus aureus, Bacillus subtilis, Bacillus cereus, Proteus mirabilis, Enterococcus faecium, Salmonella senftenberg and Salmonella typhimurium was determined. Bacterial strains were received from Hohenheim University in Stuttgart. The above microorganisms were multiplied within 24 hours and diluted in liquid substratum: Standard-I Broth and added to the solid medium: Standard-I Agar (Standard I-Nhragar, Merck No. 7881) in the amount equal to 0.5 cm3 of suspended microbes per 250.0 cm3 of agar. Then, Agar with vaccinated microbes was spilled on the Petri dishes and left to solidify. Afterwards, cylindrical wells with the radius of 4 mm were cut in the solidified bases, and filled with solutions of the tested compounds (this activity was performed twice each time and the amounts used were, respectively: 50.0 and 100.0 mg/cm3; 25.0 and 50.0 mg/cm3; 1.0 and 12.5 mg/cm3; 0.25 and 0.5 mg/cm3). The dissolvent itself was acting as the control. The samples were incubated in the temperature of 37 oC for 24 hours [21]. After the incubation, the areas of microbes growth inhibition (in mm) were measured and the minimal inhibition concentrations (MIC) were defined.

Results and discussion

The bactericidal properties of sulfonamides with free amine group in para-position in benzene ring [22] are discussed in the specialist literature. Seeking effective substances to eliminate pathogenic bacteria and, at the same time, the bacteria present in different types of pharmaceutical waste, a group of heterocyclic compounds was typed: 2-amino- -1,3,4-thiadiazole (AT) and 2-acetylamino-1,3,4-thiadiazole-5-sulfonamid (ATS), and bis(2-acetylamino-1,3,4-thiadiazole)-5,5’-disulfonamide (BATD). In order to achieve this aim, studies on biological activity of all the derivatives of 1,3,4-thiadiazole for selected environmental bacteria: Bacillus subtilis, Bacillus cereus, Proteus mirabilis, Enterococcus faecium, Salmonella senftenberg, Salmonella typhimurium, Escherichia coli and Staphylococcus aureus were carried out. Pure compounds contained in solid wastes and waste water from diuramid production were used in the research. Tables 1–3 present the determined activities of the tested chemical compounds for different concentrations reacting with the individual bacteria and values necessary to determine their biological activity. The values presented in the form of a table describe: R – half of the zone of bacteria growth inhibition and r – radius of the well equal to 4.0 mm, respectively. Basing on R/r, the activity of a given compound was determined for individual tested bacteria strains within the range of 1,3,4-thiadiazole concentration. As the results clearly illustrate: if the relation R/r < 1 occurs, it means low activity of the studied chemical substance. The relation R/r < 2 indicates medium and R/r ³ 2 high activity [21]. As the data demonstrate, after 24 hours of incubation at temperature 37 oC, the zone of bacteria growth inhibition expressed in the form of minimum inhibitory concentration (MIC) depended on the type of the tested compound as well as on the Influence of 1,3,4-Thiadiazole Derivatives on the Biological Activity... 1695 Table 1 ] 3 ; — – deficiency of activity. Concentration [mg/cm ; + – low activity 12.5 25.0 50.0 100.0 Activity of 2-acetamide-1,3,4-thiadiazole-5-sulfonamide (ATS) for chosen bacterial species R R/r Activity R R/r Activity R R/r Activity R R/r Activity ———————————— 4.04.0 1.0 1.03.0 ++2.5 ++ 0.83.5 4.0 0.63.0 3.0 0.9 +2.5 1.0 0.8 + 0.8 0.6 + 3.5 ++ + 3.0 + 0.9 + 2.5 4.0 0.8 3.5 3.5 0.6 + 3.0 1.0 0.9 + 0.9 0.8 + 4.0 ++ + 3.5 + 1.0 + 2.5 4.0 0.9 3.5 4.0 ++ 0.6 1.0 3.0 0.9 + 1.0 n.s. ++ 0.8 + + n.s. n.s. ++ + n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. Bacterial species Escherichia coli Bacillus cereus Staphylococcus aureus Bacillus subtilis Proteus mirabilis Enterococcus faecium Salmonella senftenberg Salmonella typhimurium Where: n.s. – non-studied; +++ – high activity; ++ – middle activity 1696 Teresa Rauckyte-¯ak and Bo¿ena Szejniuk Table 2 ] 3 ; — – deficiency of activity. Concentration [mg/cm ; + – low activity Activity of 2-amide-1,3,4-thiadiazole (AT) for chosen bacterial species 0.5 1.0 12.5 25 50 100 R R/r Activity R R/r Activity R R/r Activity R R/r Activity R R/r Activity R R/r Activity — — — 2.5 0.6 + 2.5 0.6 + 5.5 1.4 ++ 7.0 1.8 ++ n.s. n.s. n.s. 3.0 0.83.5 0.94.0 + 1.03.0 + 4.0 0.8 ++ 1.0 7.5 + 7.5 1.9 ++ 1.9 ++ 4.0 5.0 ++ 1.0 1.3 14.5 3.6 11.0 ++ ++ 2.8 +++ 6.5 +++ 6.0 11.5 1.6 1.5 2.9 10.0 2.5 ++ +++ ++ +++ 11.0 5.5 6.5 2.8 11.0 1.4 1.6 2.8 +++ ++ ++ +++ 12.0 3.0 n.s. 8.0 7.0 n.s. 2.0 +++ 1.8 n.s. +++ ++ n.s. n.s. n.s. 3.0 0.82.5 0.62.5 + 0.6 + 3.5 + 0.9 3.0 0.8 2.5 + 0.6 + 4.0 + 1.0 4.0 1.0 ++ 3.5 0.9 ++ 7.5 1.9 + 9.0 2.2 ++ 16.0 +++ 4.0 13.0 3.2 11.0 +++ 2.8 +++ 17.0 +++ 4.2 n.s. n.s. n.s. +++ n.s. n.s. n.s. n.s. n.s. n.s. Bacterial species Escherichia coli Staphylococcus aueus Bacillus subtilis Bacillus cereus Proteus mirabilis Enterococcus faecium Salmonella senftenberg Salmonella typhimurium Where: n.s. – non-studied; +++ – high activity; ++ – middle activity Influence of 1,3,4-Thiadiazole Derivatives on the Biological Activity... 1697 Table 3 n.s. n.s. n.s. ] 3 ; — – deficiency of activity. Concentration [mg/cm ; + – low activity 12.5 25.0 50.0 100.0 R R/r Activity R R/r Activity R R/r Activity R R/r Activity —— — — — 2.5 — 0.6 3.0 0.8 + + 3.0 0.8 3.5 0.9 + + n.s. n.s. n.s. n.s. n.s. n.s. ———————3.00.8+ — — 3.0 0.8 + 3.5 0.9 + 6.5 1.6 ++ 2.53.0 0.6 0.82.5 + 0.6 + 3.5 + 3.0 0.9 0.8 2.5 + 0.6 + 4.0 + 3.5 1.0 0.9 3.5 ++ 0.9 + n.s. + n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. 2.5 0.6 + — — — 2.5 0.6 + 3.0 0.8 + Activity of bis(2-acetamide-1,3,4-thiadiazole)-5,5’-disulfonamide (BATD) for chosen bacterial species Bacterial species Salmonella senftenberg Proteus mirabilis Enterococcus faecium Salmonella typhimurium Bacillus cereus Escherichia coli Bacillus subtilis Staphylococcus aueus Where: n.s. – non-studied; +++ – high activity; ++ – middle activity 1698 Teresa Rauckyte-¯ak and Bo¿ena Szejniuk bacteria strain. Table 1 puts together values referring to determined activities of ATS for the tested species of bacteria. Table 2 contains the results obtained for AT and Table 3 – these for BATD. Tables 1 and 3 present the results for the following concentrations applied in the tests: 12.5, 25.0, 50.0, and 100.0 mg/cm3. Additionally, Table 2 – presents concentrations 0.5 and 1.0 mg/cm3 for the most biologically active AT compound. Biological activity for all the derivatives of 1,3,4-thiadiazole with the concentration of 100.0 mg/cm3 were not tested for: Bacillus subtilis, Bacillus cereus, Proteus mirabilis, Enterococcus faecium, Salmonella senftenberg and Salmonella typhimurium. On the basis of the research carried out it was found that ATS shows medium activity of identical values in relation to Escherichia coli in all tested concentrations (Table 1). Low activity of ATS with concentrations: 12.5, 25.0, and 50.0 mg/cm3 was found for: Proteus mirabilis, Enterococcus faecium, Salmonella senftenberg and Salmonella typhimurium. It was found that for Staphylococcus aureus and Bacillus cereus low activity of ATS prevailed. No activity was detected for Bacillus subtilis in the whole range of concentrations. It could be the result of ATS chemical structure and the difference between its functional groups and reactive groups of Bacillus subtilis. Therefore, ATS is characterised by low biological activity in relation to the tested strains of bacteria. The next tested compound: 2-amino-1,3,4-thiadiazole (AT) demonstrates: high activity for Staphylococcus aureus with concentrations within the range 12.5–100.0 mg/cm3 (determined MIC = 12.5 mg/cm3), medium activity with concentration 1.0 mg/cm3, and low activity with concentration 0.5 mg/cm3 (Table 2). Diversified activity was found for Bacillus cereus, where high activity for concentration 50.0 mg/cm3, medium activity – for 1.0 mg/cm3 and low activity – for 0.5 mg/cm3; for Enterococcus faecium these are, respectively: high activity – MIC = 50.0 mg/cm3, medium – MIC = 12.5 mg/cm3, low MIC = 0.25 mg/cm3 (the only activity value possible to be determined at the concentration of 0.25 mg/cm3 is not included in the table); Salmonella senftenberg: MIC for high activity – 25.0 mg/cm3, MIC for medium activity – 12.5 mg/cm3, and MIC for low activity – 0.5 mg/cm3. In case of Escherichia coli, medium activity of AT was found within the range of concentrations from 1.0 to 100.0 mg/cm3 (MIC of medium activity: 1.0 mg/cm3) and low activity for 0.5 mg/cm3. Only Bacillus subtilis showed high and medium activities (MIC, respectively: 12.5 mg/cm3 and 0.5 mg/cm3) while Salmonella typhymurium showed high and low activities (MIC, respectively: 25.0 and 0.5 mg/cm3). The relation between AT and Proteus mirabilis appeared to be an interesting case in which medium activity was determined for concentrations 25.0 and 50.0 mg/cm3, low activity – for 1.0 and 12.5 mg/cm3, and zero activity for 0.5 mg/cm3. It was found that AT was characterised by the most diversified activity amongst the tested 1,3,4-thiadiazole derivatives in relation to Staphylococcus aureus, Bacillus cereus, Enterococcus faecium and Salmonella senftenberg bacteria, which could be determined by a suitable selection of MIC. Dimer bis(2-acetamido-1,3,4-thiadiazole)-5,5’-disulfonamide (BATD) is character- ised by its lack of activity or low activity in relation to the tested strains of bacteria. As in the case of ATS towards Bacillus subtillis, the dimer shows no activity and low Influence of 1,3,4-Thiadiazole Derivatives on the Biological Activity... 1699 activity for the concentration of 50.0 mg/cm3. The lack of BATD activity in relation to Bacillus subtilis may be interpreted by similarity of chemical structure of functional groups to ATS. It was found that the low activity of dimer occurred for Escherichia coli, Proteus mirabilis, Salmonella senftenberg, Enterococcus faecium, and Salmonella typhimurium. Interestingly enough, the lack of activity was observed for the concentration of 12.5 mg/cm3. The most diverse activities were observed for Staphylococcus aureus: from medium activity for the concentration of 100.0 mg/cm3, through low activity for the concentrations of 25.0 and 50.0 mg/cm3, and the lack of activity for the concentration of 12.5 mg/cm3. Medium activity was found for Bacillus cereus for the concentration of 50.0 mg/cm3, and the deficiency of activity for the concentrations of 12.5 and 25.0 mg/cm3. Figures 1–4 present the values’ dependence on the growth zone inhibition [in mm] for the given individual bacterial strains as a function relating to the concentration of 1,3,4-thiadiazole derivatives selected for the tests. Figure 1 shows the values’ dependence on the size of growth zone inhibition for Proteus mirabilis, Bacillus cereus, Bacillus subtilis, Staphylococcus aureus, Salmonella senftenberg, Salmonella typhimurium bacteria towards concentrations of biologically most active compound 2-amino-1,3.4-thiadiazole (AT). For all the tested strains of bacteria with the increase of AT concentration there is the increase in size of growth zone inhibition. The most steady increase of the growth zone inhibition was observed for Proteus mirabilis, Bacillus cereus and Bacillus subtilis. The difference between values of the growth zone inhibition was observed in case of: Salmonella senftenberg and Salmonella typhimurium for AT concentration of 25.0 mg/cm3, which for the concentration of 12.5 mg/cm3 was higher than the preceding value by 2.25 and 4.6 times, respectively. The values of growth zone inhibition of Staphylococcus aureus are worth considering. In this case a significant increase the trend was observed up to AT

35

30

25

20

15 Proteus mirabilis Bacillus subtilis 10 Salmonella senftenberg Bacillus cereus

Growth zone inhibition [mm] Staphylococcus aureus 5 Salmonella typhimurium

0 0.501.0012.50 25.00 50.00 100.00 Concentration of AT [mg/cm3 ]

Fig. 1. Effect of AT concentration on the level of the growth zone inhibition for selected strains of bacteria 1700 Teresa Rauckyte-¯ak and Bo¿ena Szejniuk concentration of 12.5 mg/cm3, and then the values of this parameter were kept on the same level. Figures 2–4 present the characteristics for Escherichia coli and Enterococcus faecium bacteria as its dependence on the growth zone inhibition in the function of ATS (Fig. 2), AT (Fig. 3), and BATD (Fig. 4) concentrations. These bacteria are considered to be an environmental sanitary indicator.

Escherichia 8 coli 7 Enterococcus faecium 6 5 4 3 2

rowth zone inhibition [mm] 1

G 0 12.5 25.0 50.0 100.0 Concentration of ATS [mg/cm3 ]

Fig. 2. Effect of ATS concentration on the level of the growth zone inhibition for E. coli and Enterococcus faecium

Together with the increase of ATS concentration, values of the growth zone inhibition for Escherichia coli remained unchanged at 8.0 mm. As far as the case of Enterococcus faecium is concerned, a different fact was observed: the value of the growth zone inhibition increased with the decrease of ATS. It might be determined that the optimal ATS concentration which inhibits the Enterococcus faecium’s growth zone is 12.5 mg/cm3. Figure 3 indicates a very steady increase of Escherichia coli growth zone inhibition with the increase of AT concentration within the range of 0.5–100.0 mg/cm3. A more

30 Escherichia coli 25 Enterococcus faecium 20

15

10

5

rowth zone inhibition [mm]

G 0 0.25 0.501.0012.50 25.00 50.00 100.00 Concentration of AT [mg/cm3 ]

Fig. 3. Effect of AT concentration on the level of the growth zone inhibition for E. coli and Enterococcus faecium Influence of 1,3,4-Thiadiazole Derivatives on the Biological Activity... 1701 rapid increase of the Enterococcus faecium growth zone inhibition (even up to 25.5 mm) is observed with the increase of AT concentration within the range from 0.25 to 50.0 mg/cm3. A small increase in the value of Escherichia coli and Enterococcus faecium growth zone inhibition (by one unit only) is characterised by the increase of BATD concentration within the range from 25.0 to 50.0 mg/cm3 for Enterococcus faecium, and from 12.5 mg/cm3 (50.0) to 100.0 mg/cm3 for Escherichia coli (Fig. 4).

Escherichia 6 coli Enterococcus 5 faecium

4

3

2

rowth zone inhibition [mm] 1

G 0 12.5 25.0 50.0 100.0 Concentration of BATD [mg/cm3 ]

Fig. 4. Effect of BADT concentration on the level of the growth zone inhibition for E. coli and Enterococcus faecium

For each strain of the tested bacteria, a control test of organic solvent used for dissolving the chemical compounds was carried out. The tests were conducted in order to define a possible effect of the solvent used on the tested microorganisms. The studied microbes are the representatives of pathogenic microorganisms that are generally identified in the environment [23] and belong to the group of the most thermo-tolerable bacteria. Only in case of Proteus mirabilis and Salmonella senftenberg bacteria an occasional minimum bright area of diameter not exceeding the sensitivity of the method was found.

Conclusion

On the basis of the research it was found out that 2-amino-1,3,4-thiadiazole (AT) presented the highest bacteriological activity in relation to all the tested micro- organisms. It was found that the activity inhibiting the growth of the tested bacteria in this case amounted to 0.25 mg/cm3. However, bis(2-acetylamino-1,3,4-thiadiazole- -5,5’-disulfonamide) (BATD) was characterized by the lowest bacteriological activity, because MIC for this compound was 12.5 mg/cm3. 2-amino-1,3,4-tiadiazol (AT) turned up to be the most effective as far as inhibiting the growth of the Salmonella typhimurium and Salmonella senftenberg bacteria is concerned. These microbes belong to the Enterobacteriaceae family which contains numerous species and strains living on different organisms and environments [24]. 1702 Teresa Rauckyte-¯ak and Bo¿ena Szejniuk

Salmonella typhimurium is a representative of the particularly dangerous microbes causing many pathogenic infections. The data indicate that in order to inhibit the growth of microbes belonging to Salmonella type, AT should be applied in the dosage of up to 25.0 mg/cm3 due to its high activity in this range of concentration. Also, the activity of this reagent with the Bacillus type microorganisms appeared to be quite effective, especially in the case of the tested Bacillus subtilis species. As it results from the data, its high activity was observed even in the case of the 12.5 mg/cm3 concentration, and medium activity was found for the concentration of 0.25 mg/cm3. The tested species of bacilli, which belong to the Bacillus type are the examples of the microorganisms living in the environment rich in organic substance characteristic for soil. Furthermore, from the clinical point of view they are a source of allergic chronic diseases and alimentary toxicoses [24]. The obtained results indicate that in order to eliminate Bacillus subtilis bacteria, high bactericidal activity was observed for the optimal AT concentration of 12.5 mg/cm3, but medium activity was observed even for 0.25 mg/cm3. It should be noticed that in case of determining bactericidal activity for this group of bacteria, the deficiency of activity was found in case of Bacillus subtilis for ATS and low activity in case of BATD for 50.0 mg/cm3. Escherichia coli and Enterococcus faecium bacteria were used as indicators of the environmental microorganisms (Fig. 2–4). The activity of ATS and AT, as far as their interaction with Escherichia coli is concerned, was found at the medium level for each tested concentration ranging respectively: from 12.5 to 100.0 and from 0.5 to 100.0 mg/cm3. The presence of these bacteria in the environment, especially in the surface waters, is a basic quality indicator of the biologically clean waters and allows for classifying them into their respective quality classes. However, Enterococcus faecium bacteria which belong to Enterococcus type are the microbes characteristic for the environment as far as the pollution with fecal streptococci is concerned [23]. Furthermore, their presence points at different types of the environmental pollution with the substances of organic origin [25]. As far as their influence on Enterococcus Faecium microorganisms activity is concerned the tested compounds were characterized by low effectiveness in relation to ATS and BATD. It was found that AT activity is concentration-based and demonstrates high, medium and low activity with the con- centrations: 50 mg/cm3, 12.5 to 25.0 mg/cm3 and 0.25 to 0.5 mg/cm3, respectively. These data proved that the tested thiadiazole derivative could be selectively used to eliminate pollutions of fecal origin. Furthermore, it can be found that properly selected AT concentrations allow to eliminate selectively such microorganisms as Salmonella senftenberg, Bacillus cereus and Staphylococcus aureus.

References

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[4] Fenelon L.E., Mumtaz G. and Ridgway G.L.: The in-vitro susceptibility of Chlamydia pneumonia. J. Antimicrob. Chemother. 1990, 26, 763–767. [5] Varvarason A., Tantili-Kakoulidou A., Siatra-Papastasikoudi T. and Tiligada E.: Synthesis and biological evaluation of indole containing derivatives of thiosemicarbazide and their cyclic 1,2,4-tria- zole and 1,3,4-thiadiazole analogs. Arzneim. Forsch. 2000, 50, 48–54. [6] Pintilie O., Profire L., Sunel V., Popa M. and Pui A.: Synthesis and antimicrobial activity of some new 1,3,4-thiadiazole and 1,2,4-triazole compounds having a D,L-methionine moiety. Molecules 2007, 12, 103–113. [7] Faroumadi A., Mirzaei M. and Shafiee A.: Synthesis and antituberculosis activity of 2-aryl-1,3,4-thia- diazole derivatives. Pharmazie 2001, 56, 610–612. [8] Mamolo M.G., Falagiani V., Zanpier D., Vio L. and Banfi F.: Synthesis and antimycobacterial activity of [5-(pyridin-2-1,3,4-thiadiazol-2-yl-thio)]-acetic acid arylidene-hydrazide derivatives. Farmaco 2001, 56, 587–592. [9] Clerici F., Pocar D., Guido M., Loche A., Perlini V. and Brufoni M.: Synthesis of 2-amino-5-sulphonyl- -1,3,4-thiadiazole derivatives and evaluation of their antidepressant and anxiolytic activity. J. Med. Chem. 2001, 44, 931–936. [10] Onkol T., Cakir B. and Sahin M.F.: Synthesis and antinociceptive activity of 2-[(2-oxabenzo- thiazolin-3-yl)-methyl]-5-aminoalkyl/aryl-1,3,4-thiadiazole. Turk. J. Chem. 2004, 28, 461–466. [11] Shenone S., Bruno O., Ranise A., Bondavalli W., Falcone G., Giordano L. and Vitelli M.: 3-Arylsulpho- nyl-5-arylamino-1,3,4-thiadiazol-2(3H)ones as anti-inflammatory and analgesic agents. Bioorg. Med. Chem. 2001, 9, 2149–2153. [12] Moise M., Sunel V., Profire L., Popa M., Desbrieres J. and Peptu C.: Synthesis and biological activity of some new 1,3,4-thiadiazole and 1,2,4-triazole compounds containing a phenylalanine moiety. Molecules 2009, 14, 2621–2631. [13] Labanauskas L., Kalcas V., Uderenaite E., Gaidelis P., Brukstus A. and Dauksas V.: Synthesis of 3-(3,4-dimethoxyphenyl)-1H-1,2,4-triazole-5-thiol and 2-amino-5-(3,4-dimethoxyphenyl)-1,3,4-thiadia- zole derivatives exhibiting anti-inflammatory activity. Pharmazie 2001, 56, 617–619. [14] Palaska E., Sahin G., Kelincen P., Durlu N.T. and Altionax G.: Synthesis and anti-inflammatory activity of 1-acyl thiosemicarbazides, 1,3,4-oxadiazoles, 1,3,4-thiadiazoles and 1,2,4-triazole-3-thiones. Farmaco 2002, 57, 101–107. [15] Roblin R.O. and Clapp J.W.: The preparation of heterocyclic sulfonamides. J. Amer. Chem. Soc. 1950, 72, 4890. [16] Rauckyte T. and ¯ak S.: An attempt to apply Fenton system in wastewater pretreatment from acetazolamide production. Ecol. Chem. Eng. 2006, 13(10), 1133–1141. [17] M³ochowski J.: Chemistry of heterocyclic compounds, PWN, Warszawa 1994 (in Polish). [18] Sandström J.: Recent advances in the chemistry of 1,3,4-thiadiazoles. Adv. Heterocycl. Chem. 1968, 9, 165–209. [19] Ainsworth C.: The reaction of thiosemicarbazide with orthoesters. J. Amer. Chem. Soc. 1956, 78, 1973. [20] Sarbu C., Hodisan T., Gocan S. and Liteanu C.: Analysis of acetazolamide by thin layer chromato- graphy. Farmacia (Bucharest) 1981, 29, 105–108. [21] Ionita A., Moscovici M., Popa C., Vamanu A., Popa O. and Dinu L.: Screening of yeast and fungal strains for lypolytic potential and determination of some biochemical properties of microbial lipases. J. Mol. Catal. B: Enzim. 1997, 3, 147–151. [22] Kowalik-Jankowska T.: Coordination ability of sulfonamide bond in complexes of copper (II) ions. Wiad. Chem. 1996, 50(1–2), 143–153. [23] Strauch D.: Animal waste as risk for animal and human health. Proc. 9th Int. Congr. Animal Hygiene ISAH’97, Helsinki 1997, 597–607. [24] Ko³odyñski J.: Fundamentals of bacteriology, 1998, Wyd. UW Wroc³aw (in Polish). [25] Philipp W., Grunwald R., Pinkepank T. and Böhm A.: Hygienic problems in anaerobic fermentation of slurry together with different industrial and municipal wastes in rural areas (Co-fermentation). Proc. 9th Int. Congr. Animal Hygiene ISAH’97, Helsinki 1997, 608–611. 1704 Teresa Rauckyte-¯ak and Bo¿ena Szejniuk

WP£YW POCHODNYCH 1,3,4-TIADIAZOLI NA AKTYWNOŒÆ WYBRANYCH BAKTERII ŒRODOWISKOWYCH

1 Wydzia³ Technologii i In¿ynierii Chemicznej, 2 Wydzia³ Biologii i Hodowli Zwierz¹t Uniwersytet Technologiczno-Przyrodniczy w Bydgoszczy

Abstrakt: Przedstawiono wyniki badañ mikrobiologicznych, prowadzonych z udzia³em mikroorganizmów, wystêpuj¹cych m.in. w œrodowisku biologicznym œcieków gospodarczo-bytowych oraz w osadach œcieko- wych z komunalnych oczyszczalni. Badano wp³yw wybranych zwi¹zków heterocyklicznych pochodnych 1,3,4-tiadiazolu na wra¿liwoœæ bakterii aerobowych Gram-dodatnich i Gram-ujemnych. Do badañ aktywnoœci biologicznej wytypowano szczepy bakterii œrodowiskowych: Escherichia coli, Staphylococcus aureus, Bacillus subtilis, Bacillus cereus, Proteus mirabilis, Enterococcus faecium, Salmonella senftenberg i Salmo- nella typhimurium. Dla badanej grupy bakterii najbardziej aktywnym zwi¹zkiem chemicznym okaza³ siê 2-amino-1,3,4-tiadiazol. Stwierdzono, ¿e odpowiednio dobrane stê¿enia tego zwi¹zku pozwalaj¹ na selektyw- n¹ eliminacjê takich drobnoustrojów, jak: Salmonella senftenberg, Bacillus cereus, Staphylococcus aureus i Enterococcus faecium. W przypadku Escherichia coli najni¿sze stê¿enie hamuj¹ce wzrost bakterii (MIC) stwierdzono na poziomie 12,5 mg/cm3 (wyznaczone wzglêdem 2-acetyloamino-1,3,4-tiadiazolo-5-sulfo- namidu i bis(2-acetyloamino-1,3,4-tiadiazolo)-5,5’-disulfonamidu), a dla 2-amino-1,3,4-tiadiazolu 0,5 mg/cm3. Podobne wyniki uzyskano w przypadku bakterii Enterococcus faecium z tym, ¿e dla 2-amino-1,3,4-tiadiazolu MIC < 0,25 mg/cm3, a dla bis(2-acetylamino-1,3,4-tiadiazolo)-5,5’-disulfonamidu MIC = 25,0 mg/cm3. S³owa kluczowe: pochodne 1,3,4-tiadiazolu, badania mikrobiologiczne, wskaŸnik sanitarny œrodowiska ECOLOGICAL CHEMISTRY AND ENGINEERING A

Vol. 18, No. 12 2011

Krystyna PRZYBULEWSKA1, Anna STOLARSKA2 and Daria G£¥BOWSKA1

CHANGE OF PROLINE CONTENT IN SELECTED SOIL FUNGI AS AFFECTED BY OSMOTIC STRESS

ZMIANA ZAWARTOŒCI PROLINY U WYBRANYCH GRZYBÓW GLEBOWYCH POD WP£YWEM STRESU OSMOTYCZNEGO

Abstract: The study aimed at demonstrating intracellular proline synthesis under osmotic stress conditions in selected soil fungi on the example of Trichoderma sp. and Trichotecium roseum. Effect of the increase of sodium chloride (NaCl) salinity, in concentrations of 1 to 1000 mmol × dm–3 PDA medium, on proline content in the fresh matter of mycelium cultured on medium was examined. The increase of medium salinity affects production of osmoregulatory substances in the form of proline in selected soil fungi. Its content in mycelium depends on salt (NaCl) concentration in medium as well as on species. Intracellular proline synthesis increased starting with the least salinity. As osmotic stress increased, proline content in mycelium almost quadrupled in Trichoderma sp. and quintupled in Trichotecium roseum. Keywords: fungi, salinity, proline

Fungi play an important role in decomposition of organic matter and humification processes in soil. Their number and activity depend largely on different factors, both natural and anthropogenic ones, such as temperature, pH or osmotic pressure. Salinity induces increase of osmotic pressure, which limits water uptake or even makes it impossible and can be a reason of growth arrest or even of cell destruction [1–6]. Microorganisms show certain adaptative mechanisms and adaptabilities which allow them to outlast unfavourable conditions in environment [7–9]. Microorganisms are capable of intracellular accumulation of osmoregulatory substances which allow preserving a turgor indispensable for proper cell functioning [1, 10–12]. These substances include different organic compounds such as amino acids, peptides,

1 Department of Microbiology and Biotechnology of Environment, West Pomeranian University of Technology, ul. J. S³owackiego 17, 71–434 Szczecin, Poland, phone: +48 91 449 64 24, email: krystyna.przy- [email protected] 2 Department of Plant of Physiology, West Pomeranian University of Technology, ul. J. S³owackiego 17, 71–434 Szczecin, Poland. 1706 Krystyna Przybulewska et al saccharides, alcohols and others [13–14]. One of these compounds is proline which can be absorbed from environment by microorganisms or synthesised intracellularly by them to tolerate easily growth inhibition [1, 15–19]. The study aimed at demonstrating intracellular proline synthesis on the example of selected soil fungi under osmotic stress conditions.

Material and methods

Two soil fungi, ie Trichderma sp. and Trichotecium roseum (Photo 1 and 2) were used in the study. Species affinity was determined with microscopic methods and was done by gas chromatography fatty acid methyl ester (FAME) analysis performed by Microbial ID (Newark, DE, USA). Effect of the osmotic stress resulting from sodium chloride (NaCl) salinity in concentrations of 1, 10, 100 and 1000 mmol × dm–3 PDA medium was examined. Mycelium cultured on salt-free medium was a control. Experiment was set up in three replications. Culture incubation was carried out on solid media at 24 oC for 2 weeks. Mycelium was homogenised in 10 % aqueous solution of sulfosalicylic acid in order to determine proline content.

Photo 1. Trichotecium roseum Photo 2. Trichderma sp.

Homogenate was filtered through Whatman No. 2 filter paper and filled up to a volume of 10 cm3. Proline content was determined with the method of Bates et al [20]. Data were converted and given in mg of proline × g–1 of mycelium fresh matter. The obtained results were analysed statistically, using analysis of variance and testing the factors with the Duncan’s test. Also a Pearson’s correlation coefficient was calculated between medium salinity increase and mycelium proline content.

Results and discussion

The number and activity of fungi depends largely on various factors, both natural and anthropogenic ones [7–9]. Osmotic stress, resulting from the increase of medium salinity, has a significant effect on their development and enzymatic activity [4–6]. Change of Proline Content in Selected Soil Fungi... 1707

Based on the analysis of results, it is possible to state that the increase of medium salinity with sodium chloride (NaCl) significantly affected intracellular proline content in the mycelium of examined species. Increase in its content as affected by osmotic stress differed depending on the fungi examined (Fig. 1).

10 Trichderma sp. Trichotecium roseum 8

6

–1

× LSD0.05 I = 0.133 LSD = 0.229 m 0.05 II 4 LSD0.05 I×II = 0.333

Proline [ g g f.m.]

2

0 0 1 10 100 1000 NaCl [mmol× dm–3]

Fig. 1. Proline content in mycelium under sodium chloride (NaCl) medium salinity conditions

Proline content in the mycelium cultured on control medium without NaCl addition amounted to 1.32 mg × g–1 f.m. in Trichoderma sp., whereas its content was by 39 % larger in Trichotecium roseum. Salt addition to medium in the form of NaCl induced a significant increase of the content of examined amino acid irrespective of the species of examined fungus. The least amount of salt introduced into medium, ie 1 mmol × dm–3, increased proline content by 15 % in Trichoderma sp. and by 24 % in Trichotecium roseum in relation to control (Photo 2). A tenfold and hundredfold increase of medium salinity increased proportionally its content in the mycelium of both examined species. Proline content in mycelium fresh matter was by 70–80 % larger at medium salinity with 10 mmol NaCl × dm–3, while it exceeded its amount two- and a half-fold in relation to its content in the mycelium cultured on control medium. The largest proline content, ie 8.45 mg × g–1 f.m., was found in Trichotecium roseum cultured on the most salinated medium (1000 mmol NaCl × dm–3). Significantly smaller proline content (33 %) under largest osmotic stress conditions was found in case of the second examined fungus, ie Trichoderma sp. Magnitude of the synthesis of osmoregulatory substances of that sort depends on the type of microorganisms and in case of this study depends even on fungus species. On average, its content was almost 20 % larger in Trichotecium roseum than in Trichoderma sp. (Fig. 2). The carried out study confirms information that also soil fungi, as exampled by Trichoderma sp. and Trichotecium roseum, are capable of intracellular accumulation of specific osmoregulatory substances which are stress proteins, in this case proline, under 1708 Krystyna Przybulewska et al

600 Trichderma sp. Trichotecium roseum 450

300

% of control

150

0 K 1 10 100 1000 average

Salinity (NaCl)

Fig. 2. Proline content in mycelium under sodium chloride (mmol NaCl × dm–3) medium salinity conditions expressed as percent of control medium salinity increase conditions. These results are corroborated by other authors in studies on bacteria or other eukaryotic organisms [1, 15–18]. In addition, the obtained regression equation, ie y = 0.0043x + 2.252, confirmed by significant correlation coefficient r = 0.84, points to the presence of dependence between medium salinity increase and mycelium proline synthesis.

Conclusions

1. Increase of medium salinity affects production of osmoregulatory substances in the form of proline in selected soil fungi. Its content in mycelium depends on salt (NaCl) concentration in medium as well as on fungus species. 2. Intracellular proline synthesis increased starting with the least medium salinity (1 mmol NaCl × dm–3) and amounted on average to 20 % when compared with control. As salinity increased, its content increase in mycelium almost quadrupled in Tricho- derma sp. and quintupled in Trichotecium roseum under the strongest osmotic stress conditions.

References

[1] Csonka L.N.: Physiological and genetic responses of bacteria to osmotic stress. Microbiol Rev. 1989, 53, 121–147. [2] Lopez C.S. Heras H., Garda H., Ruzal S. Sanchez-Rivas C. and Rivas E.: Biochemical and biophysical studies of Bacillus subtilis envelopes under hyperosmotic stress. Int. J. Food Microbiol. 2000, 55(1–3), 137–142. [3] Koch S. Oberson G. Eugster-Meier E., Meile L. and Lacroix Ch.: Osmotic stress induced by salt increases cell yield, autolytic, and survival of lyophilization of Lactobacillus delbrueckii supsp. lactis. Int. J. Food Microbiol. 2007, 117(1), 36–42. [4] Przybulewska K., Stolarska A. and B³aszak M.: Wp³yw zasolenia pod³o¿a chlorkiem sodu (NaCl), azotanem sodu (NaNO3) i ich mieszanin¹ na aktywnoœæ enzymatyczn¹ wybranych grzybów glebowych. Czêœæ II. Hydroliza t³uszczu. Wyd. Uczelniane UTP, Bydgoszcz 2008, 150–151. Change of Proline Content in Selected Soil Fungi... 1709

[5] Przybulewska K., Stolarska A. and B³aszak M.: Wp³yw zasolenia pod³o¿a chlorkiem sodu (NaCl), azotanem sodu (NaNO3) i ich mieszanin¹ na aktywnoœæ enzymatyczn¹ wybranych grzybów glebowych. Czêœæ III. Hydroliza skrobi. Ecol. Chem. Eng. A 2009, 16(4), 405–410. [6] Przybulewska K. and Wieczorek A.: Effect of medium salinity with sodium chloride (NaCl), sodium nitrate (NaNO3) and their compound on the enzymatic activity of selected soil fungi. Part 1. Protein hydrolysis. Ecol. Chem. Eng. A. 2007, 14(11), 1198–1202. [7] Boch J., Kempf B. and Bremer E.: Osmoregulation in Bacillus subtilis: synthesis of osmoprotectant glycine betaine from exogenously provided choline. J. Bacteriol. 1994, 176, 5364–5371. [8] Paw³owski S., Gulewicz P. and Grajek W.: Wp³yw ciœnienia osmotycznego na stan fizjologiczny komórek bakteryjnych. Post. Biol. Komór. 2002, 29(3), 435–448. [9] Kogej T., Stein M., Volkmann M., Gorbushina A.A., Galinski E.A. and Gunde-Cimerman N.: Osmotic adaptation of the halophilic fungus Hortaea werneckii: role of osmolytes and melanization, Micro- biology 2007, 153, 4261–4273. [10] Kempf B. and Brehmer E.: Uptake and synthesis of solutes as microbial stress responses to high-osmolality environments. Arch. Microbiol. 1998, 170, 426–431. [11] Poolman B. and Glaasker E.: Regulation of compatible solute accumulation in bacteria. Molecular Microbiol., 1998, 29, 397–407. [12] Mager W.H. and Varela J.C.S.: Osmostress response of the yeast Saccharomyces. Molec. Microbiol. 1993, 10, 253–258. [13] Galinski E.A. and Truper H.: Microbial behaviour in salt-stressed ecosystems. FEMS Microbial. Rev. 1994, 15, 95–108. [14] Galiñska E.A.: Osmoadaptation in bacteria. Adv. Microbial. Physiol. 1995, 37, 273–328. [15] Whatmore A.M., Chudek J.A. and Reed R.H.: The effects osmotic upshock on the intercellular solute pools of Bacillus subtilis. J. Gen. Microbiol. 1990, 136, 2527–2535. [16] Chris A., Zeeshan M., Abraham G. and Prasad S.M.: Proline accumulation in Cylindrospermum sp. Environ. Exper. Bot. 2006, 57, 154–159. [17] Lucht J. and Bremer E.: Adaptation of Escherichia coli to high osmolarity environments: osmoregulation of the high-affinity glycine betaine transport system proU. FEMS Microbiol. Rev. 1994, 14(1), 3–20. [18] Kavahara Y., OhsumiI T., Yoshihara Y. and Ikeda S.: Proline in the osmoregulation of Brevibacterium lactofermentum. Agr. Biol. Chem. 1989, 53, 2475–2479. [19] Von Blohn C., Kempf B., Kappes R.M. and Bremer E.: Osmostress response in Bacillus subtilis: Characterization of a proline Uptake system (OpuE) regulated by high osmolarity and alternative transcription factor sigma b. Mol. Microbiol. 1997, 25, 175–187. [20] Bates L.S.: Rapid determination of free proline for water stress studies. Plant Soil 1973, 39, 205–207.

ZMIANA ZAWARTOŒCI PROLINY U WYBRANYCH GRZYBÓW GLEBOWYCH POD WP£YWEM STRESU OSMOTYCZNEGO

Katedra Mikrobiologii i Biotechnologii Œrodowiska Zachodniopomorski Uniwersytet Technologiczny w Szczecinie

Abstrakt: Celem badañ by³o wykazanie syntezy wewn¹trzkomórkowej proliny w warunkach stresu osmotycznego u wybranych grzybów glebowych na przyk³adzie Trichoderma sp. i Trichotecium roseum. Badano wp³yw stresu osmotycznego w wyniku zasolenia chlorkiem sodu (NaCl), w stê¿eniu od 1 do 1000 mmol × dm–3 po¿ywki PDA na zawartosæ proliny w œwie¿ej masie grzybni wyros³ej na pod³o¿u. Wzrost zasolenia pod³o¿a wp³ywa na wytwarzanie substancji osmoregulacyjnych w postaci proliny u wybranych grzybów glebowych. Jej zawartoœæ w grzybni jest zale¿na od stê¿enia soli NaCl w pod³o¿u oraz od gatunku. Synteza proliny wewn¹trzkomórkowej zwiêksza³a siê pocz¹wszy od najmniejszego zasolenia. W miarê wzrostu stresu osmotycznego zawartoœæ proliny w grzybni zwiêkszy³a siê prawie czterokrotnie u Trichoderma sp. i piêciokrotnie Trichotecium roseum.

S³owa kluczowe: grzyby, zasolenie, prolina

ECOLOGICAL CHEMISTRY AND ENGINEERING A

Vol. 18, No. 12 2011

Joanna JARMU£-PIETRASZCZYK1, Marta KAMIONEK1 and Ewelina MALINOWSKA2

OCCURRENCE OF ENTOMOPATHOGENIC FUNGI AND NEMATODES ON PASTURES IN CENTRAL AND NORTHERN POLAND

WYSTÊPOWANIE GRZYBÓW I NICIENI ENTOMOPATOGENNYCH NA PASTWISKACH W CENTRALNEJ I PÓ£NOCNEJ POLSCE

Abstract: Entomopathogenic fungi and nematodes as important elements of the natural environment focussed the attention of researchers. Both groups of organisms possess an ability to infect and reduce the number of insects at various stages of their growth. This ability was used in the production of biopreparations. For this reason, the studies were aimed at estimating species composition and population density of entomopathogenic fungi and nematodes living in a given ecosystem. Experiments were performed in soils obtained from pastures. Fungi: Metharisum anisopliae, Bauveria basianna, Pecylomyces fumosoroseus as well as and nematodes of the family Steinernematidae were isolated from soil samples. Keywords: entomopathogenic fungi and nematode, soil

Entomopathogens comprise a very numerous group of microorganisms including fungi and nematodes. Under natural conditions these organisms were found in various ecosystems. They are an important factor decreasing population density of eg plant pests. Arthropods are their main food base though they do not show distinct food preferences [1–4]. The most frequent entomopathogenic fungi in Poland are the species of the genera: Peacilomyces, Bauveria, and Metarhizium. Apart from entomopathogenic nematodes, these organisms are a very efficient factor controlling arthropod populations and the first discovered pathogens of insects [2, 5, 6]. They were used to control pests already at the end of the 19th century. Intensification of agricultural production and common use of plant protection chemicals inclined researchers to search for alternative methods of plant protection. The last decade has brought many studies on these groups

1 Division of Zoology, Warsaw University of Life Sciences – SGGW, ul. Ciszewskiego 8, 02–786 Warszawa, Poland, phone: +48 22 593 66 28, fax: +48 22 593 66 23, email: [email protected] 2 Student of the Division of Zoology, Warsaw University of Life Sciences – SGGW, ul. Ciszewskiego 8, 02–786 Warszawa, Poland. 1712 Joanna Jarmu³-Pietraszczyk et al of organisms which contributed to the commercial use and production of bioprepara- tions based on local isolates. Species composition and density of entomopathogenic fungi and nematodes depends on factors like: soil type and moisture, human impact and land use in a given area [7–11].

Material and methods

Soil samples collected in 2007 and 2008 from variously managed pastures were used in the experiment. Mixed samples were taken with the soil cane to the depth of 20 cm. The first sample originated from barren pasture in Niekursk which was extensively grazed by sheep. The second study area was a pasture in Konradow grazed by slaughter cattle of the Hereford race and the third pasture was situated in Spala. Since 1985 this pasture has been unmanaged. Entomopathogenic fungi and nematodes were isolated from soil samples with the method of Zimmermann (1986) [12] using trap insects. The trap organisms were the G. mellonella caterpillars from the culture bred in the Department of Zoology, WULS. The experiment was carried out at 25 oC for 25 days for each soil sample. The first control was performed 5 days after the set up of the experiment and later the samples were controlled every 2–3 days. Dead insects were removed from soil samples and replaced by live ones. Dead insects were transferred to Petri dishes to estimate the reason of their death, to obtain complete growth of mycelium on the skin surface or to multiplicate entomopathogenic nematodes for estimation of their species composition. The obtained results were statistically processed with the SPSS 11 statistical software.

Results and discussion

The use of pastures affected species composition of isolated pathogens and intensity of their occurrence. Three most frequent species of entomopathogenic fungi were isolated from the studied samples were: P. fumosoroseus, B.bassiana, and M. anisopliae (Table 1). Apart from fungi, entomopathogenic nematodes of the family Steiner- nematidae were also found in studied area. M. anisopliae was the most frequent in pastures in Spala and Niekursk. The second frequent fungal species in these pastures was P. fumosoroseus. The least frequent species was B. bassiana (Table 1).

Table 1 The occurrence of entomopathogenic fungi and nematode in the soil [%]

Niekursk Konradow Spala Entomopathogenic fungi B. bassiana 2.0 0.7 10.0 P. fumosoroseus 6.2 24 36.7* M. anisopliae 17.4* 0 31.1 Saprofic fungi 10.0 43.3 6.7 Entomopathogenic nematode (Steinernematidae) 48.6 27.3* 7.8 Other biotic factor 16.1 4.7 7.8 p < 0.05 for the test ANOVA. Occurrence of Entomopathogenic Fungi and Nematodes on Pastures... 1713

The rate of insect infection by nematodes is largely determined by the size of the nematode population in soil and by their ability to infect the host. Studies of three soils from different pastures showed high variability of trap insect infection by nematodes (Table 1). Most entomopathogenic nematodes were isolated from the soil from Niekursk in the first week of the experiment. Isolating entomopathogenes from soil presents problems with rich soil microflora (saprophagous fungi, plant pathogens, various bacteria and saprophytes). The use of trap insects enables isolation of most frequent species and their further culture under laboratory conditions [1–4, 12, 13] to estimate their species composition. Many authors underline that the occurrence of entomopathogens varies seasonally and their com- position depends on sampling date. Entomopathogenic nematodes attack their prey most often in summer and late autumn. This was confirmed in the present experiment. Autumn samples from Niekursk contained more nematodes which infected insects before fungal infection. According to Ropek (2005) [14] the occurrence of spores of entomopathogenic fungi in soil may significantly decrease nematode infections. Fungal spores infecting an insect produced enzymes that inhibit the growth of mutualistic bacteria living in symbiosis with nematodes. Another factor affecting differentiation of pathogens was the type of pasture used. Pastures in Niekursk and Konradow are used permanently in contrast with that in Spala. Organic fertilisation affects mainly the occurrence of entomopathogenic nematodes [3, 9–11] and less intensively the abundance of fungi [2, 5, 15, 16].

Conclusions

Entomopathogenic fungi were mainly isolated from the soil from pastures in Spa³a and Niekursk. The dominating species was M. anisopliae. Nematodes of the family Steinernematidae were less frequent. No representatives of Heterorhabditis species were isolated.

References

[1] Bajan C. and Kmitowa K.: Polish Ecol. Stud. 1997, 23(3–4), 135–136 and 149–151. [2] Ba³azy S.: Biotechnologia 2000, 3(50), 11–32. [3] Bednarek A.: Ekologiczne uwarunkowania aktywnoœci biologicznej nicieni entomofilnych w œrodowisku glebowym agrocenoz. Wyd. SGGW, Warszawa 1990, 11–30. [4] Boguœ M.: Biotechnologia 2000, 3(50), 33–46. [5] Ignatowicz S.: Nowoczesne Rolnictwo 1998, 4, 44–45. [6] Miêtkiewski R.: Ochrona Roœl. 1992, 11, 7–8. [7] Jaworska M., Ropek D., Mazur K. and Filipek-Mazur B.: Acta Agrophys. 2001, 52, 79– 85. [8] Ropek D.: Zesz. Nauk Akad. Roln. im. H. Ko³³¹taja w Krakowie. 2005, 30, 1–26 and 146–169. [9] Matuska J. and Kamionek M.: Ecol. Chem. Eng. A 2008, 15(4–5), 383–387. [10] Jarmu³-Pietraszczyk J., Tkaczuk C., Kamionek M. and Pezowicz E.: Ecol. Chem. Eng. A 2008, 15(4–5), 349–353. [11] Pezowicz E. Kamionek M., Jarmu³ J., Ecol. Chem. Eng. 2008, 15(4–5), 389–392. [12] Zimmermann G. J.: Appl. Entomol., 1986, 102, 213–215. [13] Miêtkiewski R., Miêtkiewska Z. and Tkaczuk C.: Zesz. Nauk Wy¿. Szk. Roln.-Pedag. w Siedlacach, Rol., 1995, 37, 179–185. 1714 Joanna Jarmu³-Pietraszczyk et al

[14] Martyniuk S. and Stachyra A.: Nawozy i nawo¿enie. 2006, 4(29), 135–140. [15] Kamionek M.: Wp³yw pestycydów na Steinernema feltiae (Filipjev) i inne nicienie entomopatogeniczne, Wyd. SGGW, Warszawa 1992, 9–8.

WYSTÊPOWANIE GRZYBÓW I NICIENI ENTOMOPATOGENNYCH NA PASTWISKACH W CENTRALNEJ I PÓ£NOCNEJ POLSCE

1 Katedra Biologii Œrodowiska Zwierz¹t, Zak³ad Zoologii 2 Student – Katedra Biologii Œrodowiska Zwierz¹t, Zak³ad Zoologii Szko³a G³ówna Gospodarstwa Wiejskiego w Warszawie

Abstrakt: Polska jako kraj o zró¿nicowanej topografii stwarza dogodne warunki do rozwoju oraz zró¿nicowania fauny. Do niej zaliczamy m.in. grzyby i nicienie entomopatogenne, które s¹ wa¿nym elementem œrodowiska. Wa¿n¹ rol¹ jest ich zdolnoœæ do infekowania ró¿nych stadiów rozwojowych owadów i ich redukcja. Te umiejêtnoœci s¹ wykorzystywane w produkcji biopreparatów. Z tego powodu prowadzone badania mia³y na celu okreœlenie sk³adu gatunkowego oraz wielkoœæ populacji grzybów i nicieni entomo- patogennych wystêpuj¹cych w okreœlonym ekosystemie. Doœwiadczenie przeprowadzono na glebach po- zyskanych z pastwisk. Z badanych próbek glebowych wyizolowano takie grzyby, jak: M. anisopliae, B. basianna, P. fumosoroseus oraz nicienie z rodziny Steinernematidae. S³owa kluczowe: nicienie entomopatogenne, grzyby entomopatogenne, gleba ECOLOGICAL CHEMISTRY AND ENGINEERING A

Vol. 18, No. 12 2011

Wiera MICHALCEWICZ1*, S³awomir STANKOWSKI2, Ma³gorzata GA£CZYÑSKA3 and Marzena GIBCZYÑSKA3

SUCCESSIVE INFLUENCE OF FLUIDAL ASHES ON GENERAL NUMBER OF BACTERIA, ACTINOMYCETES AND FUNGI IN POT EXPERIMENTS

NASTÊPCZY WP£YW POPIO£ÓW FLUIDALNYCH NA OGÓLN¥ LICZBÊ BAKTERII, PROMIENIOWCÓW I GRZYBÓW W BADANIACH WAZONOWYCH

Abstract: The aim of conducted studies was to determine the impact of fluidal ashes obtained from coal combined with fermented sewage sludge and straw and using effective microorganisms, on the overall abundance of bacteria, fungi, actinomycetes and coli bacteria. The experiments were conducted in a pot environment. An increase in the number of bacteria and actinomycetes in the samples containing fluidal ashes and various organic components was observed in comparison with the first year of studies. However, the number of coli bacteria, compared with the first two years of the experiment, was significantly reduced. Keywords: fluidal ash, bacteria, fungi, actinomycetes, coli bacteria

Because of physicochemical properties such as very high pH and low organic matter content, slag waste is not a suitable environment for the development of micro- organisms. Therefore, when it is used in agriculture, it becomes necessary to enrich it with various types of organic materials, such as composted sewage sludge, straw or effective microorganisms (ceramic EM-X and EM-1), which would also be a source of microflora [1, 2]. The aim of this study was to compare the successive effect of fluidal ashes from CHP Zeran combined with fermented sewage sludge and straw, using effective microorganisms, on the number of bacteria, fungi, actinomycetes and coli bacteria.

1 Department of Microbiology and Environmental Biotechnology, West Pomeranian University of Technology, ul. J. S³owackiego 17, 71–434 Szczecin, Poland, email: [email protected] 2 Department of Agronomy, West Pomeranian University of Technology, ul. J. S³owackiego 17, 71–434 Szczecin, Poland, email: [email protected] 3 Department of General and Ecological Chemistry, West Pomeranian University of Technology, ul. J. S³o- wackiego 17, 71–434 Szczecin, Poland, email: [email protected] * Corresponding author. 1716 Wiera Michalcewicz et al

Material and methods

In the pot experiment the granulometric composition of loamy and slightly acidic soil (pHKCl 5.13) was used. The experiment was set up by the method of complete randomization in four replications. The scheme of the experiment included eight variants of fertilizing: 1 – control (soil), 2 – control II (fluidal ash), 3 – soil + sewage sludge + straw (4:2:1), 4 – fluidal ash + sludge + straw (4:2:1), 5 – soil + sewage sludge + straw (4:2:1) + microbiological formulation EM-1 (15 dm3 × ha–1), 6 – fluidal ash + sludge + straw (4:2:1) + microbiological formulation EM-1 (15 dm3 × ha–1), 7 – soil + sewage sludge + straw (4:2:1) + microbiological formulation EM-1 (15 dm3 × ha–1)+ EM ceramic powder-X (40 dm3 × ha–1), 8 – fluidal ash + sludge + straw (4:2:1) + microbial formulation EM-1 (15 dm3 × ha–1) + EM ceramic powder-X (40 dm3 × ha–1). Soil samples for microbiological analysis were taken after harvesting the crop (test plant Festulolium, variation Felopa). In the collected samples of ash and soil, the following parameters were determined: the number of bacteria on Bunte-Roviry’s base [3], the number of actinomycetes on Cyganow’s base [4] and the number of fungi on Martin’s base [5]. Because of the fact that digested sludge was used, the number of coli bacteria of faecal form was determined according to PN-77, C – 04615, Sheet 07, on Endo nourishment [6]. Cultures incubated at room temperature (20 oC) for 3 to 7 days. The number of coli bacteria was determined after 24 h of incubation at 37 oC. The results were subjected to univariate analysis of variance and then on the basis of Tukey’s test LSD0.05 values were calculated and homogenous groups were distinguished at significance level of P = 0.05.

Results and discussion

The obtained results are presented in Figs. 1, 2, 3 and 4, homogeneous groups are given. Analyzing the obtained results it should be noted that in comparison with the first year of studies a gradual increase in the number of bacteria in subsequent years was observed. The increase was from 2 to 10 times higher in comparison to the number of bacteria in 2007. Most of these microorganisms were found in samples containing fluidal ash, sewage sludge, straw and microbiological formulations. The number of actinomycetes also increased in comparison with the first year of study in all examined soil and ash samples. The largest, statistically significant, increase in the number of these bacteria was observed in samples with the addition of sewage sludge, straw and microbiological formulations: EM-1 and EM-X. The significant increase in the number of these microorganisms could be related to the antiacidic influence of the added ashes. The reaction is, in fact, one of the factors influencing the dynamics of the development of soil actinomycetes [7, 8]. The number of microscopic fungi in comparison to the year 2007 increased significantly in the control soil, in the soil with sewage sludge and straw and in the sample with fluidal ash, sediment, straw and microbiological formulations. In other trials the number of fungi, in most cases, was lower in comparison to the amount measured in the second year of the experiment. Successive Influence of Fluidal Ashes on General Number of Bacteria... 1717

250 2007 2008 2009 c c d 200 b b cd bcd 150 b abc d abc ab 100 a a

6 a a

10 c.f.u./g d.m. of soil 50 cd c d b f a ab e 0 12345678LSD0.05 Variants of experience

Fig. 1. The number of bacteria in examined samples of soil and fluidal ash Variants of experiments: 1 – control (soil), 2 – control (fluidal ash), 3 – soil + sewage sludge + straw (4:2:1), 4 – fluidal ash + sewage sludge + straw (4:2:1), 5 – soil + sewage sludge + straw (4:2:1) + microbial formulation EM-1 (15 dm3 × ha–1), 6 – fluidal ash + sewage sludge + straw (4:2:1) + microbial formulation EM-1 (15 dm3 × ha–1), 7 – soil + sewage sludge + straw (4:2:1) + microbial formulation EM-1 (15 dm3 × ha–1) + EM-X Ceramic Powder (40 dm3 × ha–1), 8 – fluidal ash + sewage sludge + straw (4:2:1) + microbial formulation EM-1 (15 dm3 × ha–1) + EM-X Ceramic Powder (40 dm3 × ha–1)

300 2007 2008 2009 ab 250

200 c bc a a abc abc 150 ab ad ad a d 100

4 a a

10 c.f.u./g d.m. of soil b a 50 c b a a a a a a 0 12345678LSD0.05 Variants of experience

Fig. 2. The number of actinomycetes in examined samples of soil and fluidal ash Explanations: see Fig. 1.

The number of coli bacteria, as an indicator of sanitary contamination, ranged from about 250 to 1200 in the first year of studies, up to about 2–20 cells in 1 g d.m. of soil and fluidal ash sample in the third year of research (Fig. 4). Compared with the control soil, statistically most of these bacteria were observed in the soil with sewage sludge and straw and in the combination of soil, sewage sludge, straw and microbiological formulation EM-X. The statistically lowest content of coli occurred in the control soil and fluidal ash. Summarizing the results of three-year studies on the fitness of fluidal ashes for agricultural purposes and soil recultivation, it should be noted that the addition of 1718 Wiera Michalcewicz et al

30 c 2007 2008 2009

ad a 20 bc ab a a c bc bc c 10 d e 4 ab ad abc ab a b ab 10 c.f.u./g d.m. of soil ab ab a abc

0 12345678LSD0.05 Variants of experience

Fig. 3. The number of fungi in examined samples of soil and fluidal ash Explanations: see Fig. 1.

c 5000 2007 2008 2009 4000 b b 3000

2000

e a a

1 c.f.u./g d.m. of soil a 1000 a d ad a a a c b b abc d a d a bc d 0 ab 12345678LSD0.05 Variants of experience

Fig. 4. The number of coli bacteria in examined samples of soil and fluidal ash Explanations: see Fig. 1. organic materials, such as composted sewage sludge, which is rich in microorganisms or microbial preparations EM-1 or EM-X to ashes or soils with ashes, caused an statistically significant increase in the number bacteria and actinomycetes. Studies by other authors [9–11] confirm the stimulative effect of fermented sewage sludge on the number and activity of many groups of microorganisms. Also the addition of microbiological formulations EM-1 or EM-X, caused in particular the increase in the number of bacteria and actinomycetes. As indicated in a few studies, the EM formulations, containing several strains of microorganisms isolated from soil, classified in different taxons, and having different physiological and biochemical features [12] have demonstrated the beneficial influence both on the functioning of a soil ecosystem and the intensity of growth and development of plants. The results of conducted studies indicate that the addition of effective microorganisms to soil or fluidal ash increased the number of microorganisms, and hence, caused the intensification of metabolic and biochemical processes in soil, and, indirectly, affected pedogenetic abilities of micro- Successive Influence of Fluidal Ashes on General Number of Bacteria... 1719 organisms. Enrichment of ash in the microbiological formulation (effective micro- organisms) accelerated the mineralization of soil organic compounds and thereby increased the biological activity of soil [13, 14]. The obtained results indicate that coal ashes can be used for fertilizing purposes.

Conclusion

1. Compared with the first year of the experiment an increase in the overall number of bacteria and actinomycetes in soil and ash samples was found. 2. There was a significant decrease in the number of coli bacteria in comparison to the first year of studies.

References

[1] Barabasz W., Szmigiel A., Albiñska D. and Barabasz J.: Wp³yw nawo¿enia osadami œciekowymi na mikroflorê ryzosfery pod upraw¹ wierzby energetycznej. Gospodarka odpadami, K. Szymañski (ed.), Monografia, t. 3, Komitet Chemii Analitycznej PAN, Koszalin 2007, 141–148. [2] Higa T.: Effective microorganisms, concept and recent advances in technology. Proc. Conference on Effective Microorganisms for a Sustainable Agriculture and Environment. 4th Int. Conf. on Kyusei Nature Farming, Bellingham – Washington, USA 1998, 247–248. [3] Bunt J.S. and Rovira A.D.: J. Soil. Sci. 1955, 6, 119–128. [4] Cyganov V.A., ukov R.A. and Timofeleva K.A.: Mikrobiologija 1964, 33(5), 863–869. [5] Martin J.P.: Soil Sci. 1950, 6, 215–217. [6] Burbianka M. and Pliszka A.: Mikrobiologia ¿ywnoœci. PWN, Warszawa 1971. [7] Kurek E.: Zesz. Probl. Post. Nauk Roln. 2002, 482, 307–316. [8] Wolna-Maruwka A. and Pilarski K.: Ochr. Œrod. Zasob. Natural. 2010, 42, 212–224. [9] Furczak J. and Joniec J.: Polish J. Soil Sci. 2002, 35, 59–67. [10] Nowak A. and B³aszak M.: Zesz. Nauk. UP we Wroc³awiu 2006, 546, 271–278. [11] Wolna-Maruwka A., Sawicka A. and Sty³a K.: Zmiany liczebnoœci wybranych grup drobnoustrojów w glebie o zró¿nicowanym nawo¿eniu organicznym, VIII Ogólnopolskie Symp. Nauk.-Techn. “Bio- technologia Œrodowiskowa”, Wis³a-Jarzêbata 2005, 41–50. [12] Mau P.: Fantastische Erfarungen mit EM, Publ. EMIKO, Germany 2003. [13] Bieliñska E. and Baran S.: In¿. Roln. 2009, 6(115), 7–15. [14] Bieliñska E., Stankowski S. and Wêgorek T.: Zastosowanie testów automatycznych do oceny mo¿liwoœci przyrodniczego wykorzystania popio³ów fluidalnych z wêgla kamiennego. Mat. Miêdzynarod. Konf. Nauk.-Techn., S³awsko 16–18 IV 2008.

NASTÊPCZY WP£YW POPIO£ÓW FLUIDALNYCH NA OGÓLN¥ LICZBÊ BAKTERII, PROMIENIOWCÓW I GRZYBÓW W BADANIACH WAZONOWYCH

1 Zak³ad Mikrobiologii i Biotechnologii Œrodowiska 2 Katedra Agronomii 3 Zak³ad Chemii Ogólnej i Ekologicznej Zachodniopomorski Uniwersytet Technologiczny w Szczecinie

Abstrakt: Celem przeprowadzonych badañ by³o okreœlenie nastêpczego wp³ywu popio³ów fluidalnych z wêgla kamiennego w po³¹czeniu z przefermentowanym osadem œciekowym i s³om¹, przy u¿yciu efektywnych mikroorganizmów, na ogóln¹ liczebnoœæ bakterii, grzybów, promieniowców oraz bakterii z grupy coli. Badania prowadzono w warunkach wazonowych. Stwierdzono wzrost liczebnoœci bakterii 1720 Wiera Michalcewicz et al i promieniowców w próbkach zawieraj¹cych popió³ fluidalny oraz ró¿ne komponenty organiczne w po- równaniu z pierwszym rokiem badañ. Natomiast liczebnoœæ bakterii z grupy coli w porównaniu z dwoma pierwszymi latami doœwiadczenia uleg³a istotnemu zmniejszeniu. S³owa kluczowe: popió³ fluidalny, bakterie, grzyby, promieniowce, bakterie z grupy coli ECOLOGICAL CHEMISTRY AND ENGINEERING A

Vol. 18, No. 12 2011

Katarzyna GLEÑ

EFFECT OF FOLIAR FERTILIZERS AND THEIR MIXTURES ON PHYTOPATHOGENIC FUSARIUM FUNGI

WP£YW NAWOZÓW DOLISTNYCH I ICH MIESZANIN NA GRZYBY FITOPATOGENNE Z RODZAJU FUSARIUM

Abstract: The paper focuses on the response of phytopathogenic Fusarium fungi to various concentrations of foliar fertilizers: Mikrovit Fe, Mikrovit Zn, urea, magnesium sulphate, and the mixtures of Mikrovit Fe + urea + magnesium sulphate, and Mikrovit Zn + urea + magnesium sulphate added to the medium. Under in vitro conditions, the analysis determined the influence of the foliar fertilizers on linear growth, biomass increment and sporulation of the following fungi: Fusarium poea, Fusarium sulphureum and Fusarium culmorum. Mikrovit Zn revealed the strongest fungistatic properties among the tested foliar fertilizers. Applied to the medium in 1.0 mm3/cm3 concentration, it very strongly inhibited the linear growth (91.93–94.17 %) and sporulation of all tested fungi and most strongly limited biomass increments in F. poea and F. sulphureum, whereas mixtures of Mikrovit Zn and Mikrovit Fe with urea and magnesium sulphate revealed slightly weaker fungistatic effect. Urea applied in 1.0 mm3/cm3 concentration reduced increments of the test fungi biomass in the range from 56.73 to 64.03 %, while magnesium sulphate, as the only one among the fertilizers used for the experiment, stimulated surface growth, biomass increment and sporulation process in all tested fungi. It should be remembered that in the agrocenoses the effect of foliar fertilizers on fungi infecting plants is more complex and conditioned by many factors. Therefore, it is necessary to conduct further research on the influence of foliar application of fertilizers on plant healthiness. Keywords: Fusarium, foliar fertilizers, linear growth, biomass, sporulation

Among phytopathogenic organisms fungi of Fusarium genus deserve special attention, since they are the most common in the environment where they infect many plant species causing various diseases which are difficult to control, such as fusarium wilts, take-all diseases or dry rot [1–2]. In the opinion of many authors [3–5] fusarioses are particularly dangerous in cereal crops since they not only contribute to yield losses, but infect grains with mycotoxins. Chemical plant protection is the most efficient method to combat these diseases, but it may negatively affect the quality of raw plant materials and pollute the environment. Several authors reported that increasingly

1 Department of Agricultural Environment Protection, University of Agriculture in Krakow, al. A. Mic- kiewicza 21, 31–120 Kraków, Poland, phone: +48 12 662 44 00, email: [email protected] 1722 Katarzyna Gleñ common foliar fertilization not only positively affects the crop yield and its quality but may also efficiently prevent and protect plants against infectious diseases [6–12]. The investigations were conducted to test whether commonly applied foliar fertilizers Mikrovit Fe, Mikrovit Zn, magnesium sulphate, urea and their mixtures (Mikrovit Fe + magnesium sulphate + urea, and Mikrovit Zn + magnesium sulphate + urea) may limit the development of the following phytopathogenic fungi: Fusarium poea (Peck) Wollenw., Fusarium culmorum (W.G. Smith) Sacc. and Fusarium sulphureum Schlecht. under in vitro conditions.

Materials and methods

The fertilizers were tested on the phytopathogenic fungi selected from the collection owned by the Agricultural Environment Protection Department: Fusarium poea, Fusarium culmorum and Fusarium sulphureum, isolated from infected wheat kernels. Each of the studied fertilizers, ie Mikrovit Fe (Fe–3%,N–4.50 %, pH – 3.2), Mikrovit Zn (N – 4.50 %, Zn – 3.5 %, pH – 2.0), magnesium sulphate (Mg – 15.65 %, S – 17.20 %, pH – 6.8) and urea (N – 46 %) and Mikrovit Fe + magnesium sulphate + urea, and Mikrovit Zn + magnesium sulphate + urea) were added to PDA medium with pH = 6.32 to obtain their medium concentrations of 0.1 mm3/cm3 (field dose) and 1.0 mm3/cm3. Subsequently, the tested fungus inoculum was supplied to Petri dishes containing the consolidated medium with added tested fertilizers. The experiment was conducted in five replications for each fertilizer combination and for each individual tested fungus. Petri dishes with the medium without fertilizers provided the control. The fungi were cultured in a thermostat at 23 oC. Daily increments of the fungi colonies served to compute the coefficient of the tested fungi linear growth rate in each fertilizer combination and on the control.

A b b T =+1 ++... x D d1 d x where: T – linear growth rate coefficient, A – mean of colony diameter measurements [mm], D – number of days since the experiment outset, b1, ..., bx – increment of colony diameter since the last measurement [mm], d1, ..., dx – number of days since the last measurement. After three weeks of fungi culturing on PDA media with added foliar fertilizers and on the control, the number of spores was assessed in the Thome hemocytometer. The fungi biomass growth was maintained in 300 cm3 Erlenmayer flasks on 100 cm3 of the modified PDA medium (without agar-agar) with the foliar fertilizers added in the same concentrations as in the experiment on Petri dishes. The effect of individual foliar fertilizers on the linear growth of the studied phytopathogenic fungi was presented as a difference between fungus colony diameter on the control Petri dishes and fungus colony diameter on the Petri dishes with individual fertilizer concentrations, and converted into the inhibition-stimulation coefficient acc. to Abbot [3]. Biomass Effect of Foliar Fertilizers and Their Mixtures on Phytopathogenic Fusarium Fungi 1723 increments were assessed in the same way. The results were elaborated statistically using ANOVA and the significance of differences was assessed by means of the t-Student test.

Results and discussion

For their development, fungal organisms need macroelements and microelements which they use for hyphae formation and production of numerous biologically active compounds and enzymes [14–15]. Conducted experiments allowed to determine that surface growth, biomass increment and sporulation of the tested fungus species were modified by the kind of foliar fertilizer and its concentration. Moreover within the same Fusarium genus, the tested fungi species differ with their sensitivity to the applied fertilizer preparations. The fact was confirmed by the Authors’ and other previous investigations [16–21]. Among the tested foliar fertilizers, Mikrovit Zn applied in 1.0 mm3/cm3 concentration revealed the strongest fungistatic effect, almost completely (91.94–94.12 %) inhibiting the linear growth and blocking sporulation process in all tested fungal organisms (Tables 1, 3).

Table 1

Coefficients of rate (T) and inhibition of tested fungi linear growth

Concentration F. poea F. sulphureum F. culmorum Foliar fertilizers 3 3 [mm /cm ] [T] [%] [T] [%] [T] [%] 1.0 23.77 56.74 18.45 64.03 21.33 61.20 Urea 0.1 55.45 6.35 44.80 20.79 52.28 16.80 1.0 73.06 +5.07* 81.01 +26.02+ 49.44 21.17 Magnesium sulphate 0.1 60.40 1.98 49.77 16.07 89.50 +38.30+ 1.0 79.48 0.95 39.80 26.70 61.46 15.02 Mikrovit Fe 0.1 80.60 +0.43 70.98 20.50 77.30 +11.02+ 1.0 1.00 94.17 1.00 91.93 1.55 94.18 Mikrovit Zn 0.1 61.29 35.94 62.90 +6.05 37.70 43.22 Mikrovit Fe + urea + 1.0 9.95 84.17 24.16 54.43 6.10 86.75 magnesium sulphate 0.1 59.58 31.18 63.09 +9.69 32.64 49.65 Mikrovit Zn + urea + 1.0 29.68 58.90 29.8 41.90 10.58 80.46 magnesium sulphate 0.1 80.10 0.20 52.1 8.62 34.03 48.34 Control 80.25 59.63 70.75

LSD0.05 for fertilizer kind 1.67 2.11 2.31

LSD0.05 for fertilizer concentration 1.13 1.14 1.24 * + denotes linear growth stimulation.

On the other hand inhibition of F. culmorum and F. sulphureum biomass growth reached 67.88 % and 68.83 %, respectively (Table 2). 1724 Katarzyna Gleñ

Table 2 Coefficient of biomass increment coefficient depending on foliar fertilizer [%]

F. poea F. sulphureum F. culmorum Foliar fertilizers Concentration [mm3/cm3] 1.0 0.1 1.0 0.1 1.0 0.1 Urea 44.60 5.35 53.25 41.48 22.92 10.91 Magnesium sulphate 17.85 +7.14 +17.46+ 4.36 +21.83+ +11.64+ Mikrovit Fe 73.21 73.21 46.58 58.22 52.40 55.31 Mikrovit Zn 91.07 55.35 68.33 39.30 67.88 50.22 Mikrovit Fe + urea + magnesium sulphate 69.64 75.00 60.00 56.77 66.96 64.05 Mikrovit Zn + urea + magnesium sulphate 73.21 70.53 56.77 53.31 71.32 65.50 + denotes biomass increment stimulation.

A strongly inhibiting effect on Fusarium fungi was also observed after the application of English foliar fertilizer Yeald to the medium, which similar as Mikrovit Zn contained zinc [21]. Combined application of Mikrovit Zn with urea and magnesium sulphate in the Authors’ own experiments more weakly affected the studied phyto- pathogenic fungi species. In the conducted experiment magnesium sulphate was the only fertilizer whose supplement in the medium, particularly in the higher concentration (1.0 mm3/cm3), stimulated hyphae surface growth and biomass increment in all test fungi species. Moreover it favoured sporulation, primarily in F. poea and F. culmorum (Table 3).

Table 3 The impact of foliar fertilizers on test fungi sporulation (quantity in 1 cm3 ×106)

F. poea F. sulphureum F. culmorum Foliar fertilizers Concentration [mm3/cm3] 1.0 0.1 1.0 0.1 1.0 0.1 Urea 0.16 0.30 8.96 2.78 1.10 0.59 Magnesium sulphate 12.04 17.23 1.55 3.24 2.10 2.80 Mikrovit Fe 0.30 0.35 13.87 1.52 14.60 3.27 Mikrovit Zn — 0.22 — 1.55 — 2.12 Mikrovit Fe + urea + magnesium sulphate 0.05 0.20 0.15 0.4 0.87 2.30 Mikrovit Zn + urea + magnesium sulphate 3.00 0.10 0.17 2.45 0.30 1.00 Control 9.90 2.17 1.72

Therefore, it may be assumed that the availability in the medium of other elements, such as magnesium or sulphur alleviates the fungistatic effect of zinc. On the other hand, on media with added 1.0 mm3/cm3 (field dose) of urea, limited surface growth of the tested fungi in the range from 56.73 % to 64.03 % and their biomass increment (22.93–53.255) were observed (Tables 1, 2). Irrespective of which concentration of this Effect of Foliar Fertilizers and Their Mixtures on Phytopathogenic Fusarium Fungi 1725 fertilizer was applied, a considerable reduction of the produced spore number was observed in F. poea and F. culmorum (Table 3), as well as modified colour of aerial mycelium (naturally pink became white). Mikrovit Fe revealed weak fungistatic properties, particularly in a short experiment on Petri dishes. No matter which concentration was applied, this fertilizer limited the linear growth of F. sulphureum by 23.6 % and F. culmorum only by 2.0 %. A variable effect of iron on Fusarium fungi was demonstrated also by other authors [16, 22], since iron may inhibit growth of fungal pathogens at limited element availability and stimulate at the element excess in the environment. On the other hand, Mikrovit Fe mixture with urea and magnesium sulphate had stronger effect, which may have resulted from accumulation of the fungistatic properties of its components (Tables 1–3). The number of F. sulphureum macroconidia on the medium containing 1.0 mm3/cm3 of this fertilizer mixture was thirteen times lower than in the control (Table 3), while the components of this mixture, ie urea and Mikrovit Fe applied separately in 1.0 mm3/cm3 concentration, strongly stimulated spore formation in F. sulphureum. The total number of spores produced by the phytopathogens evidences their infection potential, therefore, at reduced spore number the risk of plant infection diminishes [14].

Conclusions

Strong fungistatic properties, particularly of Mikrovit Zn and its mixtures with commonly used fertilizers, ie urea and magnesium sulphate and urea used separately, as presented in the paper, may find applications in the agricultural practice. It should be expected that application of these foliar fertilizers in cultivation of plants requiring zinc feeding, ie vegetables and fruit trees may result in less frequent occurrence of fungal diseases, especially when they are caused by fungi of Fusarium genus. Moreover, fungistatic properties of foliar fertilizers may undoubtedly contribute to reduce the amount of herbicides used for plant protection. Therefore, it is necessary to undertake research on the effect of foliar fertilizers on plant healthiness.

References

[1] Cook R.J.: Fusarium: Diseases, biology and taxonomy, The Pennsylvania State University Press, Park–London 1981. [2] £awecki T.: Agrotechnika 2005, 5(521), 28–30. [3] Che³kowski J.: Hodow. Roœl. Nasien. 1995, 4, 26–27. [4] Tomczak M., Che³kowski J., Kostecki M., Wiœniewska H. and Goliñski P.: Materia³y X Konferencji: Grzyby mikroskopowe – badania genetyczne i molekularne nad patogenami roœlin i ich metabolitami, Poznañ 2000. [5] Jañczak C.: Ochr. Roœl. 2006, 51(8), 35–37. [6] Nowosielski O.: Ochr. Roœl. 1986, 7, 14–16. [7] £aszcz E., Irzyk M., Klimach A. and Wieczorek W.: Mat. Symp.: Nowe kierunki w fitopatologii, Kraków 1996, pp. 99–102. [8] Bolig³owa E.: Acta Agrophys. 2003, 85, 99–106. [9] Chwil S. and Szewczuk C.: Acta Agrophys. 2003, 85, 117–124. [10] Magdziak R. and Ko³odziej B.: Acta Agrophys. 2003, 85, 319–329. 1726 Katarzyna Gleñ

[11] Jab³oñski K.: Acta Agrophys. 2003, 85, 137–143. [12] Gleñ K.: Ecol. Chem. Eng. A. 2008, 15(4–5), 331–336. [13] Kowalik R. and Krechniak E.: Szczegó³owa metodyka biologicznych i laboratoryjnych badañ œrodków grzybobójczych, [in:] Materia³y do metodyki badañ biologicznej oceny œrodków ochrony roœlin, IOR, Poznañ 1961. [14] Kirlay Z., Klement Z. and Solymosy F.: Fitopatologia. Wybór metod badawczych. PWRiL, Warszawa 1977. [15] PrzeŸdziecki Z., Wojciechowska-Kot H., Miko³ajska I. and Murawa D.: Acta Acad. Agricult. Tech., Olsztyn 1991, 53, 229–239. [16] Chmiel M.J.: Chem. In¿. Ekol. 2006, 13(1–2), 11–15. [17] Gleñ K. and Bolig³owa E.: Chem. In¿. Ekol. 2006, 13(1–2), 29–36. [18] Gleñ K. and Bolig³owa E.: Chem. In¿. Ekol. 2007, 14(9), 933–939. [19] Kotliñska T.: Mat. 28 Sesji Naukowej IOR, Poznañ 1988, pp. 293–297. [20] Gleñ K.: Ecol. Chem. Eng. A. 2008, 15(1–2), 47–54. [21] Bolig³owa E. and Gleñ K.: Acta Agrophys. 2003, 85, 107–116. [22] Gleñ K. and Bolig³owa E.: Chem. In¿. Ekol. 2006, 13(8), 743–749.

WP£YW NAWOZÓW DOLISTNYCH I ICH MIESZANIN NA GRZYBY FITOPATOGENNE Z RODZAJU FUSARIUM

Katedra Ochrony Œrodowiska Rolniczego, Wydzia³ Rolniczo-Ekonomiczny Uniwersytet Rolniczy im. Hugona Ko³³¹taja w Krakowie

Abstrakt: Badano reakcjê grzybów chorobotwórczych z rodzaju Fusarium na dodatek do pod³o¿a hodowlanego ró¿nych stê¿eñ nawozów dolistnych: Mikrovit Fe, Mikrovit Zn, mocznik, siarczan magnezu oraz mieszanin: Mikrovitu Fe + mocznik + siarczan magnezu i Mikrovitu Zn + mocznik + siarczan magnezu. W warunkach in vitro oceniano wp³yw nawozów dolistnych na wzrost liniowy, przyrost biomasy i zarod- nikowanie grzybów: Fusarium poea, Fusarium sulphureum i Fusarium culmorum. Spoœród badanych nawozów dolistnych Mikrovit Zn odznacza³ siê najsilniejszymi w³aœciwoœciami fungistatycznymi. Zaaplikowany do pod³o¿a hodowlanego w stê¿eniu 1.0 mm3/cm3 bardzo silnie hamowa³ rozrost powierzchniowy (91.93–94.17 %) i zarodnikowanie wszystkich testowanych grzybów oraz najsilniej ogranicza³ przyrost biomasy F. poea i F. sulphureum. Natomiast nieco s³absz¹ efektywnoœæ fungistatyczn¹ wykazywa³y mieszaniny: Mikrovitu Zn oraz Mikrovitu Fe z mocznikiem i siarczanem magnezu. Mocznik zastosowany w stê¿eniu 1.0 mm3/cm3 ogranicza³ przyrosty biomasy grzybów testowych w zakresie od 56.73 do 64.03 %. Z kolei siarczan magnezu, jako jedyny spoœród zastosowanych w doœwiadczeniu nawozów, stymulowa³ wzrost powierzchniowy, przyrost biomasy oraz proces sporulacji wszystkich grzybów testowych. Nale¿y pamiêtaæ, ¿e w agrocenozach oddzia³ywanie nawozów dolistnych na grzyby pora¿aj¹ce roœliny jest bardziej z³o¿one i uwarunkowane wieloma czynnikami. Dlatego istnieje potrzeba przeprowadzania badañ nad wp³ywem aplikacji nalistnej nawozów na zdrowotnoœæ roœlin. S³owa kluczowe: Fusarium, nawozy dolistne, wzrost liniowy, biomasa, zarodnikowanie ECOLOGICAL CHEMISTRY AND ENGINEERING A

Vol. 18, No. 12 2011

Ma³gorzata NABRDALIK1 and Katarzyna GRATA1

INFLUENCE OF THE CULTURE CONDITIONS ON LIPOLYTIC ACTIVITY OF Bacillus cereus AND Bacillus mycoides

WP£YW WARUNKÓW ŒRODOWISKA NA AKTYWNOŒÆ LIPOLITYCZN¥ SZCZEPÓW Bacillus cereus I Bacillus mycoides

Abstract: The aim of the research was the evaluation of lipolytic activity of B. cereus and B. mycoides strains, in reference to carbon source, pH and the temperature. In the research, two strains of Bacillus cereus and Bacillus mycoides each, isolated from the soil and water, were applied. The sources of carbon in culture media were fatty substrates: tributyrin, Tween 40, Tween 60, Tween 80 and glucose. The lipolytic activity was measured by means of titration at pH ranging from 5 to 8 and the temperature ranging from 30 oC to 60 oC. The results were presented in the units of lipolytic activity [U cm–3]. In the conducted research, the amount of liberated lipolytic activity depended on the type of fatty substrate in the medium, pH and the temperature. The strains under study showed the lowest activity at pH 5 and 6, and the highest at pH 7 and 8. In these conditions, most of the strains showed the lipolytic activity, even in case of the lack of fatty substrate in the medium. The highest amount of lipolytic activity was liberated at pH 8 in the medium with Tween 40, and the highest results (0.88 [U cm–3]) were noted for the soil strain B. cereus. When analysing the influence of the temperature on the lipolytic activity, it was stated that the highest amount of lipolytic activity was noted at 30 and 40 oC, and the lowest at 50 and 60 oC. The best results were obtained for most of the strains at 30 oC, in medium with Tween 40, and the most active was the soil strain of B. mycoides (0.88 [U cm–3]). The exception is B. cereus, as it liberated 1.38 [U cm–3], in the medium with glucose. Taking into account all analysed sources of carbon and parameters, it seems that the most active were B. mycoides strains. Keywords: Bacillus cereus, Bacillus mycoides, lipases, tributyrin, Tween

Lipases are an important group of biotechnologically valuable enzymes. They are defined as hydrolases of glycerol esters EC 3.1.1.3, and are the enzymes of high catalytical potential. They are produced by plants, animals and microorganisms, of which the last group remains in the centre of attention. Many kinds of bacteria possess

1 Department of Biotechnology and Molecular Biology, University of Opole, ul. kard. B. Kominka 6, 45–035 Opole, Poland, phone: +48 77 401 60 56, email: [email protected] 1728 Ma³gorzata Nabrdalik and Katarzyna Grata the ability to produce them, among others bacteria of Bacillus kind [1, 2]. A common interest in bacterial lipases is connected with their role as biocatalysts in many biochemical processes. They have diverse applications in a wide variety of industries ranging from detergent, oleochemical, organic synthesis, dairy, fat and oil modification to pharmaceutical. They have many applications for stereospecific hydrolysis and synthesis of a wide variety of technologically valuable esters [1, 2]. The interest in microbial lipase production has risen in the last decades, because of its large potential in environmental protection as wastewater treatment as well as decomposition and removal of oil substances. Effective breakdown of solids and the clearing and prevention of fat blockage or filming in waste systems are important in many industrial operations. Bacterial lipases are also involved in solution of such environmental problems as the breakdown of fats in domestic sewage and anaerobic digesters [3]. As shown by data in literature [2, 4–6], they are varied in terms of their enzymatic activity, which depends on the species of microbes and the culturing conditions (eg pH of the growth medium, temperature, source of nitrogen and presence of lipids in the medium). Therefore, screening of microorganisms with lipolytic activities could facilitate the discovery of novel lipases. The aim of undertaken research was the evaluation of lipolytic activity of Bacillus cereus and Bacillus mycoides, isolated form the natural environment, in reference to carbon source, pH and the temperature.

Materials and methods

The object of the study were 4 Bacillus strains: –2Bacillus cereus strains marked as: A96 and G10, isolated from soil and water, respectively, –2Bacillus mycoides strains marked as: A134 and G3, isolated from soil and water, respectively. The inoculum was produced in the basal medium consisted of 1.0 g × dm–3 yeast × –3 × –3 × –3 extract, K2HPO4 3.0 g dm ,KH2PO4 2.0 g dm , (NH4)2SO4 2.0 g dm and × × –3 MgSO4 7H2O 0.5 g dm . The sources of carbon in culture media were the following fatty substrates: tributyrin, Tween 40, Tween 60, Tween 80 and glucose. The cultures were maintained in Erlenmeyer flasks of 250 cm3 capacity containing 50 cm3 of respective growth medium with an inoculum of density equal to E = 2, obtained from the 48-hour culture on a nutrient broth. Incubation was conducted on a rotary shaker for 2 days at 30 oC. Samples were collected after 2 days of culturing and centrifugated for 20 min at 4000 rpm. The extracellular lipolytic activity was marked in the obtained supernatant by means of titration towards the same substrates as the ones added to the growth media (the proper treatment). In the control treatment the supernatant was replaced with water. Lipolytic activity was estimated at pH ranging from 5 to 8, and at temperature ranging from 30 to 60 oC. The amount of liberated fatty acids was determined by titration with 0.05 M NaOH solution against 2 % phenolophthalein as an indicator, and calculated as Influence of the Culture Conditions on Lipolytic Activity of Bacillus cereus... 1729 a subtraction between the proper treatment and the control treatment results. The results were presented in the units of lipolytic activity. The unit was expressed as the amount of μmoles of 0.05 M NaOH required to neutralize fatty acids liberated by the lipases contained in 1 cm3 of post-culture liquid within 1 minute. The lipolytic activity was expressed in the unit U cm–3.

Results and discussion

In the presented paper, 4 bacterial strains of Bacillus kind were screened for their ability to synthesize lipolytic enzymes on culture media containing different source of carbon, at pH ranging from 5 to 8 and the temperature ranging from 30 to 60 oC. In conducted tests, lipolytic activity depended on the carbon source in the medium, pH and the temperature, while individual B. cereus and B. mycoides strains showed varied activity in exocellular lipases production. For the soil strain B. cereus A96, a reverse correlation between their lipolytic activity and the temperature has been observed (Fig. 1) and it has been stated that the temperature growth caused a decrease in lipolytic activity U. The highest values were obtained at 30 oC in the presence of glucose as carbon source – 1.38 U cm–3, and the lowest at 60 oC in the medium of Tween 80 and glucose – 0.13 U cm–3. The highest, 10-fold, decrease in the lipolytic activity, has been recorded on the medium with glucose, while Tween 40 seemed to be the most stable and favourable source of the fatty substrate for B. cereus A96 (Fig. 1).

1.6 tributyrin 1.4 Twenn 40

–3 1.2 Twenn 60 1.0 Twenn 80 glucose 0.8 0.6

0.4

Lipolytic activity [U cm ] 0.2 0.0 30 40 50 60 Temperature [oC]

Fig. 1. Influence of temperature on the lipolytic activity of Bacillus cereus A96

In the presented paper, there has been no clear relationship found between the lipolytic activity and the temperature in case of B. cereus G10 strain isolated from water (Fig. 2). However, the highest activity has been recorded at 30 oC and the lowest at 60 oC, regardless of the carbon source. The most favourable source of the fatty substrate was Tween 40, where the highest values of the activity have been noted, regardless of the temperature. The highest value of 0.71 U cm–3 was noted at 30 oC. The least favourable source of carbon was Tween 80 and glucose, as irrespective of the 1730 Ma³gorzata Nabrdalik and Katarzyna Grata

1.6 tributyrin 1.4 Twenn 40

–3 1.2 Twenn 60 1.0 Twenn 80 glucose 0.8 0.6

0.4

Lipolytic activity [U cm ] 0.2 0.0 30 40 50 60 Temperature [oC]

Fig. 2. Influence of temperature on the lipolytic activity of Bacillus cereus G10 temperature, obtained results were the lowest. Also, the activity of exocellular hydrolases has not been noticed at 50 oC and 60 oC in case of Tween 80, and at 60 oCin case of medium containing glucose (Fig. 2). Growing temperature did not promote exocellular production of lipases in case of B. mycoides A134 and G3 strains (Fig. 3 and 4). The exception has been noted for B. mycoides A134 strain on the medium with glucose, where the lipolytic activity grows from the initial value of 0.83 U cm–3 at 30 oCto 1.46 U cm–3 at 40 oC, to reach the value of 0.46 U cm–3 at the temperature 60 oC (Fig. 3).

1.6 tributyrin 1.4 Twenn 40

–3 Twenn 60 1.2 Twenn 80 1.0 glucose 0.8 0.6

0.4

Lipolytic activity [U cm ] 0.2 0.0 30 40 50 60 Temperature [oC]

Fig. 3. Influence of temperature on the lipolytic activity of Bacillus mycoides A134

The most favourable medium for B. mycoides A134 strain, among media containing fatty substrate as the source of carbon, is medium with Tween 40. Recorded values of the lipolytic activity were the highest at all temperatures, when compared with other fatty substrates. The lipolytic activity at 30 oC amounted 0.88 U cm–3 and decreased slightly to the value of 0.83 U cm–3 at 60 oC. The least favourable fatty substrate was Tween 80, as at 30 oC measured activity amounted only 0.33 U cm–3, and at 50 oC the strain did not show exocellular activity of hydrolases (Fig. 3). Influence of the Culture Conditions on Lipolytic Activity of Bacillus cereus... 1731

In case of B. mycoides G3 strain the most favourable was the medium with Tween 40, similarly to B. mycoides A134 (Fig. 4). The highest values of the lipolytic activity have been obtained in the temperature ranging between 30 and 60 oC and amounted 0.83–0.67 U cm–3. Similar changes, to the aforementioned, have been noted in reference to the medium with glucose, but recorded values of the lipolytic activity have been significantly lower. At the temperature of 30 oC the lipolytic activity amounted 0.58 U cm–3, grew up to the value of 0.71 U cm–3 at 40 oC and reached the value of 0.33 U cm–3 at the highest temperature. The lowest difference in the lipolytic activity, at respective temperatures, have been obtained for the media with the following fatty substrates: tributyrin, Tween 40 and Tween 60 (Fig. 4).

1.6 tributyrin 1.4 Twenn 40

–3 1.2 Twenn 60 1.0 Twenn 80 glucose 0.8 0.6

0.4

Lipolytic activity [U cm ] 0.2 0.0 30 40 50 60 Temperature [oC]

Fig. 4. Influence of temperature on the lipolytic activity of Bacillus mycoides G3

Presented paper considers also the influence of pH on the lipolytic activity of tested B. cereus and B. mycoides strains, with reference to the source of carbon in the culture medium. Some correlation has been observed in most cases, regardless to the strain tested and the culture medium, namely pH increase in the range between 5–8 promotes the increase in the lipolytic activity (Figs. 5–8).

1.6 tributyrin 1.4 Twenn 40

–3 1.2 Twenn 60 Twenn 80 1.0 glucose 0.8 0.6

0.4

Lipolytic activity [U cm ] 0.2 0.0 5678 pH

Fig. 5. Influence of pH on the lipolytic activity of Bacillus cereus A96 1732 Ma³gorzata Nabrdalik and Katarzyna Grata

The most favourable culture media for the soil strain B. cereus A96 are those containing tributyrin and Tween 40, as the source of the fatty substrate. The gradual growth of the lipolytic activity from the initial amount of 0.29 U cm–3 at pH 5, up to the amount of 0.71 U cm–3 at pH 8 was observed in case of tributyrin. The lipolytic activity on Tween 40 was initially lower, when compared with tributyrin, and finally obtained the highest recorded value for the strain under study, equal to 0.88 U cm–3 at pH 8. B. cereus A96 did not show the lipolytic activity at pH between 5–6 on the culture medium with glucose. It was recorded only at pH 7 and amounted 0.13 U cm–3, growing up to the value of 0.63 U cm–3 at pH 8 (Fig. 5). In case of B. cereus G10 strain, the most effective source of carbon in the process of exocellular lipases biosynthesis, were also fatty substrates under study (Fig. 6), for which recorded values of the lipolytic activity did not differ significantly at pH range between 5–7. The differences were observed at pH 8, where the highest value of 0.71 Ucm–3 was obtained in the medium with Tween 40, and the lowest value of 0.38 Ucm–3 in the medium with Tween 60. Similarly to the soil strain, the lipolytic activity was not recorded if the medium did not contain the fatty substrate and glucose was the source of carbon. Such case was observed at pH between 5–6. At pH 7 and 8 measured values of the activity amounted 0.17 and 0.46 U cm–3 respectively (Fig. 6).

1.6 tributyrin 1.4 Twenn 40

–3 1.2 Twenn 60 1.0 Twenn 80 glucose 0.8 0.6

0.4

Lipolytic activity [U cm ] 0.2 0.0 5678 pH

Fig. 6. Influence of pH on the lipolytic activity of Bacillus cereus G10

In own research, when analysing the influence of pH on the lipolytic activity, it has been noted that bacterial strains B. mycoides A134 and G3 preferred different sources of fatty substrates in comparison with B. cereus strains. The highest values of the lipolytic activity have been obtained on the culture media with addition of Tween 80, Tween 60 and Tween 40 (Fig. 7 and 8). B. mycoides A134 strain revealed the highest lipolytic activity at pH 5–7 on the medium with Tween 80, and the recorded values ranged between 0.21–0.54 U cm–3. However, in the presence of Tween 40 lipolytic activity of B. mycoides A134 was the highest and amounted 0.75 U cm–3. Influence of the Culture Conditions on Lipolytic Activity of Bacillus cereus... 1733

The strain did not show the ability to produce exocellular lipases at pH between 5–7, with no fatty substrate in the culture medium. Only at pH 8, the activity was noted at the level of 0.75 U cm–3 in case of the medium with glucose and it was one of the highest values obtained for the strain (Fig. 7).

1.6 tributyrin 1.4 Twenn 40

–3 1.2 Twenn 60 Twenn 80 1.0 glucose 0.8 0.6

0.4

Lipolytic activity [U cm ] 0.2 0.0 5678 pH

Fig. 7. Influence of pH on the lipolytic activity of Bacillus mycoides A134

The last strain under study – B. mycoides G3, preferred Tween 60 and Tween 80 as the sources of the fatty substrate (Fig. 8). The value of lipolytic activity on Tween 60, was growing from the amount of 0.54 U cm–3 atpH5to1.0Ucm–3 at pH 8, which was the highest value recorded in the experiment. In Tween 80 case, the initial value of lipolytic activity at pH 5 and 6 was slightly higher and amounted 0.63 U cm–3 and 0.75 Ucm–3, respectively. At pH 8, it amounted 0.92 U cm–3 and was a bit lower when compared to the amounts obtained on the medium with Tween 60. It seems, that B. mycoides G3 strain, as the only one among all tested, uses Tween 60 as the most effective source of the fatty substrate in the process of extracellular lipases biosynthesis. When analysing the influence of pH on the lipolytic activity of B. mycoides G3 strain, it has been noted, that it was the only active strain even if the medium did not

1.6 tributyrin 1.4 Twenn 40

–3 1.2 Twenn 60 Twenn 80 1.0 glucose 0.8 0.6

0.4

Lipolytic activity [U cm ] 0.2 0.0 5678 pH

Fig. 8. Influence of pH on the lipolytic activity of Bacillus mycoides G3 1734 Ma³gorzata Nabrdalik and Katarzyna Grata contain fatty substrate. The value of its lipolytic activity in the presence of glucose increased from the value of 0.17 U cm–3 atpH5to0.83Ucm–3 at pH 8 (Fig. 8). Lipase production is influenced by temperature, pH and medium composition. The process of lipases biosynthesis conducted by strains B. cereus and B. mycoides was the most intensive at the temperature ranging between 30–40 oC. Hasan et al [7], Shaoxin et al [8] and Alkan et al [9] have also found similar results in case of B. sp. FH5, B. cereus C71 and B. coagulans. For B. cereus C71 [8] the lipase was active at temperature ranging between 30–45 oC with its most intensive activity at 33 oC. Whereas, for B. coagulans [9] and B. sp. FH5 [7], maximum activity have been observed at 37 oC. The protein nature of enzyme under study means that pH will affect the ionization state of the amino acids which determines the primary and secondary structure of the enzyme and hence, controls its overall activity [10]. The optimum activity of lipase was observed at pH between 7.0–8.0. Isolates of B. sp. have been found to produce lipolytic enzymes under alkaline conditions [9]. Lipase from B. subtilis and B. licheniformis have been of particular interest because they exhibit optimal activity and stability at extreme alkaline pH values greater than 9.5 [11]. These enzymes, however are thermolabile. Another lipase produced by B. sp. RSJ-1 has shown maximum activity at pH between 8.0–9.0 [10]. These results are in contrast to those of lipase from the B. cereus and B. mycoides under study, which are thermotolerant but display maximum activity at moderate alkaline pH between 7.0–8.0. Hasan et al [7] and Alkan et al [9] found similar results from B. sp. FH5 and B. coagulans, respectively. Lipases were defined as the enzymes hydrolyzing long-chain acyglycerols (³ 10 carbon atoms). However, it is known that most of lipases are also active on short-chain fatty acid esterase [8]. In own research, the biosynthesis of exocellular lipases in most cases, was more effective in medium with fatty substrate than with the addition of glucose. This may prove, that the synthesis of these enzymes is induced by the lipids. In case of fatty substrates, the highest values of lipolytic activity have been noted mostly in the presence of Tween 40 (C10) as the source of carbon, and noted values of lipolytic activity were higher in case of B. mycoides strains. Similarly, lipase from B. cereus C71 [8] showed higher activity toward substrate with C12 than C16 and C18. These results are in contrast to the ones obtained for the lipase from B. sp. FH5 tested by Hasan et al [7]. In the present study it has been found that maximum lipase levels were obtained when Tween 80 (C18) was used as a source of lipid which served both as a carbon source and an inducer for a lipase production. A low lipase level in a medium with glucose has also been reported by Hasan et al [7]. Actually, the variation in enzyme production at different temperatures or pH values and medium composition resulted from bacterial strains specificity.

Conclusions

The research proved significant diversity of lipolytic activity of B. cereus and B. mycoides strains, towards the source of carbon, pH and the temperature analysed in the experiment. Based on the obtained results following conclusions were drawn: Influence of the Culture Conditions on Lipolytic Activity of Bacillus cereus... 1735

1. The highest amount of lipolytic activity were liberated by the strains at 30 oC and pH equal to 8. Under these conditions the most favourable medium was with the addition of Tween 40, as the source of fatty substrate. 2. Strains under study were active even if there was no fatty substrate in the growth medium. 3. The most active were B. mycoides strains. Additionally, because of its wide substrate specificity, the enzyme can be used not only for short-chain fatty acids (contained by tributyrin) but also for medium-chain fatty acids (contained in Tween 40), which will greatly broaden its environmental applications. 4. Individual strains of B. cereus and B. mycoides showed diversity in their lipolytic activity, which was influenced by the environment from which they were isolated.

References

[1] Mrozik A. Hubert-Kocurek K., Nowak, B. and £abu¿ek S.: Microbial lipases and their significance in the protection of the environment. Post. Mikrobiol. 2008, 47, 1, 43–50. [2] Sharma R., Chisti Y. and Banerjee U.Ch.: Production, purification, characterization, and applications of lipases. Biotechnol. Adv. 2001, 19, 627–662. [3] Hasan F., Shah A.A. and Hameed A.: Industrial applications of microbial lipases. Enzym. Microb. Tech. 2006, 39, 235–251. [4] Bradoo S., Saxena R. K. and Gupta R.: Two acidothermotolerant lipases from new variants of Bacillus spp.. World J. Microbiol. Biotechnol. 1999, 15, 87–91. [5] Ertuárul S., Dönmez G. and Takaç S.: Isolation of lipase producing Bacillus sp. from olive mill wastewater and improving its enzyme activity. J. Hazard. Mater. 2007, 149, 720–724. [6] Mrozik A., Hubert-Kocurek K. and £abu¿ek S.: Lipazy bakterii Pseudomonas i Burkholderia oraz ich wykorzystanie w biotechnologii. Post. Mikrobiol. 2006. 45, 1, 19–26. [7] Hasan F., Shah A.A. and Hameed A.: Influence of culture conditions on lipase production by Bacillus sp FH5. Ann. Microbiol. 2006, 56(3), 247–252 [8] Shaoxin C., Lili Q. and Bingzhao S.: Purification and properties of enantioselective lipase from a newly isolated Bacillus cereus C71. Process Biochem., 2007, 42, 988–994. [9] Alkan, H., Baysal Z., Uyar F. and Dogru M.: Production of lipase by a newly isolated Bacillus coagulans under solid-state fermentation using melon wastes. Appl. Biochem. Biotechnol. 2007, 136(2), 183–191. [10] Sharma R., Soni S.K., Vohra R.M., Gupta L.K., Gupta J.K.: Purification and characterization of a thermostable alkaline lipase from a new thermophilic Bacillus sp. RSJ-1. Process Biochem. 2002, 37, 1075–1084. [11] Nthangeni M. B., Patterton H. G., van Tonder A., Vergeer W. P. and Litthauer D.: Over-expression and properties of a purified recombinant Bacillus licheniformis lipase: a comparative report on Bacillus lipases. Enzym. Mikrob. Tech. 2001, 28, 705–712.

WP£YW WARUNKÓW ŒRODOWISKA NA AKTYWNOŒÆ LIPOLITYCZN¥ SZCZEPÓW Bacillus cereus I Bacillus mycoides

Samodzielna Katedra Biotechnologii i Biologii Molekularnej Uniwersytet Opolski

Abstrakt: Celem podjêtych badañ by³a ocena aktywnoœci lipolitycznej szczepów B. cereus oraz B. mycoides w zale¿noœci od Ÿród³a wêgla, pH oraz temperatury. Do badañ wykorzystano 2 szczepy Bacillus cereus oraz 2 szczepy Bacillus mycoides wyizolowane z gleby i wody. ród³em wêgla w po¿ywkach by³y substraty t³uszczowe: tributyryna, Tween 40, Tween 60, Tween 80 oraz glukoza. Aktywnoœæ lipolityczn¹ oznaczono w zakresie pH od 5 do 8 oraz temperaturach od 30 do 60oC metod¹ miareczkow¹, a wyniki podano w jed- 1736 Ma³gorzata Nabrdalik and Katarzyna Grata nostkach aktywnoœci enzymatycznej [U cm–3]. W przeprowadzonym doœwiadczeniu aktywnoœæ lipolityczna uzale¿niona by³a od rodzaju substancji t³uszczowej zawartej w pod³o¿u, pH oraz temperatury. I tak, badane szczepy B. cereus oraz B. mycoides wykazywa³y najmniejsza aktywnoœæ przy pH 5 oraz 6. Natomiast najwiêksz¹ aktywnoœæ stwierdzono przy pH 7 i 8. W tych warunkach wiêkszoœæ szczepów wykazywa³a aktywnoœæ lipolityczn¹ nawet przy braku substratu t³uszczowego w pod³o¿u. Najwiêksze wartoœci aktywnoœci lipolitycznej uzyskano przy pH 8 na pod³o¿u z dodatkiem Tween 40, a najwiêksze wartoœci (0,88 U cm–3) uzyskano dla glebowego szczepu B. cereus. Analizuj¹c wp³yw temperatury na aktywnoœæ lipolityczn¹ stwierdzono, i¿ najwy¿sz¹ aktywnoœæ odnotowano w temperaturze 30 i 40 oC, a najmniejsz¹ w 50 i 60 oC. Najwiêksze wartoœci, dla wiêkszoœci szczepów, uzyskano w temperaturze 30 oC na pod³o¿u z dodatkiem Tween 40, gdzie najbardziej aktywnym okaza³ siê glebowy B. mycoides (0,88 U cm–3). Wyj¹tek stanowi glebowy B. cereus, dla którego wartoœæ aktywnoœci lipolitycznej na pod³o¿u z dodatkiem glukozy wynosi³a a¿ 1,38 U cm–3. Uwzglêdniaj¹c wszystkie analizowane Ÿród³a wêgla i parametry, najaktywniejszymi by³y szczepy B. mycoides. S³owa kluczowe: Bacillus cereus, Bacillus mycoides, lipazy, tributyryna, Tween ECOLOGICAL CHEMISTRY AND ENGINEERING A

Vol. 18, No. 12 2011

Adam RADKOWSKI1

EFFECT OF MICROELEMENT FERTILIZATION ON THE QUALITY AND NUTRITIONAL VALUE OF THE MEADOW SWARD HAY PART II. THE CONTENT OF MACROELEMENTS*

WP£YW NAWO¯ENIA MIKROELEMENTAMI NA JAKOŒÆ I WARTOŒÆ POKARMOW¥ SIANA RUNI £¥KOWEJ CZ. II. ZAWARTOŒÆ MAKROELEMENTÓW

Abstract: The one-factor field experiment was designed by means of random block sampling in four replicants (fields of 2.0 × 5.0 m area). The experimental field was located on the acid brown soil (type II B) of the V quality class. The study was conducted in the years 2006–2008 in the individual farm in Pilica administrative district, in Zawiercie county, in the region of Krakow–Czestochowa Jura, at the altitude of 320 m. The kind of microelement fertilizer was the determining factor in the study. During the experiment foliar fertilizers were applied in the form of single microelements (copper, zinc, manganese) as well as multicomponent preparation – Plonvit P, containing elements in the form of chelates. It was stated that applied fertilization had the most spectacular effect on the sodium content in the meadow sward. As a result of foliar application of multicomponent preparation, copper and manganese significant increase of average content of this element was observed in comparison with the non-fertilized field (2.34; 1.98 and 1.09-time higher values, respectively). Moreover, it was found that foliar treatment with examined microelements caused elevation of calcium and magnesium level by 46.0 % and 45.7 %, respectively. Treatment with the multicomponent fertilizer also resulted in the increased phosphorous content above the standard level. Additionally, applied foliar fertilizers narrowed the proportions between the sum of univalent and divalent cations in the meadow sward. Keywords: meadow sward, fertilization with microelements, chemical composition

The high level of nitrogen, phosphorus and potassium treatment as well as intensive agricultural production affects the enhancement of microelements uptake by plants, what results in their deficiency both in soil and in plants. The amounts of microelements that are retained by the soil during the application of basic mineral and organic fertilizers usually cannot cover the plant requirements for these components. Therefore,

1 Department of Grassland, University of Agriculture in Krakow, al. A. Mickiewicza 21, 31–120 Kraków, Poland, phone: +48 12 662 43 61, fax: +48 12 633 62 45, email: [email protected] * Part I – Ecol. Chem. Eng. A 2011, 18(8), 1111–1115. 1738 Adam Radkowski there is a need to apply microelement fertilizers [1–3]. Although these elements are absorbed in small quantities they are essential for the proper proceeding of many biochemical and physiological processes in plants [4]. Microelements significantly influence not only the yield level but most of all they positively affect its quality as well as the content of organic and mineral compounds in forage [5, 6]. Therefore, the aim of the three-year long field experiment was an estimation of the effect of foliar fertilization with microelements in the form of chelated multicomponent preparation or single microelements on the content of macroelements in the meadow sward and the proportions between them.

Materials and methods

The study was conducted in the years 2006–2008 on the individual agricultural farm in Solca, in the Pilica administrative disctrict. The experiment was designed by the method of random block sampling, in four replicants, on the brown, acid soil (pHKCl = 5.2) of the V quality class. The soil contained medium levels of assimilable potassium, manganese and zinc and were poor in assimilable phosphorus and copper. During the vegetation period (April–September) the total rainfall amounted to 338.1 mm; 375.4 mm and 320.3 mm, and the mean temperatures reached the values of 15.2, 14.3 and 14.9 oC, respectively for the years 2006, 2007 and 2008. During the experiment plants were treated with foliar preparation of 14 % zinc chelate (chelator – EDTA+DTPA) in a dose of 100 g Zn × ha–1, manganese chelate 14 % Mn (chelator – EDTA+DTPA) in a dose of 100 g Mn × ha–1, copper chelate 12 % Cu (chelator – EDTA+DTPA) in a dose of 60 g Cu × ha–1 as well as Plonvit P in a dose of2dm3 × ha–1. The fertilizers were applied for each regrowth. Zinc, manganese and copper were dosed in such proportions to achieve the equal contents of the relevant microcomponents in the single fertilizers and in the multi- component preparation. Plonvit P is a concentrated, multicomponent, microelement fertilizer containing in a single dose of 2 dm3: 100 g Zn, 100 g Mn and 60 g Cu in the form of chelates. The spraying solutions were prepared by dissolution of the proper amounts of chelates containing microelements in such water volume to obtain the volume of working liquid corresponding to 300 dm3 × ha–1. The tap water of the medium hardness degree was used for that purpose. The first spraying with microelements was done after the beginning of the spring vegetation, the following – after the harvesting at the stage of the initial sward regrowth but not later than 3 weeks before the next mowing. During the investigation period the basic mineral fertilization was also applied: under I regrowth – 80 kg N × ha–1 and under II and III regrowths – 60 kg N × ha–1 for each regrowth in the form of ammonium × –1 saltpetre. Phosphorus was applied once in the spring, in the amount of 120 P2O5 ha as a triple superphosphate and potassium – under the first and third regrowths in × –1 a dosage of 60 kg K2O ha (for each regrowth) as 57 % potassium salt. The experimental fields had the area of 10 m2. In the collected plant material the chemical composition of forage, the content of dry matter by drying at 105 oC, phosphorus and magnesium were determined – Effect of Microelement Fertilization on the Quality and Nutritional Value... 1739 colorometrically by the vanadium-molybdenic method, potassium, sodium and calcium – by the flame photometry [7]. On the basis of the obtained results the proportions of the univalent to divalent sum of cations (K + Na) : (Mg + Ca) in the meadow sward were calculated. All results were subjected to the analysis of variance and verified using the Tukey test at the significance level of a = 0.05.

Results and discussion

The conducted study revealed that fertilization with microelements significantly affected the chemical composition of the meadow sward. Foliar application of both the single elements and the multicomponent Plonvit P, which contain the elements in the form of chelate complexes, had the most spectacular influence on the sodium content in the meadow flora (Table 1).

Table 1

The weighted mean of macroelement content and the ionic proportions in the meadow sward as affected by the fertilization with microelements (mean for three years)

Fertilized objects Examined parameter Multicomponent Mean LSD0.05 Control Cu Zn Mn fertilizer P content 2.34 3.27 3.08 3.25 3.04 3.00 0.37 [g × kg–1 d.m.] K content 16.31 22.20 16.71 18.99 16.94 18.23 3.97 [g × kg–1 d.m.] Ca content 3.51 7.71 4.31 6.07 4.00 5.12 2.59 [g × kg–1 d.m.] Mg content 1.03 1.81 1.68 1.64 1.32 1.50 0.31 [g × kg–1 d.m.] Na content 0.120 0.402 0.358 0.132 0.251 0.253 0.150 [g × kg–1 d.m.] (K + Na) : (Ca + Mg) 1.63 1.10 1.25 1.12 1.44 1.26 0.32

The highest sodium level was observed for the object treated with multicomponent fertilizer. The plants from that field contained 70% higher Na level than the plants collected from the non-fertilized object. Foliar application of multicomponent pre- paration, zinc, copper and manganese affected also the significant elevation of the mean calcium content when compared with the control object and the differences reached the values of: 55, 42, 19, 12 %, respectively. The great diversification as dependent on the foliar application of microelements was found for magnesium. The highest increase of its content was observed after the treatment with copper and zinc (besides the multicomponent fertilizer). The Mg level was 39 and 37 %, respectively higher than that of the control object. Copper is an element that takes part in the transformational processes of iron compounds in plants and affects the growth and anatomical structure 1740 Adam Radkowski of many tissues [8]. Zinc is an activator of many enzymes and plant hormones and participates in the synthesis of vitamins B, C and P. Zinc affects the growth and development and enhances the plant good condition [9]. In our investigations we stated also the increase of phosphorus and potassium content after the foliar application of microelements. As a result 26 % higher phosphorus and 13 % higher potassium levels in relation to the control objects were foun under the treatment with microelements. The quality of plant crops is estimated not only on the basis of optimal con- centrations of certain elements but also the proportions between them are of a great importance as the feeding value is taken into consideration [10, 11]. The obtained results indicate that microelement fertilization affected the decreased proportion of univalent to divalent cations. It is worth emphasizing, that the lowest value of the above mentioned ratio was found for the sward foliar fertilized with the solution of multicomponent preparation and zinc.

Conclusions

1. The application of single microelement fertilizers as well as multicomponent Plonvit P preparation highly affected the sodium content in the meadow flora. Foliar application of manganese, copper and Plonvit P caused significant elevation of the Na level by on average 52, 66 and 70 %, respectively in relation to the control object. 2. Fertilization with the examined microelements caused the elevation of calcium and magnesium content by 37 and 36 % on average, in comparison with the control object. 3. The treatment with multicomponent Plonvit P resulted in the highest increase of the examined macroelements. The application of that fertilizer affected 45 % (on average) higher content of the macroelements when compared with the control field. 4. The application of copper, zinc and manganese influenced the elevated content of the examined macroelements by 30, 26 and 23 %, respectively in relation to the control object.

References

[1] Czuba R.: Celowoœæ i mo¿liwoœæ uzupe³nienia niedoborów mikroelementów u roœlin. Zesz. Probl. Post. Nauk Roln. 1996, 434, 55–64. [2] Szewczuk C. and Michalojæ Z.: Praktyczne aspekty dolistnego dokarmiania roœlin. Acta Agrophys. 2003, 85, 19–29. [3] Ruszkowska M. and Wojcieska-Wyskupajtys U.: Mikroelementy – fizjologiczne i ekologiczne aspekty ich niedoborów i nadmiarów. Zesz. Probl. Post. Nauk Roln. 1996, 434, 1–11. [4] Michalojæ Z. and Szewczuk C.: Teoretyczne aspekty dolistnego dokarmiania roœlin. Acta Agrophys. 2003, 85, 9–17. [5] Wojcieska U.: Rola mikroelementów w kszta³towaniu fotosyntetycznej produkcyjnoœci roœlin. Post. Nauk Roln. 1985, 6, 10–24. [6] Gorlach E.: Zawartoœæ pierwiastków œladowych w roœlinach pastewnych jako miernik ich wartoœci. Zesz. Nauk. AR w Krakowie 1991, 262, Sesja Nauk. 34, 13–22. [7] Ostrowska A., Gawliñski S. and Szczubia³ka Z.: Metody analizy i oceny w³aœciwoœci gleb i roœlin. Katalog. Wyd. IOŒ Warszawa 1991, pp. 334. Effect of Microelement Fertilization on the Quality and Nutritional Value... 1741

[8] Wojciechowska-Wyskupajtys U.: Efekty dolistnego dokarmiania roœlin w œwietle referatów wyg³o- szonych na „Miêdzynarodowym Sympozjum Dolistnego Nawo¿enia” w Kairze (10–14.12.1995). Post. Nauk Roln. 1996, 5, 123–127. [9] Grzywnowicz-Gazda Z.: Wp³yw zró¿nicowanego nawo¿enia cynkiem na wysokoœæ i jakoœæ plonu ziarna jêczmienia jarego. Zesz. Probl. Post. Nauk Roln. 1983, 242, 201–209. [10] Spiak Z.: Mikroelementy w rolnictwie. Zesz. Probl. Nauk Roln. 2000, 471, 29–34. [11] Krzywy J., Baran S. and Krzywy E.: Wp³yw nawozów jednosk³adnikowych i wielosk³adnikowych na kszta³towanie stosunków jonowych K : Mg, K : (Mg + Ca), Ca:PorazN:Swroœlinach uprawnych. Zesz. Probl. Post. Nauk Roln. 2002, 484, 317–323.

WP£YW NAWO¯ENIA MIKROELEMENTAMI NA JAKOŒÆ I WARTOŒÆ POKARMOW¥ SIANA RUNI £¥KOWEJ CZ. II. ZAWARTOŒÆ MAKROELEMENTÓW

Katedra £¹karstwa Uniwersytet Rolniczy im. Hugona Ko³³¹taja w Krakowie

Abstrakt: Jednoczynnikowe doœwiadczenie polowe za³o¿ono metod¹ losowanych bloków, w czterech powtórzeniach (poletka o wymiarach 2,0 × 5,0 m). Na polu doœwiadczalnym wystêpowa³a gleba brunatna kwaœna (rz¹d II B – gleby brunatne ziemne), zaliczana pod wzglêdem bonitacyjnym do klasy V. Doœwiad- czenie prowadzono w latach 2006–2008 w indywidualnym gospodarstwie rolnym po³o¿onym w gminie Pilica, powiat zawierciañski na Jurze Krakowsko-Czêstochowskiej, na wysokoœci powy¿ej 320 m n.p.m. Czynnikiem doœwiadczenia by³ rodzaj nawo¿enia mikroelementowego. W doœwiadczeniu zastosowano dolistnie nawo¿enie pojedynczymi mikroelementami (miedŸ, cynk i mangan) oraz wielosk³adnikowy nawóz zawieraj¹cy pierwiastki w formie schelatowanej – Plonvit P. Wykazano, ¿e zastosowane nawo¿enie najwiêkszy wp³yw wywiera³o na zawartoœæ sodu w runi ³¹kowej. W wyniku dolistnego stosowania wielosk³adnikowego nawozu oraz miedzi i manganu stwierdzono znaczny wzrost œredniej zawartoœci tego pierwiastka – w porównaniu z obiektem nienawo¿onym – odpowiednio: 2,34; 1,98; i 1,09-krotnie. Ponadto stwierdzono, i¿ dolistne zastosowanie badanych mikroelementów spowodowa³o wzrost zawartoœci wapnia i magnezu odpowiednio o 46,0 i 45,7 % w porównaniu z obiektem kontrolnym. Wykazano równie¿, ¿e nawo¿enie wielosk³adnikowym nawozem spowodowa³o wzrost zawartoœci fosforu ponad wartoœæ nor- matywn¹. Ponadto zastosowane nawozy dolistne zawêzi³y stosunek sumy kationów jednowartoœciowych do sumy kationów dwuwartoœciowych w runi ³¹kowej. S³owa kluczowe: ruñ ³¹kowa, nawo¿enie mikroelementami, sk³ad chemiczny

ECOLOGICAL CHEMISTRY AND ENGINEERING A

Vol. 18, No. 12 2011

Anna KWIECIÑSKA1 and Krystyna KONIECZNY1

RECOVERY OF INDUSTRIAL WATER FROM PIG LIQUID MANURE BY MEANS OF MEMBRANE TECHNIQUES

ODZYSK WODY PRZEMYS£OWEJ Z GNOJOWICY TRZODY CHLEWNEJ Z WYKORZYSTANIEM TECHNIK MEMBRANOWYCH

Abstract: Liquid manure that is produced during high density livestock farming requires proper utilization methods. Nowadays, it is mainly used as a fertilizer or as a substrate for biogas or compost production. However, these methods are often very limited and do not allow to utilize the total amount of produced liquid manure, thus it is still treated as problematic waste. High water content in liquid manure leads to the assumption that it can be treated as a water source. This assumption is quite realistic if application of low and high pressure membrane techniques is considered. Such a solution would allow not only to recover water that could be further reused on farms, but also to obtain valuable concentrated nutrients solution which can be used as a fertilizer and easily transported to agricultural areas. The aim of the study was to determine the effectiveness of water recovery from pig liquid manure using integrated system: centrifugation/two step ultrafiltration/nanofiltration. The first step ultrafiltration was performed using PVDF membrane of cut off 100 kDa while the second step using PES membrane of cut off 10 kDa. During the polishing process ie nanofiltration hydrophilic composite membrane of cut off 200 Da was used. The effectiveness of the process was determined basing on the change of values of parameters like: + – 2– 3– 2+ 2+ BOD5 and COD, contents of TOC, IC, TC and Ntot, concentrations of NH4 ,Cl,SO4 ,PO4 ,Mg ,Ca and K+, while the capacity of the treatment was determined using volumetric permeate streams. Obtained results allow to conclude that the proposed system can be used to recover water of an industrial quality. Keywords: liquid manure, water recovery, membrane processes, ultrafiltration, nanofiltration

High density livestock farming results in the production of a significant amount of liquid manure which is a mixture of animal urea, feces and water [1–3]. Nowadays, it is mainly used as a fertilizer or a substrate for biogas or compost production. In Poland, 70 mln m3 of liquid manure is produced every year, however only ca 21 mln m3 is applied for fertilizing purposes [4], whereas the rest is treated as a highly load waste and

1 Division of Sanitary Chemistry and Membrane Processes, Institute of Water and Wastewater Engineering, Faculty of Energy and Environmental Protection, Silesian University of Technology, ul. J. Konarskiego 18, 44–100 Gliwice, Poland, phone: +48 32 237 29 81, email: [email protected] 1744 Anna Kwieciñska and Krystyna Konieczny requires special treatment methods. However, except for nutrients, liquid manure contains also a significant amount of water (ca 90–97 %) which is usually omitted product during consideration of liquid manure management methods. The development and improvement of membrane processes create the possibility of water recovery from liquid manure [5–7]. The application of low and high pressure membrane techniques would allow to obtain water of proper quality that could be reused on farms. The recovery of water would be a great advantage in areas which deal with its deficit and the concentrated fertilizing mixtures could be easily transported to agriculture areas [8–11].

Materials and methods

A50dm3 sample of a pig liquid manure was collected from a 13000 m3 lagoon localized at one of the high density livestock farm in Silesia region. Firstly, the liquid manure was centrifuged for 10 minutes at the rotational speed of 15000 rpm. The obtained supernatant was next introduced to the laboratory membrane cell by Koch (Fig. 1). The device is equipped with the 0.5 dm3 feed tank and the effective separation area of the installed flat membrane is equal to 28 cm2. All processes were carried out in the cross-flow mode.

VENT INERT GAS INLET NF/RO

FEED TANK

HEATING/ COOLING

PI FEED PUMP MEMBRANE CELL

MF/UF

PERMEATE CONCENTRATE

Fig. 1. The scheme of the laboratory installation for membrane filtration by Koch

Firstly, 100 kDa polyvinylidene fluoride (UF-PVDF-100) membrane was used for ultrafiltration of the supernatant and the permeate was introduced to the second treatment step which was carried out with the use of 10 kDa polyethersulfone (UF-PES-10) membrane. The polishing of the second ultrafiltration step permeate was performed using composite nanofiltration membrane of cut off 200 Da (NF-200). All applied membranes were provided by KOCH. The detail scheme of the process is shown in Fig. 2. Recovery of Industrial Water from Pig Liquid Manure... 1745

Crude slurry Concentrate 1

Centrifugation Supernatant UF 1° 10 min, PVDF, 15000 rpm 100 kDa, 0.3 MPa

Deposit Permeate 1

Concentrate 3 NF, composite Permeate 2 UF 2° membrane, 200 Da PES, 10 kDa, 3.0 MPa 0.45 MPa

Concentrate 2

Permeate 3 (water)

Fig. 2. The scheme of the pig liquid manure treatment process in the integrated system of centrifugation/two step ultrafiltration/nanofiltration

Following parameters were analyzed in the produced process streams: pH, conduct- + 3– ivity, COD, BOD5, contents of TOC, IC, TC, Ntot, concentrations of ions NH4 ,PO4 , – 2– 2+ 2+ + + Cl ,SO4 ,Mg,Ca and K (Na – only in nanofiltration permeate). COD, + 3– + concentrations of K ,PO4 ,Ntot. and NH4 were determined according to the methodology by Merck methodology, while BOD5 by means of the respirometric – 2– method with the use of the OXI Top WTW set. Concentrations of Cl and SO4 ions were obtained during chromatographic analysis using ionic chromatograph DX 120 by Dionex. The content of particular carbon forms was analyzed using Multi N/C analyzer by Jena Analytic. Concentrations of Ca2+ and Mg2+ were determined by means of the classical titration method while Na+ content using flame photometer. The liquid manure membrane filtration was preceded by the determination of the dependence the volumetric deionized water flux on pressure for all membranes applied. After the liquid manure treatment membranes were washed with deionized water and its volumetric flux was again measured.

Results and discussion

The liquid manure treatment was preceded by the characterization of applied membranes ie the determination of dependence of the volumetric deionized water flux on pressure. The results of the experiment are presented in Fig. 3a–c. The logarithmic dependence of the volumetric deionized water flux on pressure determined for 100 kDa membrane indicated that the increase of the transmembrane pressure above 0.4 MPa would not improve the process capacity. For other membranes ie 10 kDa and 200 Da regular, linear dependences were obtained. The study of the composition of treated streams indicated that the proposed system of integrated centrifugation/two step ultrafiltration/nanofiltration was sufficient for the 1746 Anna Kwieciñska and Krystyna Konieczny

(a) 1600 y = 497.4 ln(x) + 1620 1400 R2 = 0.899 32 1200 1000 800 600

400 200

Deionized water flux [dm /m h] 0 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 Pressure [MPa]

(b) 700 y = 921.4x + 18.696 600 2

32 R = 0.9957 500

400

300

200

100

Deionized water flux [dm /m h] 0 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 Pressure [MPa]

(c) 45 y = 11.957x – 2.112 40 R2 = 0.9872 32 35 30 25 20 15 10 5

Deionized water flux [dm /m h] 0 0.0 1.5 2.0 2.5 3.0 3.5 4.0 Pressure [MPa]

Fig. 3. The dependence of the volumetric deionized water flux on pressure: (a) UF-PVDF – 100 kDa; (b) UF-PES – 10 kDa; (c) NF – 200 Da liquid manure treatment. The improvement of the quality of treated streams was observed in every step (Table 1). However, finally obtained permeate did not fulfill the + regulations for the drinking water quality [12] as concentrations of TOC and NH4 ions, which were exceeded, and Mg2+ and Ca2+ ions were absent (Table 2). Nevertheless, it can be used as a sanitary safe industrial water for washing of farmhouses and animals as well as for fields watering. Recovery of Industrial Water from Pig Liquid Manure... 1747

Table 1

The comparison of parameters of treated streams

Crude Permeate 3 Parameter Unit Supernatant Permeate 1 Permeate 2 liquid manure (water) pH — 7.36 7.69 8.1 8.17 8.6 Conductivity mS/cm 5.96 5.84 5.75 5.6 0.675 TOC mg/dm3 1134 671 528 381 9.7 IC mg/dm3 466 437 420 370 44 TC mg/dm3 1600 1108 948 751 53.7 3 COD mgO2/dm 7560 3785 2990 2285 13.9 3 BOD5 mgO2/dm 3000 2900 1500 480 n/m* + 3 NH4 mg/dm 1350 1200 1478 1414 135 3 Ntot mg/dm 3100 2950 2950 2950 148 2– 3 SO4 mg/dm 199 192 185 49.8 0 3– 3 PO4 mg/dm 63.1 34.6 32.2 24.7 0 Cl– mg/dm3 385 373 371 311 52 K+ mg/dm3 820 805 790 790 180 Mg2+ mg/dm3 19.2 16.8 14 11 0 Ca2+ mg/dm3 100 100 88 80 0 Na+ mg/dm3 ————41 * Non measurable.

Table 2

The comparison of parameters of nanofiltration permeate and drinking water

Permeate Parameter Unit Drinking water*, ** 100-10-0.2 pH — 8.60 6.5–9.5 Conductivity mS/cm 675 2500 Smell — acceptable*** acceptable*** TOC mg/dm3 9.7 5.0 Cl– mg/dm3 52 250 2– 3 SO4 mg/dm 0 250 + 3 NH4 mg/dm 135 0.5 3 Total hardness mgCaCO3/dm 0 60–500 Na+ mg/dm3 41 200 * Permissible values of parameters of drinking water according to the Regulation of Minister of Health on drinking water quality from 20.04.2010; ** There are no regulations considering quality of water for animals watering; *** Slight, hardly sensible, determined during cool study by 4 people.

The evaluation of process capacities indicated that the liquid manure filtration caused fouling of all applied membranes as all permeate fluxes decreased during the treatment 1748 Anna Kwieciñska and Krystyna Konieczny

(Fig. 4a–c). Both types of fouling, the reversible and irreversible ones, were observed. However, the second fouling type occurred only in case of ultrafiltration and was most severe during the first step. In comparison with distilled water flux, the membrane capacity during liquid manure treatment decreased by 90 % for 100 kDa membrane, 63 % for 10 kDa membrane and 31 % for 200 Da membrane. However, while comparing initial and final liquid manure fluxes, the capacity of the first step ultrafiltration decreased by 36 %, the second step ultrafiltration by 29 % and the polishing step by 76 %.

(a) 160 140

32 120 100 80 60 40 20 Permeate flux [dm /m h] 0 0.0 0.2 0.4 0.6 0.8 1.0 1.2 Time [h]

(b) 250

200

32

150

100

50

Permeate flux [dm /m h] 0 0.0 0.1 0.2 0.3 0.4 0.5 0.6 Time [h]

(c) 35 30

32 25 20 15 10 5

Permeate flux [dm /m h] 0 0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 Time [h]

Fig. 4. The decrease of the permeate flux during liquid manure filtration: (a) UF-PVDF – 100 kDa; (b) UF-PES – 10 kDa; (c) NF – 200 Da Recovery of Industrial Water from Pig Liquid Manure... 1749

The applied washing of membranes with distilled water allowed to partially recover the initial capacity of UF membranes, while in case of the NF membrane the post-process water flux was even greater than the initial one. It was probably due to the modification of the membrane, which could have been caused by the deposition of calcium and magnesium ions and/or organic substances present in the liquid manure on the membrane surface. The washing of membranes with distilled water fluxes enabled the initial capacity recovery at the level of 39 % for 100 kDa membrane, 65 % for 10 kDa membrane and 103 % for 200 Da membrane. The comparison of initial distilled water streams, permeate streams and washing distilled water streams is presented in Fig. 5.

1400 0.3 Deionized water MPa 1200 Treated slurry Deionized water after process 1000

800

32

600 0.3 0.4 MPa Flux[dm/mh] MPa 400 0.45 0.45 MPa 0.3 MPa 200 MPa 3.0 3.0 3.0 MPa MPa MPa 0 UF-PVDF UF-PES NF composite 100 kDa 10 kDa 200 Da

Fig. 5. The comparison of volumetric fluxes of the deionized water (before and after the process) and the treated liquid manure determined for all membranes

Conclusions

The presented study focused on the recovery of the industrial water from the pig liquid manure by means of the integrated system comprised of centrifugation/two step ultrafiltration/nanofiltration. The obtained results allowed to conclude that the proposed system is suitable for that purpose and the final permeate could be reused on farm eg for animals or farmhouse washing. However, the obtained water did not fullfill the regulations for drinking water quality as concentrations of ammonium ion and TOC were exceeded and it did not contain magnesium and calcium ions. The appearnce of fouling phenomenon was observed in case of all membranes. Its capacity of greatest influence on the membrane capacity was observed in case of UF membranes. Additionally, for these membranes both forms of fouling, reversible and irreversible, were present. The washing of membranes with distilled water did not allow to recover the initial UF membranes, while for NF membranes the post-process water flux was greater than the initial one. It was explained by the possible modification of the polishing membrane surface, probably by magnesium and calcium ions and/or the organic substances. 1750 Anna Kwieciñska and Krystyna Konieczny

References

[1] Ma³a encyklopedia rolnicza, PWRiL, Warszawa 1964. [2] Kutera J.: Gospodarka gnojowic¹, Wyd. Akad. Roln. we Wroc³awiu, Wroc³aw 1994. [3] Hus S.: Chemia wody, œcieków i gnojowicy, Wyd. Akad. Roln. we Wroc³awiu, Wroc³aw 1995. [4] Charakterystyka Gospodarstw Rolnych w 2007 r., Informacje i Opracowania Statystyczne, GUS, Departament Rolnictwa i Gospodarki ¯ywnoœciowej, Departament Pracy i Warunków ¯ycia, Warszawa 2008. [5] Van der Bruggen B., Vandecasteele C., Gestel T.V., Doyen W. and Leysen R.: A review of pressure- -driven membrane processes in wastewater treatment and drinking water production, Environ. Progr. 2003, 22, 46–56. [6] Bodzek M. and Konieczny K.: Mo¿liwoœæ zastosowania technik membranowych w in¿ynierii œrodowiska, [in:] Monografie Komitetu In¿ynierii Œrodowiska PAN, Gliwice 2002, 12, 191–228. [7] Jarusutthirak C. and Amy G.: Membrane filtration of wastewater effluents for reuse: effluent organic matter rejection and fouling, Water Sci. Technol. 2001, 43(10), 225–232. [8] Fugere R., Mameri N., Gallot J.E. and Comeau Y.: Treatment of pig farm effluents by ultrafiltration, J. Membr. Sci. 2005, 255, 225–231. [9] Buelma G., Dube R. and Turgeon N.: Pig manure treatment by organic bed biofiltration, Desalination 2008, 231, 297–304. [10] Massea L., Massea D.I. and Pellerin Y.: The effect of pH on the separation of manure nutrients with reverse osmosis membranes, J. Membr. Sci. 2008, 325, 914–919. [11] Pieters J.G., Neukermans G.G.J. and Colanbeen M.B.A.: Farm-scale membrane filtration of sow slurry, J. Agricult. Eng. Res. 1999, 73, 403–440. [12] Rozporz¹dzenie Ministra Zdrowia z dn. 29.03.2007 w sprawie jakoœci wody przeznaczonej do spo¿ycia przez ludzi, DzU, nr 61, poz. 417.

ODZYSK WODY PRZEMYS£OWEJ Z GNOJOWICY TRZODY CHLEWNEJ Z WYKORZYSTANIEM TECHNIK MEMBRANOWYCH

Wydzia³ In¿ynierii Œrodowiska i Energetyki Politechnika Œl¹ska, Gliwice

Abstrakt: Gnojowica powstaj¹ca podczas wielkoprzemys³owej hodowli zwierz¹t wymaga stosowania odpowiednich metod utylizacji. Obecnie jest ona wykorzystywana jako nawóz b¹dŸ te¿ substrat do produkcji biogazu oraz kompostu. Jednak¿e metody te s¹ czêsto ograniczone i nie pozwalaj¹ na zagospodarowanie ca³kowitej iloœci powstaj¹cej gnojowicy, st¹d te¿ wci¹¿ jest ona traktowana jako uci¹¿liwy odpad. Du¿a zawartoœæ wody w gnojowicy pozwala za³o¿yæ, i¿ mo¿e byæ ona traktowana jako Ÿród³o wody. To za³o¿enie jest ca³kiem realne w przypadku zastosowania nisko i wysokociœnieniowych procesów membranowych. Takie rozwi¹zanie pozwoli³oby nie tylko na odzysk wody, który mog³aby zostaæ ponownie wykorzystana na farmie, ale tak¿e na otrzymanie wartoœciowych, stê¿onych roztworów substancji od¿ywczych, które mog³yby zostaæ wykorzystane jako nawóz i ³atwo transportowane na tereny rolnicze. Celem przeprowadzonych badañ by³o okreœlenie efektywnoœci odzysku wody z gnojowicy trzody chlewnej, wykorzystuj¹c zintegrowany system: wirowanie/dwustopniowa ultrafiltracja/nanofiltracja. Pierwszy stopieñ ultrafiltracji prowadzono z u¿yciem membrany z PVDF o cut off 100 kDa, drugi zaœ stopieñ z wykorzystaniem membrany z PES o cut off 10 kDa. Podczas etapu doczyszczania, tj. nanofiltracji zastosowano kompozytow¹ membranê hydrofilow¹ o cut off 200 Da. Efektywnoœæ procesu okreœlono, korzystaj¹c ze zmiany wartoœci parametrów procesu: BZT5, ChZT, zawartoœci OWO, WN, OW, Nca³ stê¿enia + – 2– 3– 2+ 2+ + jonów NH4 ,Cl,SO4 ,PO4 ,Mg ,Ca iK, wydajnoœæ zaœ wyznaczono na podstawie objêtoœciowych strumieni permeatów. Przeprowadzane badania pokazuj¹, ¿e zaproponowany system mo¿e zostaæ wyko- rzystany do odzysku wody o jakoœci przemys³owej. S³owa kluczowe: gnojowica, odzysk wody, procesy membranowe, ultrafiltracja, nanofiltracja ECOLOGICAL CHEMISTRY AND ENGINEERING A

Vol. 18, No. 12 2011

Karolina MIELCZAREK1, Jolanta BOHDZIEWICZ2 and Anna KWARCIAK-KOZ£OWSKA1

APPLICATION OF POLYSULFONE MEMBRANES FOR COKE-MAKING WASTEWATER TREATMENT

MEMBRANY POLISULFONOWE W OCZYSZCZANIU ŒCIEKÓW KOKSOWNICZYCH

Abstract: Considering the complicated and variable type of coke-making sewages their purification strategy is difficult to generalize and it requires an integrated system, which joins biological and physicochemical separated processes. In the paper the researches were presented, which purpose was to define the efficiency of purifying matting pressure membrane techniques – ultrafiltration and reverse osmosis. In the process of low-pressure filtration flat membranes, applied in the laboratory, were different in terms of compactness of structure and porosity. Keywords: coke-making wastewater treatment, integrated system, membranes pressure techniques, poly- sulfone ultrafiltration membranes

The aqueous-sewage management environmentally safe is the obligation of each industrial plant. The best solution would be to establish a local sewage treatment plants assuring to neutralize wastewaters. The negative effect caused by coking plants upon the environment consists in carrying industrial waters cleared in the insufficient rank to plumbing or receivers. The way of cleaning them is of great importance in aspect of nature protection. The coke-making wastewaters industry have changeable compound composition. They contain: polycyclic aromatic hydrocarbons, compounds heterocyclic, oils, tars and substances of inorganic character such as: cyanide, sulfides, sulfates, thiosulfates, ammonia and also heavy metals. One of the ways and the most frequently applied in the wastewater treatment technology and which seems to be environmentally friendly are pressure membrane techniques. It is possible to link them with classical separated processes of purifying wastewaters into so-called hybrid or integrated systems [1–9].

1 Institute of Environmental Engineering, Czestochowa University of Technology, ul. BrzeŸnicka 60a, 42–200 Czestochowa, Poland, phone: +48 34 325 73 34, email: [email protected] 2 Institute of Water and Wastewater Engineering, Silesian University of Technology, ul. J. Konarskiego 18, 44–100 Gliwice, Poland, phone: +48 32 237 15 26, email: [email protected] 1752 Karolina Mielczarek et al

Apparatus

In the process of membranes cleaning coking industry sewages a system equipped with the plate-frame membrane module of the American Osmonics company of the SEPA CF-NP type, the container of sewers with the radiator was applied as well as cone-and-float meter, the high-pressure pump and manometers and valves. A scheme of the equipment used in investigations is presented below.

Fig. 1. Scheme of the plate- and frame membrane module applied for cleaning coke-making sewage

Raw wastewater

The process of cleaning was applied in the coke plant effluents coming from Coke Plant ISD Huta Czestochowa “Koksownia” Sp. z o.o., which characteristics are display- ed in Table 1.

Table 1

Characteristics of coke-making effluents from coking plant ISD Huta Czestochowa “Koksownia” Sp. z o.o.

Raw Accompanied to receiver set the coefficients Indicator Unit wastewater of pollutions of sewages* pH [-] 8.7–10.9 6.5–9.0 3 COD mgO2/dm 4200–3100 125 3 BOD5 mgO2/dm 20–80 25 + 3 Ammoniacal nitragen mg NH4 /dm 25–104 10 * Directive of the Environment Minister from 24th of July 2006 in the matter of conditions one should fulfil which at inserting sewages to waters or soil, and in the matter of substances particularly harmful to the environment aqueous (DzU 2006, nr 137, poz. 984). Application of Polysulfone Membranes for Coke-Making Wastewater Treatment 1753

As we can observe the values of coefficients describing coke-making wastewater in comparison with standardized values are higher what makes it impossible to pour them directly in to receiver. They are also hard to biodegradation process due to very low value of BOD5/COD relationship

Analytical methods

During the first stage of researches carried out diversified polysulfone flat membranes were prepared varying from 13 to 17 % mass of the polymer in organic solvent and their appropriate porosity was outlined. In order to form of long-lasting membrane structure they were subjected to the process of conditioning, consisting in pure water filtering with the changeable membrane pressure taking out from 0.5 × 105 Pa to 3.0 × 105 Pa and linear velocity of pure water over membrane surface – 2 m/s. Membranes shaped up to the moment of stabilizing the amount of the flux of pure water. During the next part of the experiment their transport properties were defined outlining relation between the volumetric flux of pure water from and the trans- -membrane pressure changed in the scope of value from 0.5 × 105 Pa to 3.0 × 105 Pa and linear velocity of medium over the membrane surface – 2 m/s. The last stage of researches consisted in defining the possibilities of applying manufactured ultrafiltrating polysulfone membranes in cleaning coking industry sewages. The usefulness of the membranes depended on: permeability of membranes, relative permeability and the grade of removing the cargo of pollutants from wastewater. According to interlaboratory studies, purified (with the suitable polysulfone membrane singled out earlier) coke-making industry sewages were characterized by high value of ratio of pollutants which impeded the immediate carrying them to the natural receiver, they were subjected to cleaning with the reverse osmosis method applying of the Osmonics company of the SE type membrane The effectiveness of this process was assessed, as in the case of ultrafiltration taking into consideration the size of the permeate flux and the change in pollutants value of raw and purified sewages characteristics.

Results and discussion

Preparing asymmetric polysulfone membranes and marking their proper porosity

Preparing PSF membranes consisted in pouring the thin film out from membrane solution and gelation into pure water. The membrane solution was prepared by dissolving in period of 24 h polysulfone in N,N-dimethylformamide. The concentration of the polymer changed in the scope from 13 to 17 % weight what resulted in the shift of the porosity and consequently in the density of the membranes structure (Fig. 2). We can see clearly that the porosity of membranes decrease together with the increase of the concentration of the polymer and their structure became much more 1754 Karolina Mielczarek et al

75.0

74.5

74.0

73.5

73.0

72.5

Suitable porosity [%] 72.0

71.5 13 14 15 16 17 Part of polymer in membrane solution [% weight]

Fig. 2. Dependence of suitable porosity synthesized polysulfone memebranes on content polysulfone in membrane solution open. So the porosity of the PSF-13 membrane took the 74.8 %, PSF-15 of the 73.94 % and the lowest was the PSF-17 with the 72.8 %.

Transport properties of ultrafiltration polysulfone membranes

Defining the transport property of manufactured membranes consisted in outlining the relation between the temporary volume stream of pure water and the pressure. Measurements made demonstrated that the plumbing membranes productivity depend- ence on the pressure trans-membrane (Figs. 3, 4). The biggest volumetric flux of pure water was observed in PSF-13 membrane and the lowest stream was obtained in PSF-17 membrane due to the its clenched structure. For the trans-membrane pressure3×105 Pa it was 97 times smaller in comparison with

18 PSF-13 16 PSF-14 14

532

×× 12 y = –125.29x2 + 67.856x + 6.434

2 2

HO R = 0.9861 10

8 y = –4.6669x + 1.8015 R2 = 0.9392 6

4

2

Volumetric flux of water J0 10 [m /m s] 0 0.05 0.1 0.15 0.2 0.25 0.3 0.35 Pressure [MPa]

Fig. 3. Dependence of volumetric water flux on use ie pressure for membranes PSF-13 and PSF-14 Application of Polysulfone Membranes for Coke-Making Wastewater Treatment 1755

0.7

0.6 PSF-15 PSF-16 y = 1.3943x2 + 0.7456x + 0.2545 PSF-17 R2 = 0.9668 0.5

532

××

2 0.4

HO y = –1.2x2 + 1.6069x + 0.0778 R2 = 0.9804 0.3

0.2 y = –1.3657x2 + 0.1058x + 0.0612 R2 = 0.9237 0.1

Volumetric flux of water J 10 [m /m s]

0 0 0.05 0.1 0.15 0.2 0.25 0.3 0.35 Pressure [MPa]

Fig. 4. Dependence of volumetric water flux on use ie pressure for membranes PSF-15, PSF-16 and PSF-17 the flux water for the PSF-13 membrane, in the same conditions of the filtration. In the D case of all membranes relations JH2O =f( P) described quadratic equations and high value of rates proved the correct selection of the regression line (function).

Coke-making wastewater purification in integrated system of ultrafiltartion-reverse osmosis

Investigations carried out in this stage were supposed to allow to outline a poly- sulfone ultrafiltrating which would be describe itself with the big efficient and would assure the biggest grade of removing charge of pollutants from cleaning wastewater. Coke-making industry sewages was subjected to cleaning with low pressure filtration outlining for each of membranes the relation between experimental temporary flux and the time filtration (Figs. 5, 6). The biggest flux post-treat was observed in the process of the low-pressure filtration of the PSF-13 membrane. Its value took 2.73 × 10–5 m3/m2 × s out after stabilizing. The fastest decrease of the stream was observed for this membrane at the same time during what was caused by the biggest intensity of the fouling process. In the 125 first minutes the flux permeate diminished 5 times. Different post-treat properties were observed in PSF-17 membrane which had the lowest stream and in comparison with PSF-13 47.1 % smaller. Also for this membrane the smallest stream decrease was registered. Changes estimated as the relationship of the experimental volumetric momentary flux, in their relative permeability were also appointed in the ultrafiltration process of cleaning coking industry sewages for examined membranes permeate (Jv) to the initial 1756 Karolina Mielczarek et al

16

PSF-13 14 PSF-14

12

532

×× 10

u

8

6

4

2

Volumetric flux of permeate J 10 [m /m s]

0 0 60 120 180240 300 360 420 480 540 600 Time [min]

Fig. 5. Dependence of temporary experimental volumetric flux on time treatment coke-making ultrafiltartion process at membranes PSF-13 and PSF-14

0.45 PSF-15 0.40 PSF-16 PSF-17 0.35

532 0.30

××

u 0.25

0.20

0.15

0.10

Volumetric flux of permeate0.05 J 10 [m /m s]

0 0 60 120 180 240 Time [min]

Fig. 6. Dependence of temporary experimental volumetric flux on time treatment coke-making ultrafiltartion process at membranes PSF-15, PSF-16 and PSF-17 flux of pure water (J0). The change in the permeability of the relative membrane is bound with intensity of the phenomenon setting on its surface and inside pore of the low-pressure process for the membranes fouling separation, into lowering, in the considerable way relative permeability of the membrane. Many factors have effect on to Application of Polysulfone Membranes for Coke-Making Wastewater Treatment 1757 the step of membrane polluting, and the most important are: affinity of chemical substances presented in sewages in comparison with polymer membrane material and the size and the structure of their particles. In Figs. 7 and 8 changes in the relation for the relative permeability of membranes were compared from the time low-pressure filtration carry out for cleaning coking industry sewages. It has been proved that the lowest relative permeability had PSF-13 membrane and the highest PSF-16. It was demonstrated that the relative permeability of examined

0.8

0.7 PSF-13 PSF-14

0 0.6

J

v 0.5

0.4

0.3

0.2

Relative penetrability (J / )

0.1

0 050 100 150 200 250 300 350 400 450 500 550 600 Time [min]

Fig. 7. Compare dependence of changes relative penetrability polysulfone membranes PSF-13 and PSF-14 on time treatment coke-making ultrafiltration process

1.2

PSF-15 1.0 PSF-16 PSF-17

0

J

v 0.8

0.6

0.4

Relative penetrability (J / ) 0.2

0 050 100 150 200 Time [min]

Fig. 8. Compare dependence of changes relative penetrability polysulfone membranes PSF-15, PSF-16 and PSF-17 on time treatment coke-making ultrafiltration process 1758 Karolina Mielczarek et al membranes is a function of the density of their structure. The PSF-13 membrane the most opened structure is relatively easily undergoing the fouling process. The pores are blocked quickly by sewages due to the big diameters. Together with the height membrane polymeric contents their porosity is decreasing and the relative permeability is increasing. Another lowering was observed in the PSF-17 membrane. It is possible to example this fact with the smallest porosity of this membrane that caused layer covering in the form of pollutants as a result in a smaller intensity and blocks the pores of “secondary membrane” which final has the smaller membrane permeability in com- parison with membranes with blocked times. The assessment criterion for the effectiveness of low-pressure wastewater filtration was apart from the size of the volumetric flux permeate and relative permeability, step of removing of pollutants. The level of cleaning coke-making wastewater was related with the change of value of the following parameters: COD, TC and TOC, which was typical for “raw” and cleaned sewers. For the PSF-16 membrane additionally a dry substance remains after roasting and conductivity. From prepared for five polysulfone membranes, as it was expected, none assured the high level of removing charge of pollutants from cleaned sewages appropriately what made it impossible in consequence their immediate carry to the natural environment (Table 2).

Table 2

The stage of removal in process the cargo of pollutions treatment coke-making sewages in ultrafiltration process with use of polysulfone membranes

Raw Degree of removal Membrane Indicator Unit Permeate wastewater pollutions [%] 3 COD mgO2/dm 3266 2939 10 PSF-13 TC mgC/dm3 720.95 690.7 4.2 TOC mgC/dm3 495.58 481.7 2.8 3 COD mgO2/dm 3038.5 2488.5 18.1 PSF-14 TC mgC/dm3 718.2 638.5 11.1 TOC mgC/dm3 483.2 450.8 6.7 3 COD mgO2/dm 3808 3027.4 20.5 PSF-15 TC mgC/dm3 707.3 600.5 15.1 TOC mgC/dm3 494.6 448.6 9.3 3 COD mgO2/dm 2754 2087.5 24.2 TC mgC/dm3 616.9 515.1 16.5 TOC mgC/dm3 390.7 340.3 12.9 3 PSF-16 DS g/dm 6.28 6.09 3 Compounds of inorganic substances g/dm3 0.605 0.35 42 3 COD mgO2/dm 2754 1556 43.5 PSF-17 TC mgC/dm3 616.9 70.7 88.6 TOC mgC/dm3 390.7 63.3 83.8 TC – Total carbon; TOC – Total organic carbon; COD – Chemical Oxygen Demand. Application of Polysulfone Membranes for Coke-Making Wastewater Treatment 1759

As the most suitable was accepted PSF-16 membrane with 16 % weight of the polymer in membrane solution. Its choice was determaid by the the volumetric flux size of permeate flux, which in comparison with the membrane about the most clenched structure (PSF-17) was over three times higher (DP=3.0×105 Pa). Besides that this memebrane had the highest relative permeability. After using this membrane purified sewages were characterized by the following indexes of pollutants: COD – 2087.5 mgO2/dm3, the concentration of total carbon TO of 515.1 mgC/dm3, total organic carbon of 340.3 mgC/dm3 and ammoniacal nitrogen 98 + 3 mgNH 4/dm . As it can be seen all the values exceeded the acceptable standardized levels. Sewages left after the process of ultrafiltration treatment were subjected to in additional cleaning of the reverse osmosis with the method on polymer for SE membrane. Changes in the volumetric flux of pure water and sewages were measured during the investigations and we can concluded, that outlined fluxes during carry out the process (pure water and permeate) moved close. It proves the fact that the applied osmotic membrane is characterized by a big density (Fig. 9).

0.43

0.41 y = –5E-18x + 0.416 2 0.39 R = –8E-17

0.37

532 0.35

×× u 0.33

0.31 y = –2E-06x32 + 0.0002x + 0.0076x 2 Volumetric temporary flux R = 0.9876 0.29

of permeate J 10 [m /m s] 0.27 de-ionized water coke wastewater 0.25 0204060 Time [min]

Fig. 9. Dependence of temporary volumetric treatment coke-making wastewater and pure water on time high pressure process

Compared indicators cleaned in the integrated system ultrafiltration-reverse osmosis of pollutants in sewages were described in Table 3. Unfortunately in this way cleaned effluents did not fulfil qualitative norms presented into the Directive of the Environment Minister from 24th July 2006. Concerning the conditions that should be at inserting sewages into waters or soil, and in the matter of substances particularly harmful to the environment aqueous (Log. Act 2006 number 137 item 984). They stated the exceeding of the acceptable concentration of ammoniacal + 3 nitrogen passed 2-ratio (22.4 mgNH4 /dm ) that is way the sewages should subjected to the stripping process before carry them to the natural receiver. 1760 Karolina Mielczarek et al

Table 3 The efficiency of cleaning in integrated system ultrafiltration-reverse osmosis coke-making sewages

Cleaned sewages Value Raw UF-PSF-16 RO-SE of coefficients Indicator Unit waste- for introduced Stage Stage water sewages to Value of removal of Value of removal of land waters pollutions [%] pollutions [%] 3 COD mgO2/dm 2754 2087.5 24.2 74.3 97.3 125 9TC mgC/dm3 616.9 515.1 16.5 29.3 94.3 ns. TOC mgC/dm3 390.7 340.3 12.9 12.1 96.4 ns. Ammoniacal + 3 nitrogen mgNH4 /dm 131.6 98.0 25.5 22.4 83.0 10 TC – Total carbon; TOC – total organic carbon; ns.– not standardized.

Conclusions

1. Coke-making sewages sequential system of ultrafiltration-reverses osmosis applied in investigations didn’t provide properly high degree of treatment. 2. From prepared five polymeric low-pressure filtration membranes the most profitable was a PSF-16 membrane showed itself for 16 % weight contents of the polymer in membrane solution. Since none of membranes assured the high level of coke-making industry wastewater purification the choice was determined by the size of the volumetric flux permeate, which in comparison with the membrane with the most clenched structure (PSF-17) was over three times higher (DP=3.0×105 Pa) and had highest relative permeability. 3. Sewages cleaned in the process of the reverse osmosis were characterized by too + 3 high concentration on the level, of ammonia 22.4 mgNH4 /dm . They should before carry them to the natural receiver, be subjected to the stripping process.

Acknowledgement

The researches were carried out by BW 401/201/08.

References

[1] Minhalma M. and De Pinho M.N.: Integration of nanofiltration/steam stripping for the treatment of coke plant ammoniacal wastewater. J. Membr. Sci. 2004, 242(1–2), 87–95. [2] Lai P., Zhao H., Wang Ch. and Ni J.: Advanced treatment of coking wastewater by coagulation and zero-valent iron processes. J. Hazard. Mater. 2007, 147(1–3), 232–239. [3] Minhalma M. and De Pinho M.N.: Development of nanofiltration/steam stripping sequence for coke plant wastewater treatment. Desalination 2002, 149(1–3), 95–100. [4] Zhang M., Tay J.H., Qian Y. and Gu X.S.: Coke plant wastewater treatment by fixed biofilm system for COD and NH3-N removal. Water Res. 1998, 32(2), 519–527. [5] Jianlong W., Xiangchun Q., Libo W., Yi Q. and Hegemann W.: Bioaugumentation as a tool to enhance the removal of refractory compound in coke plant wastewater. Process Biochem. 2002, 38(5), 777–781. Application of Polysulfone Membranes for Coke-Making Wastewater Treatment 1761

[6] Ghose M.K.: Complete physico-chemical treatment for coke plant effluents. Water Res. 2002, 36(5), 1127–1134. [7] Caetano A.T.: Existing industrial application: results and perpectives – Membrane Technology: application to industrial wastewater treatment. Kluwer Academic Publisher, Dordrecht 1995. [8] Mulder M.: The use of membrane processes in industrial problems. An introduction – Membrane Processes in separation and purification. Kluwer Academic Publisher, Dordrecht 1994. [9] Wiessner A., Remmler M., Kuschk P. and Stottmeister U.: The treatment of a disposited lignite pyrolysis wastewater by adsorption using activated carbon and activated coke. Colloids Surf. A, 1998, 139(1), 91–97.

MEMBRANY POLISULFONOWE W OCZYSZCZANIU ŒCIEKÓW KOKSOWNICZYCH

Instytut In¿ynierii Œrodowiska, Wydzia³ In¿ynierii i Ochrony Œrodowiska Politechnika Czêstochowska

Abstrakt: Ze wzglêdu na z³o¿ony i zmienny sk³ad œcieków koksowniczych strategia ich oczyszczania jest trudna do uogólnienia i wymaga prowadzenia tego procesu w uk³adach zintegrowanych, kojarz¹cych biologiczne i fizykochemiczne procesy jednostkowe. W artykule omówiono badania, których celem by³o okreœlenie efektywnoœci oczyszczania œcieków koksowniczych w uk³adzie kojarz¹cym ciœnieniowe techniki membranowe, a mianowicie ultrafiltracjê i odwrócon¹ osmozê. W procesie niskociœnieniowej filtracji zastosowano wytwarzane w laboratorium p³askie membrany polisulfonowe ró¿ni¹ce siê zwartoœci¹ struktury i porowatoœci¹. S³owa kluczowe: oczyszczanie œcieków koksowniczych, uk³ady zintegrowane, ciœnieniowe techniki membra- nowe, polisulfonowe membrany ultrafiltracyjne

ECOLOGICAL CHEMISTRY AND ENGINEERING A

Vol. 18, No. 12 2011

Justyna HACHO£1 and El¿bieta BONDAR-NOWAKOWSKA1

ECOLOGICAL RISK CLASSIFICATION IN THE REGULATED AND CONSERVED WATERCOURSES

KLASYFIKACJA RYZYKA EKOLOGICZNEGO W CIEKACH REGULOWANYCH I KONSERWOWANYCH

Abstract: The subject of the following study is ecological risk in regulatory and maintenance works conducted in small and medium-sized lowland watercourses. Risk has not been identified well enough. It results from the lack of the data to assess its level objectively. The following research presents a proposal of solving the problem. The results of the field work conducted between 2007 and 2008 on 10 Lower Silesian lowland watercourses form a basis for this analysis. The research included hydromacrophytes identification and the degree of the bottom coverage by these aquatic plants. The following study showed that in a result of regulatory and maintenance works quality and quantity alternations in aquatic plants communities were observed. The analysis of these alternations enabled assigning measures to the factors of considered risk. It served as a basis for describing the matrix of risk classification. Risk classification method suggested in the study may be useful in designing plans concerning ecological risk management and at the determination of the safety management rules in case of technical interferences in the watercourses. Keywords: aquatic vascular plants, ecological risk, maintenance works, watercourses regulation

Introduction

The research on regulatory and conservation works impact on the watercourses biocenosis show it is very variable. The most often effects of the following works are quality and quantity changes in plant and animal communities in the watercourse [1–8]. The level of these changes is dependent on both technical and environmental factors [5–7, 9, 10]. All these should be taken into account when planning regulation or

1 Institute of Environmental Development and Protection, Wroclaw University of Environmental and Life Sciences, pl. Grunwaldzki 24, 50–363 Wroc³aw, Poland, phone: +48 71 320 55 85, fax: +48 71 320 55 74, email: [email protected] 1764 Justyna Hacho³ and El¿bieta Bondar-Nowakowska conservation. In spite of knowledge base accessible in this matter there is no method allowing for biocenosis changes forecast being a result of technical works occurring in the river bed. The lack of such a tool results in the possibility of the assessment of the project after it has been done. Decisions taken by designers and contractors are assessed after the works completion. Taking into account that the effects of some decisions are irreversible for the environment it is crucial to take fast actions on solving the problem to assure environmental safety. The following research shows the proposal of using an assessment of ecological risk to cover the problem. The term “risk” defines a degree of exposure to harmful events and their possible consequences [11, 12]. Determination of the level of ecological risk in regulated and conserved watercourses serves a possibility of changes forecast in river beds ecosystems [13–15]. The aim of this study is the determination of the principle to assess and classify ecological risk of regulatory and conservation works basing on one element of watercourse ecosystem – hydromacrophytes. These water plants are good bioindicators of the quality of water environment [16–18]. Therefore, they are one of the basic factors considered in an assessment of the ecological state of flowing waters [19].

Study objects

Field work was performed during vegetation periods in 2007 and 2008 for 10 small and medium Lower Silesian watercourses. These watercourses were divided into 34 experimental sections 100 meters long each. Detailed characteristic of particular sections is showed in the Table 1. The sections had similar climatic (moderate, transitional between maritime and continental), geological (Foresudetic Monocline built of Permian and Trias rocks) and soil (Luvisols formed from loess and brown soil) conditions [20–22]. Adjacent field was used agriculturally with a domination of arable lands and grasslands. Most of experimental sections was not shadowed. Only some of them were slightly shadowed. Water in experimental sections was contaminated neither with urban or industrial wastes. Particular sections varied with the degree of antrophogenic transformation – 11 were located in the watercourses where conservatory works were done while other 13 were located in regulated watercourses. Each watercourse had one section where no works were conducted. In the following comparative analyses they served as reference points. Conservation works in the examined watercourses included: manual scything of scarps and riverside zones, mechanical elutriate of the bottom with removal of the aquatic plants and reparation of the scarps’ strengthening. Regulatory works showed deeper interference in the river bed itself. As a result of these works changes in parameters of vertical and cross-sections and the method of the banks protection occurred. In most cases these works were mechanically performed. Ecological Risk Classification in the Regulated and Conserved Watercourses 1765

Table 1 Study sections

Unmodified (U), Water- Number Bottom conserved (C) course Inclination Slope River of study width Substrate or regulated (R) depth of the slope protection section [m] section [m] 1 C 5 3.5 sand/gravel 1:1.5 fascines 2 C 3 2 gravel/stones 1:2 fascines Czarna 3 C 3 2 gravel/stones 1:2 fascines Woda 4 C 3 2 gravel/stones 1:2 fascines 5 C 3 2 gravel/stones 1:1 fascines 6 U 7 1.5 sand 1:1 non-protected 7 U 3 1.2 sand 1:1.5 non-protected 8 R 3 0.6 sand 1:3 fascines Dobra 9 R 6.8 2 sand 1:0.2 gabions 10 R 2 2 sand 1:2 stone coating 11 C 2 3 organic 1:1.5 non-protected Oleszna 12 C 2.4 1.36 organic 1:0.8 non-protected 13 U 1.5 2 organic 1:1.5 non-protected 14 U 7 2.06 sand 1:1.5 non-protected Olesnica 15 R 4 2.27 organic 1:1 fascines 16 U 8 1.8 organic 1:1 non-protected Orla 17 R 10 1.8 organic 1:0.5 fascines Potok 18 C 2 3.5 sand/stones 1:2 fascines Sulistro- 19 U 2 3.5 sand/stones 1:2 non-protected wicki 20 C 2 3.5 sand/stones 1:2 fascines 21 C 10 2.15 sand 1:2 fascines 22 U 5 2.15 sand 1:2 non-protected Smortawa 23 R 3 1.5 sand 1:2 fascines 24 R 3 1.5 sand 1:2 fascines 25 U 5 3 sand 1:1.5 non-protected Sleza 26 R 5 3 concrete 1:0.2 stone concrete 27 R 5 1.5 sand/gravel 1:1.5 stone coating 28 C 1.5 1.92 sand 1:1.5 non-protected Zalina 29 U 1.5 2.11 sand 1:1 non-protected 30 R 2 2 organic 1:1 fascines 31 R 3.3 1.4 sand 1:1.5 fascines 32 U 3 2.2 sand 1:2 non-protected Zurawka 33 R 3 2 sand 1:2 fascines 34 R 3 2.2 sand 1:2 fascines

Methods

In the framework of the following field work macrophytes species was identified on the examined sections and the degree of the bottom coverage with them was de- 1766 Justyna Hacho³ and El¿bieta Bondar-Nowakowska termined. All hydromacrophytes rooted in water for at least 90 % of the vegetation period and plants flowing naturally on the water surface or under it were taken into account. Five levels Braun-Blanquet scale was used for the determination of the density degree [23]. In order to assess species variety in the examined sections Shannon-Wiener Index – H [24] was calculated, considering both number of species and their coverage regularity [10]. Statistical analysis of the results was done using Statistica v. 9.1. programme. Risk level connected with conservation and regulatory works performance was defin- ed according to the following formula [11]:

R = P × S In this formula: P – stands for the probability of change of the hydromacrophyte environment as a result of the performed works, S – stands for the susceptibility of changes in the aquatic plant species composition. These factors were ascribed with different measurements. They were indicated by the field work results. In both cases 5 levels scales were used. Product of P and S parameters formed a basis of the risk level assessment. It was follow- ed by two parametric matrix of the risk assessment [25]. According to accepted scales, matrix was marked due to observed changes in the plants community concerning works conducted in the examined watercourse. Risk classification was based on this analysis.

Results and discussion

In the examined sections 20 species of aquatic macrophytes were determined altogether. This number is small in comparison with the results obtained from other authors in similar studies [5, 26, 27]. The small number of species of aquatic plants may be due to the fact that most watercourses underwent some kind of technical interference in the past as well as due to agricultural settings in the surroundings. Furthermore the small rivers have a lower species richness of aquatic plants than large rivers [26]. The following species were found: Alisma plantago-aquatica L., Berula erecta (Huds.) Coville., Butomus umbellatus L., Callitriche sp., Ceratophyllum demersum L., Elodea canadensis L., Glyceria Maxima (Hartm.) Holmb., Hydrocharis morsus ranae L., Myosotis palustris (L.) L. em. Rchb., Lemna minor L., Nuphar lutea (L.) Sibth. & Sm., Phalaris arundinacea L., Phragmites communis Trin., Potamogeton pectinatus L., Potamogeton filiformis Pers., Sagittaria sagittifolia L., Sparganium emersum Rehmann, Sparganium erectum L. em. Rchb. s.s., Spirodela polyrrhiza (L.) Schleid., Typha angustifolia L. According to Method of Macrophytes Rivers Assessment (MMOR) these species have a wide or medium wide ecological scale and low or medium index value – W measured at 1 or 2 [28]. Figure 1 shows statistic data referring to aquatic plants occurrence in the examined river sections. Ecological Risk Classification in the Regulated and Conserved Watercourses 1767

(a)MedianaPercentile 25–75 %Min.–Max (b) Mediana Medium ± Error std. Min.–Max 10 2.2 2.0 8 1.8 1.6 6 1.4 1.2 4 1.0 0.8 2 0.6 0.4

Shannon-Wiener Index 0 0.2

Numer of aquatic plant species 0.0 –2 –0.2 UnmodifiedConservedRegulated Unmodified Conserved Regulated

Fig. 1. Diversification of the number of species (a) and species variety Shannon-Wiener Index (b) in the examined sections

The highest mean number of species was observed in natural sections while in modified sections especially those which were regulated a lot less. It is widely believed that any technical interference within a watercourse’s channel adversely affects its biocoenosis [5, 29–33]. Values of the diversity Shannon-Wiener Index determined in compared sections were slightly different (Fig. 1b). In unmodified and conserved examined sections they were similar while in regulated watercourses they were considerably lower. The mean index value in unmodified, conserved and regulated river beds was calculated at 0.98; 0.95 and 0.5, respectively. Ecological risk factors were defined and classified using results of the following research. While assessing P factor, determining the possibility of alternations in the number of aquatic plants species, a strong relation with a range of performed works in the river bed was taken into account. Observations made during the research allowed to acknowledge a five degree scale.

Table 2 Scale of the changes occurrence probability in the aquatic plant communities in the river beds as a result of the watercourses regulation and maintenance works – P

Probability of changes occurence Point scale Description scale 1 Slope mowing, river bed elutriation with the removal of aquatic plant life Very low 2 Slope mowing, river bed elutriation with the removal of the aquatic plant life, reparation and Low the strengthening of fascine 3 Changes in cross-section parameters, modification of the scarps incline to 1:1.5, additional Medium strengthening of a riverbank’s foundation with fascine construction 4 Changes in cross-section parameters, the strengthening of scarps with stone or stone mattress High gabions 5 Changes in cross-section parameters, modification of watercourse’s vertical plane with hori- Very high zontal scarps, the strengthening of slopes with box gabions or retaining walls

Particular levels in the scale defining risk factor S – changes in the number of species as a result of works conduction were determined on the basis of comparison of species 1768 Justyna Hacho³ and El¿bieta Bondar-Nowakowska composition of macrophytes aquatic communities in the sections where technical works were and were not done. In the sections where no human interference was present 15 species (Fig. 2) were found and 10 out of 15 showed medium index value (W = 2) while other species revealed low value (W = 1). None section was defined with stenotopic species of a high index value (W = 3).

12

10

8 Alisma plantago-aquatica Berula erecta Butomus umbellatus 6 Elodea canadensis Lemna minor 4 Glyceria Maxima Nuphar lutea Myosotis palustris Sagittaria sagittifolia Phalaris arundinacea Sparganium emersum 2 Potamogeton pectinatus Spirodela polyrrhiza

Number of aquatic plant species Sparganium erectum Typha angustifolia 0 12 W

Fig. 2. Index value of the species occurring in unmodified sections

The following research enables to claim that the least severe effects of conducted works occurred in watercourses with wide ecological scale species. The more species with low index value the better they adapt to new, technically changed habitat conditions. On the basis of this relation the effects of conservatory and maintenance works were linked with aquatic macrophytes species composition in particular water- courses (Table 3). Table 3 The scale of the consequences of the watercourses regulation and maintenance works in the river bed for the aquatic plant communities – S

Susceptibility of changes of aquatic plant species composition Point scale Description scale 1 No hydromacrophytes in the watercourse Lack 2 1–3 species of hydromacrophytes with low and medium index values ofW=1orW=2in Mild the watercourse 3 More than 4 species of hydromacrophytes dominated with taxons of a low index value Moderate ofW=1inthewatercourse 4 More than 4 species of hydromacrophytes dominated with taxons of a medium index value Severe ofW=2inthewatercourse 5 Taxons with the high index value ofW=3inthewatercourse Very severe

Classifications of considered risk factors showed in Tables 2 and 3 formed a basis to compile ecological risk matrix in both maintained and conserved watercourses (Fig. 3). Ecological Risk Classification in the Regulated and Conserved Watercourses 1769

5 5 10152025

4 4 8 12 16 20

3 3691215

2 246810

Probability (P) 1 12345 123 4 5

Susceptibility of changes (S)

Fig. 3. A matrix of ecological risk in maintained and regulated watercourses

The following matrix shows the level of risk may be placed between 1 and 25 points. Information obtained during the field work were used for determination of point range for small risk – usually accepted, medium and high – unaccepted. Each section located in modified watercourses were defined with the area of matrix where observed changes in plants communities took place. This data was presented on both matrixes – for watercourses where conducted works comprised of the river bed conservation (Fig. 4) and for regulated watercourses (Fig. 5). Both matrixes were also marked with the direction of the observed changes referring to the number of species and Shannon- -Wiener Index.

5

4

3

Probability (P) == +=+–– 2 18, 20 1, 2, 3, 4, 5 ++ +++––

++ = + 1 11, 12 28 21 ++ + +

123 4 5

Susceptibility of changes (S)

Fig. 4. A matrix of ecological risk for maintained study sections [] – shows a trend of changes in the number of species: “+” – increase, “–” – decrease, “=” – no changes; 28 – the number of the examined section; [] – shows a trend of changes in the Shannon-Wiener Index: “+” – increase, “–” – decrease

Observed alternations in aquatic plants communities concerned enlarging or lowering the number of species and species biodiversity index. Directions of these changes were marked in the Figs. 4 and 5 as upper and lower index given above the number. They show corresponding examined section. In case of conservatory works the most common 1770 Justyna Hacho³ and El¿bieta Bondar-Nowakowska situations were those with a growth in both values (Fig. 4). It shows that properly executed maintenance work, including mowing the banks, the removal of plant life from the river bottom and its dredging, in fact does allow the watercourse to function properly, without causing a permanent loss of aquatic plant communities [1, 5, 14, 34–38]. Maintenance works do not cause the disappearance of islands and oxbow lakes, do not change the route of the river bottom and do not restrict the watercourse’s capacity during overflowing [4]. Regulated sections in most cases show that the result of river beds regulation is lowering the number of species of aquatic plants and the values of Shannon-Wiener Index. According to Gunkel [30] river regulation is a major controlling factor for the aquatic ecosystems. Conventional river control causes the changes in hydraulic characteristics of the river. The hydrological regime has been widely recognized as an important factor controlling colonization of streambeds by macrophytes [38]. If for these works risk assessment according to proposed method risk level would be contained in the right, upper matrix area (Fig. 5).

–– 5 26 9 ––

–– 4 27 10 ––

+–== –– – – – 3 30, 31, 33, 34 8, 15, 17, 23, 24 +––––+––+

Probability (P) 2

1

123 4 5

Susceptibility of changes (S)

Fig. 5. A matrix of ecological risk for regulated study sections [] – shows a trend of changes in the number of species: “+” – increase, “–” – decrease, “=” – no changes; 34 – the number of the examined section; [] – shows a trend of changes in the Shannon-Wiener Index: “+” – increase, “–” – decrease

Figure 6 shows the matrix where both maintenance and regulatory works were considered. It was a basis for ecological risk classification taking into account its 3 levels – low, medium and high risk (Fig. 7). The analysis of Fig. 7 reveals that the limits between fixed levels of risk are as follows: – low risk – £ 4 points, – moderate risk – 6–8 points, – high risk – ³ 12 points. Basing on the research it was impossible to determine the risk level for 5, 9 and 10 points. Therefore, it is necessary to conduct further research, including study objects Ecological Risk Classification in the Regulated and Conserved Watercourses 1771

Low risk – – 5 26 9 Medium risk – – High risk – – 4 27 10 – –

+–== –– – – – 3 30, 31, 33, 34 8, 15, 17, 23, 24 +––+ –– + – –

Probability (P) == +=+–– 2 18, 20 1, 2, 3, 4, 5 ++ +++––

++ = + 1 11, 12 28 21 ++ + +

1 2 3 4 5

Susceptibility of changes (S)

Fig. 6. The ecological risk classification based on the alternations of aquatic plants communities of maintained and regulated watercourses [] – shows a trend of changes in the number of species: “+” – increase, “–” – decrease, “=” – no changes; 28 – the number of the examined section; [] – shows a trend of changes in the Shannon-Wiener Index: “+” – increase, “–” – decrease

Unknown risk 5 51015 20 25 Low risk 4 4812 16 20 Medium risk 3 36912 15 High risk

2 246810

Probability (P) 1 12345 123 4 5

Susceptibility of changes (S)

Fig. 7. A matrix showing the three ecological risk levels of maintained and regulated watercourses with various composition of aquatic plant communities and the different degree of anthropogenic transformation. Conducted research and analyses should be considered as pilotage for the following project. Better recognition requires further observations performed in the communities of plants and animals in watercourses being a subject to water works.

Conclusions

1. The research conducted on 10 lowland watercourses found in Lower Silesia, where maintenance and regulatory works were done, showed these works resulted in alternations in the number of aquatic plants species and the index of their biodiversity. The range of performed works and ecological tolerance of the plants occurrence in watercourses had a big influence on the size of these changes. 1772 Justyna Hacho³ and El¿bieta Bondar-Nowakowska

2. The results of the following research allowed for determination and classification of ecological risk factors in five level scales – the possibility of alternations in species in the communities of hydromacrophytes and the level of these changes. 3. Based on the risk matrix, the ecological risk was then classified in three different levels. Low risk was determined for those values ranging from 1 to 4, medium risk was found between 6 and 8, while high risk was determined at values of 12 points or more. 4. The performed analysis found that maintenance works are connected with low or moderate risk, whereas regulatory works comprise of moderate or high risk area. 5. In order to limit adverse alternations of the aquatic plants community, a risk assessment should be performed when planning any maintenance or regulatory works. It requires a detailed environmental assessment of the river bed and a detailed analysis of the range of the planned works.

References

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KLASYFIKACJA RYZYKA EKOLOGICZNEGO W CIEKACH REGULOWANYCH I KONSERWOWANYCH

Instytut Kszta³towania i Ochrony Œrodowiska Uniwersytet Przyrodniczy we Wroc³awiu

Streszczenie: Przedmiotem pracy jest ryzyko ekologiczne w robotach regulacyjnych i konserwacyjnych wykonywanych na ma³ych i œrednich ciekach nizinnych. Ryzyko to nie jest jeszcze dobrze rozpoznane. Wynika to m.in. z braku podstaw do obiektywnej jego oceny. W pracy przedstawiono propozycjê rozwi¹zania tego problemu. Podstawê analizy stanowi¹ wyniki badañ terenowych prowadzonych w latach 2007–2008, na 10 nizinnych ciekach Dolnego Œl¹ska. Badania obejmowa³y identyfikacjê wystêpuj¹cych w korycie gatunków naczynio- wych roœlin wodnych oraz okreœlenie stopnia pokrycia przez nie dna. Wykaza³y one, ¿e w wykonanych prac zachodz¹ zmiany jakoœciowe i iloœciowe w zbiorowiskach naczyniowych roœlin wodnych. Analiza tych zmian pozwoli³a na przypisanie miar czynnikom rozpatrywanego ryzyka. Stanowi³o to podstawê opracowania macierzy klasyfikacji ryzyka. Zaproponowana w pracy metoda klasyfikacji ryzyka mo¿e byæ przydatna w opracowywaniu planów zarz¹dzania ryzykiem ekologicznym oraz przy okreœlaniu zasad zarz¹dzania bezpieczeñstwem œrodowiska przyrodniczego w przypadkach ingerencji technicznych w korytach cieków. S³owa kluczowe: ryzyko ekologiczne, naczyniowe roœliny wodne, roboty konserwacyjne, regulacja cieków Indexes

CONTENTS OF VOLUME 18 OF “ECOLOGICAL CHEMISTRY AND ENGINEERING A”

SPIS TREŒCI TOMU 18 MIESIÊCZNIKA CHEMIA I IN¯YNIERIA EKOLOGICZNA A”

1

1. Anita BIESIADA, Agnieszka NAWIRSKA, Alicja KUCHARSKA and Anna SOKÓ£- -£ÊTOWSKA – Chemical Composition of Pumpkin Fruit Depending on Cultivar and Storage ...... 9 2. Maria GAWÊDA and Zofia NIZIO£-£UKASZEWSKA – Effect of Lead Cumulation on Boron Content in Some Vegetables ...... 19 3. Aleksander GONKIEWICZ and Ma³gorzata LEJA – Influence of Auxin NAA and Ethephon on Yield Quality of Apple (Malus domestica Borkh.) ‘Sampion’ cv. .... 27 4. Krzysztof KLAMKOWSKI, Waldemar TREDER and Anna TRYNGIEL-GAÆ – Growth and Photosynthetic Activity of Cucumber as Influenced by Different Fertilization Regimes ...... 35 5. Tomasz KLEIBER and Andrzej KOMOSA – Effect of Vegetation Period on the Quantitative Relations Between Nutritive Components in the Leaves of Anthurium (Anthurium cultorum Birdsey) ...... 43 6. El¿bieta KOZIK, Stanis³awa SZCZEPANIAK, Alicja DOMINIAK and Przemys³aw KOZAK – Reaction of Corsican Hellebore (Helleborus argutifolius Viv.) to Differentiated Nitrogen Fertilization ...... 51 7. Maria LEJA, Iwona KAMIÑSKA and Katarzyna KULCZAK – Antioxidative Properties in Grapes of Selected Cultivars Grown in Poland ...... 59 8. Monika MA£ODOBRY, Monika BIENIASZ and Ewa DZIEDZIC – Yield Structure and Content of Some Components in Fruit of Six Strawberry Cultivars ...... 67 9. Zenia MICHA£OJÆ and Halina BUCZKOWSKA – Yield and Eggplant Fruit Quality (Solanum melongena L.) Dependent on Plant Training and Nitrogen Fertlization .... 73 10. Maria POBO¯NIAK and Adam ŒWIDERSKI – Initial Research on the Influence of the Colour of the Pea Leaves on the Infestation by Thrips ...... 83 11. Marek SIWULSKI, Agnieszka JASIÑSKA, Krzysztof SOBIERALSKI and Iwona SAS-GOLAK – Comparison of Chemical Composition of Fruiting Bodies of Some Edible Mushrooms Cultivated on Sawdust ...... 89 12. Sylwester SMOLEÑ and W³odzimierz SADY – Effect of Fertilization with Entec-26 and Ammonium Nitrate on the Changes in Selected Chemical Soil Properties after Carrot Cultivation ...... 97 13. Sylwester SMOLEÑ and W³odzimierz SADY – Effect of Nitrogen Fertilizers on the Change in Selected Chemical Characteristics of Soil after Carrot Cultivation . . 105 14. Anna SZOPIÑSKA and Maria GAWÊDA – Comparison of Carrot Quality Cultivated Using Conventional, Integrated and Organic Methods ...... 113 1778 Indexes

15. Wanda WADAS – Effect of Multinutrient Complex Fertilizers on Total Nitrogen and Nitrate(V) Content in the Tubers of Very Early Potato Cultivars ...... 123 16. Dorota WALKOWIAK-TOMCZAK, Ró¿a BIEGAÑSKA-MARECIK and El¿bieta RADZIEJEWSKA-KUBZDELA – Changes in Contents of Nitrates(V) and Nitrates(III) in Small Radish Packaged and Stored in Modified Atmosphere ...... 129 17. Renata WOJCIECHOWSKA and Piotr SIWEK – Effect of Film Used in Low Tunnels and for Plant Shading on Nitrate Metabolism in Celery Stalks ...... 139 18. Krzysztof WRAGA and Monika PLACEK – Use of Media Supplemented with Compost Containing Potato Pulp in Cultivation of Egyptian Star Cluster (Pentas lanceolata (Forssk.) Deflers) ...... 147

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19. Wojciech D¥BROWSKI – Effectiveness of Constructed Wetlands for Dairy Wastewater Treatment ...... 175 20. Julitta GAJEWSKA, Piotr JACAK and Leszek BABIÑSKI – Influence of Anoxic Condition on the Composition of Microorganisms Colonized a Contemporary Wood Samples in Archaeological Site in Biskupin ...... 183 21. Katarzyna IGNATOWICZ – Graveyard – Point Source Pollution of Natural Water by Pesticides ...... 191 22. El¿bieta JEKATIERYNCZUK-RUDCZYK – Changes of Phosphorus Compounds Concentrations in Waters of Lowland Catchments with Various Anthropopression Levels ...... 201 23. Aleksander KIRYLUK – Concentrations of Nitrates(V) in Well Waters in the Rural Areas of Podlasie Province and the Assesment of Inhabitants’ Health Risk ..... 207 24. Katarzyna KOWALCZEWSKA-MADURA, Renata DONDAJEWSKA and Ryszard GO£DYN – Seasonal Changes of Phosphorus Release from the Bottom Sediments of Rusalka Lake During the Restoration Process ...... 219 25. Tadeusz MOLENDA and Damian CHMURA – Seasonal Changes in Selected Physicochemical Parameteres of Saline Water Bodies (Case Study of Retention- -Dosing Reservoir “Brzeszcze”) ...... 225 26. Marcin NIEMIEC and Barbara WIŒNIOWSKA-KIELIAN – Assessment of Heavy Metal Pollution of Rainwaters Flowing Down the Road No. 4 Taken from Retention Reservoirs ...... 235 27. Zbigniew OSADOWSKI, Krystian OBOLEWSKI, Katarzyna GLIÑSKA-LEWCZUK and Olga LOPATOVSKAYA – Phytosociological and Ecological Analysis of Lower Section of the Kwacza River Before Restoration (Slupia River Basin) . . . 241 28. Jan PAWLUCZUK – Dynamics of Mineral Nitrogen Forms Content in Ombrophilous Organic Soil and in Underground Water ...... 255 29. Marie BJELKOVÁ, Martina VETROVCOVÁ, Miroslav GRIGA and Petr ŠKARPA – Effect of Sewage Sludge in Soil on Cd, Pb and Zn Accumulation in the Linum usitatissimum L...... 265 30. Stefan TSAKOVSKI and Vasil SIMEONOV – Classification of Different-Sized Aerosol Monitoring Data ...... 275 31. Miros³aw WIATKOWSKI – Influence of Msciwojow Pre-Dam Reservoir on Water Quality in the Water Reservoir Dam and Below the Reservoir ...... 289 32. Jaros³aw ZAWADZKI and Adam TARGOWSKI – Evaluation of Efficiency Selected Secondary Sampling Methods in Soil Studies ...... 301

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33. Teresa BANASZKIEWICZ – Ecological Aspect of Addition of Different Quantity of Enzyme Preparation to Mixture for Broiler Chickens ...... 335 Indexes 1779

34. Joanna D£U¯NIEWSKA and Ryszard MAZUREK – Impact of the Distance from Black Locust (Robinia pseudoacacia L.) Shelterbelts on Soil Microflora .... 341 35. Wojciech DMUCHOWSKI and Aneta Helena BACZEWSKA – Heavy Metal Content in City Tree Leaves Used for Compost Production ...... 347 36. Katarzyna GLEÑ – Effect of Foliar Fertilizers and Their Mixtures on Phytopathogenic Fusarium Fungi ...... 353 37. Joanna JARMU£-PIETRASZCZYK, Marta KAMIONEK and Przemys³aw WILKOWSKI – Effect of Long Fertilisation on Seasonal Variability of Occurrence of Entomopathogenic Nematodes and Fungi ...... 359 38. Marta KAMIONEK, Joanna JARMU£-PIETRASZCZYK, Anna M. BADOWSKA- -KOZAKIEWICZ and El¿bieta PEZOWICZ – Influence of Lead and Cadmium Ions on the Entomopathogenic Nematodes Steinernema feltiae Filipjev ...... 365 39. Monika KOWALSKA-GÓRALSKA, Piotr £AWA and Magdalena SENZE – Impact of Silver Contained in the Nano Silver Preparation on the Survival of Brine Shrimp (Artemia salina Leach 1819) Larvae ...... 371 40. Katarzyna MALINOWSKA – Content of Assimilation Pigments in the Photosynthectic Apparatus of Maple (Acer platanoides L.) Growing In Various Site Conditions of Szczecin ...... 377 41. Ryszard MAZUREK, Agnieszka JÓZEFOWSKA and Anna PIOTROWSKA – Influence of Distance from Black Locust (Robinia pseudoacacia) Shelterbelts on Dehydrogenase Activity in Arable Soils ...... 385 42. Ma³gorzata PACHOLEWSKA and Beata CWALINA – Resistance of Acidithiobacillus ferrooxidans to As(III) and Sb(III) Ions ...... 391 43. El¿bieta PEZOWICZ, Dorota TUMIALIS and Magdalena LEŒNIAK – Investigations on the Control of Sphaerolecanium prunastri F. (Homoptera, Coccinea) by Entomopathogenic Nematodes Under Contaminated Conditions ...... 397 44. Anna PODLEŒNA – Yielding Effect of Nitrogen and Sulfur at Pot Experiment Conditions with Winter Wheat (Triticum aestivum L.)...... 401 45. Olga POLESZCZUK, Dorota TUMIALIS and El¿bieta PEZOWICZ – Influence of Low Doses of Ionizing Radiation on Young and Two Week old Invasive Larvae of Entomopathogenic Nematodes (Heterorhabditidae, Steinernematidae)...... 407 46. Adam RADKOWSKI and Iwona RADKOWSKA – Estimation of the Quality and Nutritional Value of Hay from the Selected Individual Farms Located in the Region of Krakow-Czestochowa Jura. Part II. Content of Macroelements .... 413 47. Monika RAJKOWSKA, Kamila POKORSKA and Magdalena HOLAK – Relationships Between Macro- and Microelements and Heavy Metals in Selected Organs of Rudd (Scardinius erythrophthalmus L.) from Lakes Miedwie and Zelewko ...... 419 48. Dariusz ROPEK and Krzysztof FR¥CZEK – Effect of the Solid Waste Landfill in Tarnow on the Occurrence of Beneficial Entomofauna on Horse Bean ...... 425 49. Pawe³ SKONIECZEK and Józef KOC – Role of Retention Reservoir in Sodium Migration from Agricultural and Afforested Catchment Areas ...... 435 50. Ewa STANIS£AWSKA-GLUBIAK and Jolanta KORZENIOWSKA – Tolerance of White Mustard (Sinapsis alba L.) to Soil Pollution with Several Heavy Metals . . . 445 51. Karolina STEINDOR and Bernard PALOWSKI – Cadmium and Lead Accumulation Patterns in Organs of Chosen Urban Tree Species ...... 453 52. Barbara ŒCIGALSKA and Bernadetta £ABUZ – Content of Heavy Metals in the Top Layer of Soils with Triticale Growing Thereon in One-Crop Systems ...... 461 53. Maciej WALCZAK and Aleksandra BURKOWSKA – UVB Radiation Impact on Activity of DNA and Cellular Protein Synthesis of Water Environment Bacteria ...... 469

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54. Jakub BEKIER, Jerzy DROZD and Micha³ LICZNAR – Nitrogen Transformations in Composts Produced from Municipal Solid Wastes ...... 497 1780 Indexes

55. Jacek CZEKA£A – Influence of Long-Term Sprinkling Irrigation and Nitrogen Fertilisation on Soil Nitrogen Content of a Cereal Crop Rotation ...... 507 56. Franciszek CZY¯YK and Agnieszka RAJMUND – Quantity of Nitrogen Deposited in Soil as Precipitated from Atmosphere in the Wroclaw Area during 2002–2007 . . . 515 57. Tadeusz FILIPEK and Pawe³ HARASIM – Nitrogen Content and Amino Acids Protein Composition of Grain of Winter Wheat Foliar Fertilized with Urea and Microelements Fertilizers ...... 523 58. Stefan GRZEGORCZYK, Kazimierz GRABOWSKI and Jacek ALBERSKI – Nitrogen Accumulation by Selected Species of Grassland Legumes and Herbs ...... 531 59. Gra¿yna HARASIMOWICZ-HERMANN and Janusz HERMANN – Nitrogen Bioconversion and Fodder Protein Recovery from Distillery Spent Wash ...... 537 60. Czes³awa JASIEWICZ and Agnieszka BARAN – Comparison of the Effect of Mineral and Organic Fertilization on the Composition of Amino Acids in Green Biomass Maize ...... 545 61. Andrzej KOCOWICZ and El¿bieta JAMROZ – Carbon and Nitrogen Content of Mountain Meadow and Forest Podzols and Brown Acid Soils ...... 553 62. Urij Anatoljevich MAZHAJSKIJ, Tatjana Mihajlovna GUSEVA, Andrej Valerjevich ILJINSKIJ, Svetlana Valerjevna ANDRIYANEC and Ekaterina Sergeevna GUSEVA – Influence of Heavy Metals on Microorganisms Taking Part in the Circulation of Nitrogen in Soil ...... 563 63. Zenia MICHA£OJÆ – Influence of Varied Doses and Forms of Microelements and Medium on Nitrate(V) and (III) Content in Lettuce ...... 571 64. Anna MIECHÓWKA, Micha³ G¥SIOREK, Agnieszka JÓZEFOWSKA and Pawe³ ZADRO¯NY – Content of Microbial Biomass Nitrogen in Differently Used Soils of the Carpathian Foothills ...... 577 65. Lidia OKTABA and Alina KUSIÑSKA – Mineral Nitrogen in Soils of Different Land Use ...... 585 66. Joanna ONUCH-AMBORSKA – Effect of Different Doses of Nitrogen on Soil Quality and Yield of Plants Grown in the Land Recultivated after Sulphur Mining ..... 593 67. Tomasz SOSULSKI and Marian KORC – Effects of Different Mineral and Organic Fertilization on the Content of Nitrogen and Carbon in Soil Organic Matter Fractions ...... 601 68. Tomasz SOSULSKI and Stanis³aw MERCIK – Dynamics of Mineral Nitrogen Movement in the Soil Profile in Long-Term Experiments ...... 611 69. Tamara PERSICOVA and Natalia POSHTOVAYA – Effectiveness of Bacterial Preparations and Plant Growth Regulators in the Separate and Mixed Crops of Oats, Spring Wheat and Narrow-Leaved Lupine Depending on Level of Nitrogen Nutrition ...... 619 70. Ewa SPYCHAJ-FABISIAK, Jacek D£UGOSZ and Krzysztof PI£AT – Spatial Variability of Total Nitrogen in the Surface Horizon at the Production Field Scale ...... 629 71. Alojzy WOJTAS, Ma³gorzata D¥BEK, Gra¿yna PIOTROWSKA and Tadeusz MALINOWSKI – Nitrogen in Water from Wells ...... 637

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72. El¿bieta BOLIG£OWA – Impact of Abiotic Factors on Fusarium Mycotoxin Occurrence in Cereal Grain ...... 665 73. Aldona CI¥G£O, Anna KUCZKOWSKA-KUNIAR, W³adys³aw ZAMACHOWSKI, Robert STAWARZ and Grzegorz FORMICKI – Accumulation of Heavy Metals at Early Stages of Phrynohyas resinifictrix (Goeldi) Ontogenesis ...... 673 74. Joanna D£U¯NIEWSKA and Ryszard MAZUREK – Effect of Extracts of Soils from Various Distances from Black Locust (Robinia pseudoaccacia) Shelterbelts on Trichoderma Fungi ...... 679 75. Krzysztof FR¥CZEK and Dariusz ROPEK – Impact of the Municipal Landfill Site on Bacteria Participating in Transformation of Soil Nitrogen ...... 685 76. Micha³ G¥SIOREK – Heavy Metals in Soils of Henryk Jordan Park in Krakow . . . 697 Indexes 1781

77. Joanna JARMU£-PIETRASZCZYK and Joanna PIASECKA – Effect of Active Compounds from Pesticides Applied to Soil on the California Earthworm Eisenia fetida ...... 703 78. Marta KANDZIORA-CIUPA, Ryszard CIEPA£ and Aleksandra NADGÓRSKA-SOCHA – Biomonitoring of Heavy Metals in the Bieszczady National Park Using Soil and Fagus sylvatica L. Leaves ...... 709 79. Ma³gorzata K£YŒ – Emigration Acitivity of Rice Weevil Sitophilus oryzae L. (Coleoptera, Curculionidae) in Conditions of Reduced Temperature ...... 717 80. Józef KOC and Marcin DUDA – Role of the Retention Reservoir in Szabruk for the Chlorine Ion Migration from Its Agricultural Catchment ...... 723 81. Anna KOCOÑ – Phosphorus Farming of Morphologically Different Pea (Pisum sativum) Varieties in Potassium Deficiency Soil Conditions ...... 731 82. Monika KOWALSKA-GÓRALSKA, Magdalena SENZE and Rafa³ SZA£ATA – Influence of Mine Water on Water Quality of Pelcznica River ...... 737 83. Monika KOWALSKA-GÓRALSKA and Tomasz SKWARKA – Bioaccumulation of Selenium in Chosen Water Plant from the Drawa River ...... 743 84. Kornelia KUCHARSKA, El¿bieta PEZOWICZ and Dorota TUMIALIS – Mortality and Pathogenic Properties of Heterorhabditis bacteriophora (Poinar 1976) from Nematop Biopreparation after Contact with an Insecticide ...... 749 85. Tomasz KUNIAR, Dariusz ROPEK and Tadeusz LEMEK – Impact of Multi- -Walled Carbon Nanotubes on Viability and Pathogenicity of Enthomopathogenic Nematodes ...... 757 86. Katarzyna MALINOWSKA, Ma³gorzata MIKICIUK, Jacek WRÓBEL and Ewa CZE¯YK – Influence of Cadmium on Physiological Parameters of Clone Jorr of Basket Willow (Salix viminalis L.) from Aquatic Cultures ...... 763 87. Ryszard MAZUREK and Pawe³ ZADRO¯NY – Cadmium in Soils of the Ojcow National Park ...... 771 88. Grzegorz MIKICIUK and Ma³gorzata MIKICIUK – Influence of a Polymer Supersorbent on Selected Physiological Features of Strawberry ...... 777 89. Aleksandra NADGÓRSKA-SOCHA, Ryszard CIEPA£ and Marta KANDZIORA- -CIUPA – Bioindication of Heavy Metals Pollution in the Towns: Bedzin and Czeladz ...... 785 90. El¿bieta PISULEWSKA, Ryszard PORADOWSKI and Robert WITKOWICZ – Effect of Sowing Density on the Yield and Chemical Composition of Oat Grains ...... 793 91. Olga POLESZCZUK, El¿bieta PEZOWICZ and Dorota TUMIALIS – Retrieval Irradiated Entomopahthogenic Nematodes – Steinernema feltiae (Filipiev, 1934) from the Soil ...... 801 92. Monika RAJKOWSKA and Miko³aj PROTASOWICKI – Distribution of Selected Metals in Bottom Sediments of Lakes Insko and Wisola (Poland) ...... 805 93. Dariusz ROPEK and Krzysztof FR¥CZEK – Impact of Municipal Landfill Site in Tarnow on the Occurrence of Beneficial Beetles ...... 813 94. Barbara SKWARY£O-BEDNARZ – Influence of the Contents of Total Forms of Lead on the Number of Selected Groups of Microorganisms in the Forest Soils of the Protected Zone in the Roztocze National Park ...... 821 95. Maciej WALCZAK and Aleksandra BURKOWSKA – UV Radiation Impact on Enzymatic and Respiratory Activity of Neustonic and Planktonic Bacteria ..... 827

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96. Katarzyna JAROMIN, Alia JLILATI, Marcin WIDOMSKI and Grzegorz £AGÓD – Materials, Exploatation Manners and Roughness Coefficient in Gravitational Sanitation Conduits ...... 853 1782 Indexes

97. Grzegorz £AGÓD, Henryk SOBCZUK, Zbigniew SUCHORAB and Marcin WIDOMSKI – Flow Parameters Effects on Aerobic Biodegradation of Pollutants in Sewer System ...... 865 98. Zbigniew SUCHORAB, Agnieszka ¯ELAZNA and Henryk SOBCZUK – Water Content Measurement of Building Materials Using Surface TDR Probe ...... 877 99. Lidia WOLNY and Anna KORZEKWA-WOJTAL – Effect of Polyelectrolyte Dose on the Characteristics of Sewage Sludge in Sedimentation Processes in Small-Size Wastewater Treatment Plants ...... 887 100. Anna ŒWIERCZYÑSKA, Jolanta BOHDZIEWICZ and Magdalena AMALIO-KOSEL – Activity of Activated Sludge Microorganisms in the Co-Treatment of the Leachates in the SBR Bioreactor ...... 895 101. Mariusz DUDZIAK – Effect of the Contact Angel on the Effectiveness of Mycoestrogens Removal from Water Using Nanofiltration Membranes ...... 903 102. Beata ZA£ÊSKA-CHRÓST, Lech SMOCZYÑSKI and Regina WARDZYÑSKA – Treatment of Wastewater from the Pulp and Paper Industry by Electrocoagulation in a Static System ...... 911 103. Iwona ZAWIEJA, Pawe³ WOLSKI and Lidia WOLNY – Recovering of Biogass from Waste Deposited on Landfills ...... 923 104. Anna ZWODZIAK, Izabela SÓWKA, Maria SKRÊTOWICZ, Anna WOROBIEC, Alicja NYCH, Jerzy ZWODZIAK and Rene VAN GRIEKEN – PM10, PM2.5 and PM1.0 Indoor and Outdoor Concentrations and Chemical Composition in School Environment ...... 933 105. Agnieszka BARAN, Marek TARNAWSKI and Czes³awa JASIEWICZ – Assessment of the Content and Solubility of Heavy Metals in Bottom Sediments of the Chancza Reservoir ...... 941 106. Petr ŠKARPA – Monitoring the Changes in Total Contents of Manganese, Copper and Zinc in Soils from Long-Term Stationary Experiments ...... 951 107. Krystyna PRZYBULEWSKA, Sylwia MICHA£OWSKA, Magdalena B£ASZAK and Anna STOLARSKA – Growth of Soil Fungi on Culture Media Contaminated with Selected Herbicides ...... 959 108. Agata ŒWIÊCI£O – Effect of Pesticide Preparations and Indoleacetic Acid on Yeast Saccharomyces cerevisiae Cells ...... 967 109. Alina KOWALCZYK-JUŒKO – Properties of Ash in the Combustion of Selected Energy Crops ...... 973 110. Krystyna HOFFMANN, Józef HOFFMANN and Filip DONIGIEWICZ – Influence of Concentration of Phosphoric Acid on Obtained Fodder Phosphate Quality .... 983 111. El¿bieta HUZAR, Alicja WODNICKA and Ma³gorzata DZIÊCIO£ – Analysis of Volatile Compounds in Nail Polish Removers as a Criterion of Health Hazard Determination and Commodity Evaluation ...... 991

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112. Beata GRYGIERZEC – Effect of Nitrogen Fertilization on the Quantity of Seed Yield of Selected Poa pratensis L. Cultivars ...... 1019 113. Barbara HERMAN, Robert BICZAK and Piotr RYCHTER – Root Celery Reaction on NaCl and CaCl2 Salinity ...... 1025 114. Piotr KACORZYK – Effect of the Way of Utilization and the Level of Fertilization on the Quality of Leachate Water. Part I. The Concentration of Mineral Components in Leachate Water ...... 1033 115. Miros³aw KASPERCZYK, Wojciech SZEWCZYK and Piotr KACORZYK – Estimation of the Persistence of Liming Action on the Mountain Meadow ...... 1041 116. Monika KOWALSKA-GÓRALSKA and Magdalena SENZE – Selenium Concentration in Various Carp (Cyprinus carpio L.) Organs ...... 1047 117. Monika KOWALSKA-GÓRALSKA, Tomasz SKWARKA and Jerzy FEDORSKI – Selenium Content in Hard and Soft Hair of Silesian and Holstein Race Horses . . 1053 Indexes 1783

118. Helena KUBICKA, Agnieszka PYZA, Aneta WOLSKA-SOBCZAK and Wojciech DMUCHOWSKI – Content of Selected Elements in Seedlings of Inbred Lines of Winter Rye (Secale cereale L.)...... 1059 119. Kornelia KUCHARSKA, El¿bieta PEZOWICZ, Dorota TUMIALIS and Mi³os³awa BARKOWSKA – Effect of Silver Nanoparticles on the Mortality and Pathogenicity of Entomopathogenic Nematodes ...... 1065 120. Bogdan KULIG, Jan KO£ODZIEJ, Andrzej OLEKSY, Marek KO£ODZIEJCZYK and Aleksandra SAJDAK – Influence of the Weather Conditions on Faba Bean Yielding ...... 1071 121. Anna LORENC-KOZIK, El¿bieta PISULEWSKA and Krzysztof GONDEK – Impact of Weather Conditions on Chemical Composition of the Seeds of Three Soybean Cultivars ...... 1079 122. Piotr MALCZYK and Magdalena RYDLEWSKA – Spatial Variability of Total Mercury Content in Surface Horizon of Soils of Gnieznienskie Lakeland Area .... 1087 123. Katarzyna MALINOWSKA, Barbara MARSKA and Sylwia STEPANIUK – Content of Assimilation Dyes and Water Balance in Common Dandelion Found Nearly Chemical Works “Police” ...... 1093 124. Joanna MATUSKA and Marta KAMIONEK – Invasiveness of the Entomopathogenic Nematodes Steinernema feltiae (Filipjev 1934) Isolated from Various Habitats in Poland ...... 1101 125. Ma³gorzata MIKICIUK, Katarzyna MALINOWSKA and Agnieszka KOCHANEK – Effect of an Increased Concentration of Sodium Chloride on Some Physiological Features of Lettuce (Lactuca sativa var. capitata) ...... 1105 126. Adam RADKOWSKI and Iwona RADKOWSKA – Effect of Microelement Fertilization on the Quality and Nutritional Value of the Meadow Sward Hay. Part I. The Content of Organic Components and Nutritional Value ...... 1111 127. Dariusz ROPEK and Piotr KACORZYK – Biodiversity of Soil Fauna Depending on Vegetal Cover and Fertilization ...... 1117 128. Wiera S¥DEJ and Anna NAMIOTKO – Nitrates(V) Content in Potato Tubers Cultivated Under Various Fertilization Systems ...... 1123 129. Barbara SKWARY£O-BEDNARZ – Contents of Forms of Lead in the Soils of the Protected Zone in the Roztocze National Park and Adjacent Production Areas ...... 1131 130. Dorota TUMIALIS, Marta KAMIONEK, Anna MAZURKIEWICZ and El¿bieta PEZOWICZ – Effect of Different Factors on the Nematode Heterorhabditis megidis (Poinar, Jackson and Klein, 1987) – Mutualistic Bacteria Photorhabdus luminescens (Thomas and Poinar, 1979) In Vitro Cultures ...... 1139 131. Ma³gorzata W£ODARCZYK – Kinetics of Releasing Herbicide Metazachlor from Hydrogel Microcapsules to Aquatic Environments ...... 1147

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132. Monika ARASIMOWICZ, Barbara WIŒNIOWSKA-KIELIAN and Marcin NIEMIEC – Post-Effect of Bottom Sediment Addition to the Substratum on Chemical Composition of White Mustard (Sinapis alba L.) Biomass. Part 1. Macroelements Content ...... 1175 133. Agnieszka BARAN and Czes³awa JASIEWICZ – Accumulation of Trace Elements in Black and Green Teas: Characteristics of Leaves and Brews ...... 1185 134. Dariusz BORUSZKO – Aplication of Heavy Metals and Nutrients into Natural Environment with Sewage Sludge ...... 1193 135. Wojciech D¥BROWSKI – Removal of Organic and Biogenic Compounds from Reject Water with Constructed Wetlands ...... 1203 136. Jean B. DIATTA, Katarzyna PRZYGOCKA-CYNA, Maria BIBER and Remigiusz £UKOWIAK – Assessment of Heavy Metals Contamination in Recreational Parks of Poznan ...... 1211 1784 Indexes

137. Katarzyna IGNATOWICZ and Tomasz BREÑKO – Concentration of Heavy Metals in Compost Produced from Municipal Sewage Sludge for Natural Reuse ...... 1219 138. Monika JAKUBUS and Jacek CZEKA£A – Influence of Composting Sewage Sludge on Change in Lead Content in Sequentially Separated Fractions ...... 1227 139. Czes³awa JASIEWICZ, Agnieszka BARAN and Peter KOVÁÈIK – Heavy Metal Contents and the Sanitary State as an Assessment of Radish (Raphanus sativum L.) Quality ...... 1237 140. Hanna JAWORSKA and Halina D¥BKOWSKA-NASKRÊT – Total Content of Mercury in the Soils of the Surroundings of Lafarge-Cement Plant in Malogoszcz ...... 1245 141. Micha³ KOPEÆ and Krzysztof GONDEK – Effect of 40-Year Diversified Fertilizer Experiment on Changes in Mercury Content in Grassland (Czarny Potok) ..... 1251 142. Joanna KOSTECKA and Mariola GARCZYÑSKA – Influence of Selected Insecticides on Vermicomposting of Wastes with Participation of the Earthworm Dendrobaena veneta ...... 1263 143. Stanis³aw KOWALIK and Jerzy WÓJCIK – Analysis of Chemical Properties of the Incineration Wastes and the Possibilities of Their Biological Reclamation . . . 1271 144. Krzysztof KUD – Influence of Alluvial Processes on the Circulation of Manganese and Lithium in Riparian Environments ...... 1279 145. Piotr MALCZYK and Magdalena RYDLEWSKA – Properties of Soils Surrounded Trzuskawica Lime Plant Industry S.A., Department of Kujawy ...... 1287 146. Lilla MIELNIK, Jacek CZEKA£A, Ryszard PIOTROWICZ and Piotr KLIMASZYK – Occurrence of Some Heavy Metals in Bottom Sediments of Lobelia Lakes .... 1293 147. Pawe³ MUSZYÑSKI – Sorption of the Surfactant Hyamine 1622 in Soils ..... 1301 148. Barbara PATORCZYK-PYTLIK and Aldona ZIMOCH – Content of Selenium in Alfalfa in Dependence of Its Dose and the Type of Soil ...... 1313 149. Janina PIEKUTIN – Surface Water Polluted with Petroleum-Derivative Substances in Podlasie Region ...... 1321 150. Wiera S¥DEJ and Anna NAMIOTKO – Content of Copper, Zinc and Manganese in Soil Fertilized with Municipal Solid Waste Composts ...... 1327 151. Miros³aw SKORBILOWICZ – Changes in Water Quality of Melioration Systems in Upper Narew River Catchment ...... 1339 152. Petr ŠKARPA, Lubica POSPÍŠILOVÁ, Marie BJELKOVÁ, Karel FIALA and Jaroslav HLUŠEK – Effect of Organic Matter and pH on the Mobility of Some Heavy Metals in Soils of Permanent Grasslands in the Foothills of the Hruby Jesenik Mts ...... 1347 153. Monika TABAK and Barbara FILIPEK-MAZUR – Formation of Maize Yield as a Result of Fertilization with Organic Materials ...... 1355 154. Józefa WIATER, Anna SIEMIENIUK and Joanna SZCZYKOWSKA – Influence of Low Retention Reservoir on Water Quality of Suprasl River ...... 1363 155. Miros³aw WYSZKOWSKI and Agnieszka ZIÓ£KOWSKA – Effect of Compost, Bentonite and CaO on Some Properties of Soil Contaminated with Petrol and Diesel Oil ...... 1373

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156. Ma³gorzata HAWROT-PAW, Beata SMOLIK and Agnieszka KAMIENIECKA – Preliminary Study on the Efficiency of Biodiesel Biological Decomposition with Autochthonous Soil Microflora ...... 1401 157. Krystyna HOFFMANN and Marta HUCULAK-M¥CZKA – Evaluation of Concentration of Humic Substances in Selected Raw Materials and Wastes ...... 1407 158. Hanna JAWORSKA and Halina D¥BKOWSKA-NASKRÊT – Profile Distribution and Mobility of Lead in Selected Arable Soils from Pradolina Glogowska ..... 1417 Indexes 1785

159. Jolanta JANKOWSKA, Gra¿yna Anna CIEPIELA, Roman KOLCZAREK and Kazimierz JANKOWSKI – Occurrence of Cadmium in Herbs Growing on Grassland Located Near the Highway ...... 1425 160. Andrzej JAGUŒ – Assessment of Trophic State of Inland Water (the Case of the Sola Cascade Dam Reservoirs) ...... 1433 161. Katarzyna SOBCZYÑSKA-WÓJCIK and Ma³gorzata RAFA£OWSKA – Assessment of Restored Water Bodies in a River-Lake System Based on Phosphorus Concentrations ...... 1441 162. Ewa JURKIEWICZ-KARNKOWSKA and El¿bieta BIARDZKA – Chemical and Granulometric Parameters of Bottom Sediments in the Assessment of Floodplain Water Bodies of the Lower Bug River ...... 1457 163. Alia JLILATI, Katarzyna JAROMIN, Marcin WIDOMSKI and Grzegorz £AGÓD – Some Models of Sediments Transport in Gravitational Sanitation Systems ..... 1467 164. Anna GORCZYCA, Piotr JANAS and Marek J. KASPROWICZ – Impact of the Pulsed High Magnetic Field on Fusarium culmorum (W.G. Smith) Sacc...... 1477 165. Zdzis³aw CIEÆKO, Tomasz NAJMOWICZ and Miros³aw WYSZKOWSKI – Soil Pollution with Arsenic Versus the Concentration of Magnesium in Plants ...... 1485 166. Miros³aw WYSZKOWSKI and Maja RADZIEMSKA – Effect of Some Substances on Content of Selected Components in Soils Contaminated with Chromium ..... 1497 167. Czes³awa JASIEWICZ, Marek MADEYSKI , Marek TARNAWSKI and Agnieszka BARAN – Effect of Bottom Sediment Supplement to Soil on Yield and Chemical Composition of Maize ...... 1505 168. Janina GOSPODAREK and Katarzyna GLEÑ – Influence of Heavy Metals in Soil Upon Broad Bean (Vicia faba L.) Seed Infection by Diseases and Pests ...... 1515 169. Adam RADKOWSKI and Iwona RADKOWSKA – Estimation of the Nutritional Value of Hay from Selected Individual Farms in the Region of Krakow-Czestochowa Jura. Part I. The Content of Organic Compounds and Nutritional Value ...... 1521 170. Anna CHRZAN and Maria MARKO-WOR£OWSKA – Content of Selected Traffic Pollution in Soil and Pedofauna Near Busy Traffic Roads in Krakow ...... 1527 171. Barbara FILIPEK-MAZUR and Monika TABAK – Heavy Metals Availability in Soils Exposed to Traffic Pollution ...... 1533 172. Piotr KACORZYK – Effect of the Way of Utilization and the Level of Fertilization on the Quality of Leachate Water. Part II. The Loads of Components Carried with Leachate Water ...... 1539 173. Magdalena SENZE, Monika KOWALSKA-GÓRALSKA and Iwona CZY¯OWICZ – Bioaccumulation of Aluminium in the Aquatic Environment of the Dobra River in Wroclaw ...... 1545 174. Leszek B. ORLIKOWSKI, Magdalena PTASZEK, Aleksandra TRZEWIK and Teresa ORLIKOWSKA – Occurrence of Phytophthora Species in Rivers, Canals and Water Reservoirs in Relation to Its Location, Seasonal Analysis and Fungicide Residues . . 1551 175. Mariola GARCZYÑSKA and Joanna KOSTECKA – Influence of Dimilin 25 WP on Characteristics of Earthworm Eisenia fetida Sav., Vermicomposting Organic Waste ...... 1557 176. Janina GOSPODAREK – Residual Effect of Soil Contamination with Heavy Metals on Sitona sp. Beetles Feeding on Broad Bean (Vicia faba L.)...... 1565 177. Joanna JARMU£-PIETRASZCZYK, Marta KAMIONEK and Ines KANIA – Occurrence of Entomopathogenic Fungi in Selected Parks and Urban Forests Of The Warsaw District Ursynow ...... 1571 178. Joanna MATUSKA-£Y¯WA and Marta KAMIONEK – Morphometric Changes in Heterorhabditis megidis (Poinar, Jackson and Klein 1987) After Different Contact with Lead(II) Ions ...... 1575 1786 Indexes

12

179. Marcin KOZAK, W³adys³aw MALARZ, Andrzej KOTECKI, Waldemar HELIOS and Joanna GÓRA – Follow-up Effect of Hilling on Growth and Yielding of Miscanthus (Miscanthus x giganteus Greef et Deu.) ...... 1599 180. El¿bieta SACA£A – Miscanthus – Unusual Grass: Biochemical and Physiological Characteristic: A Review ...... 1615 181. Anna GORCZYCA and Marek J. KASPROWICZ – Initial Research on the Effect of the Nanogro Plant Growth Stimulator on Fusarium culmorum (W.G. Smith) Sacc. . 1625 182. Marcin SIDORUK, Andrzej ROCHWERGER, El¿bieta SKORBI£OWICZ and Miros³aw SKORBI£OWICZ – Effect of Catchment Area Use on Lead and Zinc Accumulation in the Bottom Deposits of Lakes Ardung and Bukwald ...... 1633 183. Zdzis³aw CIEÆKO, Miros³aw WYSZKOWSKI and El¿bieta ROLKA – Aluminium Concentration in Plants Depending on Soil Contamination with Cadmium ..... 1641 184. Janina GOSPODAREK – Effect of Soil Contamination with a Mixture of Heavy Metals on Broad Bean (Vicia faba L.) Seed Quality ...... 1651 185. Pawe³ WOLSKI, Iwona ZAWIEJA and Lidia WOLNY – Impact of Temperature on Viscosity of Sewage Sludge After Conditioning ...... 1659 186. Teresa RAUCKYTE-¯AK and S³awomir ¯AK – Changeability of the Speciation Forms of Heavy Metals in Soil Subject to Many Years of Fertilization with Wastewaters from Vegetable Fat Production ...... 1667 187. Beata GRYGIERZEC – Effect of Nitrogen Fertilization on Seed Production of Lolium perenne L. Turfgrass Cultivars ...... 1675 188. Joanna KOSTECKA and Grzegorz P¥CZKA – Kitchen Wastes as a Source of Nitrogen and Other Macroelements According to Technology of Vermiculture . . . 1683 189. Teresa RAUCKYTE-¯AK and Bo¿ena SZEJNIUK – Influence of 1,3,4-Thiadiazole Derivatives on the Biological Activity of the Selected Environmental Bacteria .... 1691 190. Krystyna PRZYBULEWSKA, Anna STOLARSKA and Daria G£¥BOWSKA – Change of Proline Content in Selected Soil Fungi as Affected by Osmotic Stress ...... 1705 191. Joanna JARMU£-PIETRASZCZYK, Marta KAMIONEK and Ewelina MALINOWSKA – Occurrence of Entomopathogenic Fungi and Nematodes on Pastures in Central and Northern Poland ...... 1711 192. Wiera MICHALCEWICZ, S³awomir STANKOWSKI, Ma³gorzata GA£CZYÑSKA and Marzena GIBCZYÑSKA – Successive Influence of Fluidal Ashes on General Number of Bacteria, Actinomycetes and Fungi in Pot Experiments .... 1715 193. Katarzyna GLEÑ – Effect of Foliar Fertilizers and Their Mixtures on Phytopathogenic Fusarium Fungi ...... 1721 194. Ma³gorzata NABRDALIK and Katarzyna GRATA – Influence of the Culture Conditions on Lipolytic Activity of Bacillus cereus and Bacillus mycoides ..... 1727 195. Adam RADKOWSKI – Effect of Microelement Fertilization on the Quality and Nutritional Value of the Meadow Sward Hay. Part II. The Content of Macroelements ...... 1737 196. Anna KWIECIÑSKA and Krystyna KONIECZNY – Recovery of Industrial Water from Pig Liquid Manure by Means of Membrane Techniques ...... 1743 197. Karolina MIELCZAREK, Jolanta BOHDZIEWICZ and Anna KWARCIAK- -KOZ£OWSKA – Application of Polysulfone Membranes for Coke-Making Wastewater Treatment ...... 1751 198. Justyna HACHO£ and El¿bieta BONDAR-NOWAKOWSKA – Ecological Risk Classification in the Regulated and Conserved Watercourses ...... 1763 AUTHOR INDEX OF VOLUME 18 OF “ECOLOGICAL CHEMISTRY AND ENGINEERING A”

WYKAZ AUTORÓW PUBLIKACJI ZAMIESZCZONYCH W TOMIE 18 MIESIÊCZNIKA „ECOLOGICAL CHEMISTRY AND ENGINEERING A / CHEMIA I IN¯YNIERIA EKOLOGICZNA A”

Meaning of the digits in the index entries – (no. of issue) first page, no. of the article (in the volume contents). Sposób zapisu odnoœników hase³ – (nr zeszytu) pierwsza strona artyku³u, nr artyku³u (w spisie treœci rocznika).

ALBERSKI Jacek (4) 531, 58 BONDAR-NOWAKOWSKA El¿bieta (12) 1763, AMALIO-KOSEL Magdalena (7) 895, 100 198 ANDRIYANEC Svetlana Valerjevna (4) 563, 62 BORUSZKO Dariusz (9–10) 1193, 134 ARASIMOWICZ Monika (9–10) 1175, 132 BREÑKO Tomasz (9–10) 1219, 137 BUCZKOWSKA Halina (1) 73, 9 BABIÑSKI Leszek (2) 183, 20 BURKOWSKA Aleksandra (3) 469, 53; (5–6) 827, BACZEWSKA Aneta Helena (3) 347, 35 95 BADOWSKA-KOZAKIEWICZ Anna M. (3) 365, C 38 HMURA Damian (2) 225, 25 BANASZKIEWICZ Teresa (3) 335, 33 CHRZAN Anna (11) 1527, 170 BARAN Agnieszka (4) 545, 60; (7) 941, 105; CI¥G£O Aldona (5–6) 673, 73 (9–10) 1185, 133; (9–10) 1237, 139; (11) 1505, CIEÆKO Zdzis³aw (11) 1485, 165; (12) 1641, 183 167 CIEPA£ Ryszard (5–6) 709, 78; (5–6) 785, 89 BARKOWSKA Mi³os³awa (8) 1065, 119 CIEPIELA Gra¿yna Anna (11) 1425, 159 CWALINA Beata (3) 391, 42 BEKIER Jakub (4) 497, 54 CZEKA£A Jacek (4) 507, 55; (9–10) 1227, 138; BIARDZKA El¿bieta (11) 1457, 162 (9–10) 1293, 146 BIBER Maria (9–10) 1211, 136 CZY¯OWICZ Iwona (11) 1545, 173 BICZAK Robert (8) 1025, 113 CZY¯YK Ewa (5–6) 763, 86 BIEGAÑSKA-MARECIK Ró¿a (1) 129, 16 CZY¯YK Franciszek (4) 515, 56 BIENIASZ Monika (1) 67, 8 BIESIADA Anita (1) 9, 1 D¥BEK Ma³gorzata (4) 637, 71 BJELKOVÁ Marie (2) 265, 29; (9–10) 1347, 152 D¥BKOWSKA-NASKRÊT Halina (9–10) 1245, B£ASZAK Magdalena (7) 959, 107 140; (11) 1417, 158 BOHDZIEWICZ Jolanta (7) 895, 100; (12) 1751, D¥BROWSKI Wojciech (2) 175, 19; (9–10) 1203, 197 135 BOLIG£OWA El¿bieta (5–6) 665, 72 DIATTA Jean B. (9–10) 1211, 136 1788 Indexes

D£UGOSZ Jacek (4) 629, 70 HELIOS Waldemar (12) 1599, 179 D£U¯NIEWSKA Joanna (3) 341, 34; (5–6) 679, HERMAN Barbara (8) 1025, 113 74 HERMANN Janusz (4) 537, 59 DMUCHOWSKI Wojciech (3) 347, 35; (8) 1059, HLUŠEK Jaroslav (9–10) 1347, 152 118 HOFFMANN Józef (7) 983, 110 DOMINIAK Alicja (1) 51, 6 HOFFMANN Krystyna (7) 983, 110; (11) 1407, DONDAJEWSKA Renata (2) 219, 24 157 DONIGIEWICZ Filip (7) 983, 110 HOLAK Magdalena (3) 419, 47 DROZD Jerzy (4) 497, 54 HUCULAK-M¥CZKA Marta (11) 1407, 157 DUDA Marcin (5–6) 723, 80 HUZAR El¿bieta (7) 991, 111 DUDZIAK Mariusz (7) 903, 101 DZIEDZIC Ewa (1) 67, 8 IGNATOWICZ Katarzyna (2) 191, 21; (9–10) DZIÊCIO£ Ma³gorzata (7) 991, 111 1219, 137 ILJINSKIJ Andrej Valerjevich (4) 563, 62 FEDORSKI Jerzy (8) 1053, 117 FIALA Karel (9–10) 1347, 152 JACAK Piotr (2) 183, 20 FILIPEK Tadeusz (4) 523, 57 JAGUŒ Andrzej (11) 1433, 160 FILIPEK-MAZUR Barbara (9–10) 1355, 153; (11) JAKUBUS Monika (9–10) 1227, 138 1533, 171 JAMROZ El¿bieta (4) 553, 61 FORMICKI Grzegorz (5–6) 673, 73 JANAS Piotr (11) 1477, 164 FR¥CZEK Krzysztof (3) 425, 48; (5–6) 685, 75; JANKOWSKA Jolanta (11) 1425, 159 (5–6) 813, 93 JANKOWSKI Kazimierz (11) 1425, 159 JARMU£-PIETRASZCZYK Joanna (3) 359, 37; GAJEWSKA Julitta (2) 183, 20 (3) 365, 38; (5–6) 703, 77; (11) 1571, 177; (12) GA£CZYÑSKA Ma³gorzata (12) 1715, 192 1711, 191 GARCZYÑSKA Mariola (9–10) 1263, 142; (11) JAROMIN Katarzyna (7) 853, 96; (11) 1467, 163 1557, 175 JASIEWICZ Czes³awa (4) 545, 60; (7) 941, 105; GAWÊDA Maria (1)19, 2; (1) 113, 14 (9–10) 1185, 133; (9–10) 1237, 139; (11) 1505, G¥SIOREK Micha³ (4) 577, 64; (5–6) 697, 76 167 GIBCZYÑSKA Marzena (12) 1715, 192 JASIÑSKA Agnieszka (1) 89, 11 GLEÑ Katarzyna (3) 353, 36; (11) 1515, 168; (12) JAWORSKA Hanna (9–10) 1245, 140; (11) 1417, 1721, 193 158 GLIÑSKA-LEWCZUK Katarzyna (2) 241, 27 JEKATIERYNCZUK-RUDCZYK El¿bieta (2) G£¥BOWSKA Daria (12) 1705, 190 201, 22 GO£DYN Ryszard (2) 219, 24 JLILATI Alia (7) 853, 96; (11) 1467, 163 GONDEK Krzysztof (8) 1079, 121; (9–10) 1251, JÓZEFOWSKA Agnieszka (3) 385, 41; (4) 577, 64 141 JURKIEWICZ-KARNKOWSKA Ewa (11) 1457, GONKIEWICZ Aleksander (1) 27, 3 162 GORCZYCA Anna (11) 1477, 164; (12) 1625, 181 GOSPODAREK Janina (11) 1515, 168; (11) 1565, KACORZYK Piotr (8) 1033, 114; (8) 1041, 115; 176; (12) 1651, 184 (8) 1117, 127; (11) 1539, 172 GÓRA Joanna (12) 1599, 179 KAMIENIECKA Agnieszka (11) 1401, 156 GRABOWSKI Kazimierz (4) 531, 58 KAMIÑSKA Iwona (1) 59, 7 GRATA Katarzyna (12) 1727, 194 KAMIONEK Marta (3) 359, 37; (3) 365, 38; (8) GRIGA Miroslav (2) 265, 29 1101, 124; (8) 1139, 130; (11) 1571, 177; (11) GRYGIERZEC Beata (8) 1019, 112; (12) 1675, 1575, 178; (12) 1711, 191 187 KANDZIORA-CIUPA Marta (5–6) 709, 78; (5–6) GRZEGORCZYK Stefan (4) 531, 58 785, 89 GUSEVA Ekaterina Sergeevna (4) 563, 62 KANIA Ines (11) 1571, 177 GUSEVA Tatjana Mihajlovna (4) 563, 62 KASPERCZYK Miros³aw (8) 1041, 115 KASPROWICZ J. Marek (11) 1477, 164; (12) HACHO£ Justyna (12) 1763, 198 1625, 181 HARASIM Pawe³ (4) 523, 57 KIRYLUK Aleksander (2) 207, 23 HARASIMOWICZ-HERMANN Gra¿yna (4) 537, KLAMKOWSKI Krzysztof (1) 35, 4 59 KLEIBER Tomasz (1) 43, 5 HAWROT-PAW Ma³gorzata (11) 1401, 156 KLIMASZYK Piotr (9–10) 1293, 146 Indexes 1789

K£YŒ Ma³gorzata (5–6) 717, 79 MALARZ W³adys³aw (12) 1599, 179 KOC Józef (3) 435, 49; (5–6) 723, 80 MALCZYK Piotr (8) 1087, 122; (9–10) 1287, 145 KOCHANEK Agnieszka (8) 1105, 125 MALINOWSKA Ewelina (12) 1711, 191 KOCOÑ Anna (5–6) 731, 81 MALINOWSKA Katarzyna (3) 377, 40; (5–6) 763, KOCOWICZ Andrzej (4) 553, 61 86; (8) 1093, 123; (8) 1105, 125 KOLCZAREK Roman (11) 1425, 159 MALINOWSKI Tadeusz (4) 637, 71 KO£ODZIEJ Jan (8) 1071, 120 MA£ODOBRY Monika (1) 67, 8 KO£ODZIEJCZYK Marek (8) 1071, 120 MARKO-WOR£OWSKA Maria (11) 1527, 170 KOMOSA Andrzej (1) 43, 5 MARSKA Barbara (8) 1093, 123 KONIECZNY Krystyna (12) 1743, 196 MATUSKA Joanna (8) 1101, 124 KOPEÆ Micha³ (9–10) 1251, 141 MATUSKA-£Y¯WA Joanna (11) 1575, 178 KORC Marian (4) 601, 67 MAZHAJSKIJ Urij Anatoljevich (4) 563, 62 KORZEKWA-WOJTAL Anna (7) 887, 99 MAZUREK Ryszard (3) 341, 34; (3) 385, 41; KORZENIOWSKA Jolanta (3) 445, 50 (5–6) 679, 74; (5–6) 771, 87 KOSTECKA Joanna (9–10) 1263, 142; (11) 1557, MAZURKIEWICZ Anna (8) 1139, 130 175; (12) 1683, 188 MERCIK Stanis³aw (4) 611, 68 KOTECKI Andrzej (12) 1599, 179 MICHALCEWICZ Wiera (12) 1715, 192 KOVÁÈIK Peter (9–10) 1237, 139 MICHA£OJÆ Zenia (1) 73, 9; (4) 571, 63 KOWALCZEWSKA-MADURA Katarzyna (2) MICHA£OWSKA Sylwia (7) 959, 107 219, 24 MIECHÓWKA Anna (4) 577, 64 KOWALCZYK-JUŒKO Alina (7) 973, 109 MIELCZAREK Karolina (12) 1751, 197 KOWALIK Stanis³aw (9–10) 1271, 143 MIELNIK Lilla (9–10) 1293, 146 KOWALSKA-GÓRALSKA Monika (3) 371, 39; MIKICIUK Grzegorz (5–6) 777, 88 (5–6) 737, 82; (5–6) 743, 83; (8) 1047, 116; (8) MIKICIUK Ma³gorzata (5–6) 763, 86; (5–6) 777, 1053, 117; (11) 1545, 173 88; (8) 1105, 125 KOZAK Marcin (12) 1599, 179 MOLENDA Tadeusz (2) 225, 25 KOZAK Przemys³aw (1) 51, 6 MUSZYÑSKI Pawe³ (9–10) 1301, 147 KOZIK El¿bieta (1) 51, 6 KUBICKA Helena (8) 1059, 118 NABRDALIK Ma³gorzata (12) 1727, 194 KUCHARSKA Alicja (1) 9, 1 NADGÓRSKA-SOCHA Aleksandra (5–6) 709, KUCHARSKA Kornelia (5–6) 749, 84; (8) 1065, 78; (5–6) 785, 89 119 NAJMOWICZ Tomasz (11) 1485, 165 KUCZKOWSKA-KUNIAR Anna (5–6) 673, 73 NAMIOTKO Anna (8) 1123, 128; (9–10) 1327, KUD Krzysztof (9–10) 1279, 144 150 KULCZAK Katarzyna (1) 59, 7 NAWIRSKA Agnieszka (1) 9, 1 KULIG Bogdan (8) 1071, 120 NIEMIEC Marcin (2) 235, 26; (9–10) 1175, 132 KUSIÑSKA Alina (4) 585, 65 NIZIO£-£UKASZEWSKA Zofia (1)19, 2 KUNIAR Tomasz (5–6) 757, 85 NYCH Alicja (7) 933, 104 KWARCIAK-KOZ£OWSKA Anna (12) 1751, 197 O KWIECIÑSKA Anna (12) 1743, 196 BOLEWSKI Krystian (2) 241, 27 OKTABA Lidia (4) 585, 65 LEJA Ma³gorzata (1) 27, 3 OLEKSY Andrzej (8) 1071, 120 LEJA Maria (1) 59, 7 ONUCH-AMBORSKA Joanna (4) 593, 66 LEMEK Tadeusz (5–6) 757, 85 ORLIKOWSKA Teresa (11) 1551, 174 LEŒNIAK Magdalena (3) 397, 43 ORLIKOWSKI B. Leszek (11) 1551, 174 LICZNAR Micha³ (4) 497, 54 OSADOWSKI Zbigniew (2) 241, 27 LOPATOVSKAYA Olga (2) 241, 27 P LORENC-KOZIK Anna (8) 1079, 121 ACHOLEWSKA Ma³gorzata (3) 391, 42 PALOWSKI Bernard (3) 453, 51 £ABUZ Bernadetta (3) 461, 52 PATORCZYK-PYTLIK Barbara (9–10) 1313, 148 £AGÓD Grzegorz (7) 853, 96; (7) 865, 97; (11) PAWLUCZUK Jan (2) 255, 28 1467, 163 P¥CZKA Grzegorz (12) 1683, 188 £AWA Piotr (3) 371, 39 PERSICOVA Tamara (4) 619, 69 £UKOWIAK Remigiusz (9–10) 1211, 136 PEZOWICZ El¿bieta (3) 365, 38; (3) 397, 43; (3) 407, 45; (5–6) 749, 84; (5–6) 801, 91; (8) 1065, MADEYSKI Marek (11) 1505, 167 119; (8) 1139, 130 1790 Indexes

PIASECKA Joanna (5–6) 703, 77 ŠKARPA Petr (2) 265, 29; (7) 951, 106; (9–10) PIEKUTIN Janina (9–10) 1321, 149 1347, 152 PI£AT Krzysztof (4) 629, 70 SKONIECZEK Pawe³ (3) 435, 49 PIOTROWICZ Ryszard (9–10) 1293, 146 SKORBI£OWICZ El¿bieta (12) 1633, 182 PIOTROWSKA Anna (3) 385, 41 SKORBI£OWICZ Miros³aw (9–10) 1339, 151; PIOTROWSKA Gra¿yna (4) 637, 71 (12) 1633, 182 PISULEWSKA El¿bieta (5–6) 793, 90; (8) 1079, SKRÊTOWICZ Maria (7) 933, 104 121 SKWARKA Tomasz (5–6) 743, 83; (8) 1053, 117 PLACEK Monika (1)147, 18 SKWARY£O-BEDNARZ Barbara (5–6) 821, 94; POBO¯NIAK Maria (1) 83, 10 (8) 1131, 129 PODLEŒNA Anna (3) 401, 44 SMOCZYÑSKI Lech (7) 911, 102 POKORSKA Kamila (3) 419, 47 SMOLEÑ Sylwester (1) 97, 12; (1) 105, 13 POLESZCZUK Olga (3) 407, 45; (5–6) 801, 91 SMOLIK Beata (11) 1401, 156 PORADOWSKI Ryszard (5–6) 793, 90 SOBCZUK Henryk (7) 865, 97; (7) 877, 98 POSHTOVAYA Natalia (4) 619, 69 SOBCZYÑSKA-WÓJCIK Katarzyna (11) 1441, POSPÍŠILOVÁ Lubica (9–10) 1347, 152 161 PROTASOWICKI Miko³aj (5–6) 805, 92 SOBIERALSKI Krzysztof (1) 89, 11 PRZYBULEWSKA Krystyna (7) 959, 107; (12) SOKÓ£-£ÊTOWSKA Anna (1) 9, 1 1705, 190 SOSULSKI Tomasz (4) 601, 67; (4) 611, 68 SÓWKA Izabela (7) 933, 104 PRZYGOCKA-CYNA Katarzyna (9–10) 1211, 136 SPYCHAJ-FABISIAK Ewa (4) 629, 70 PTASZEK Magdalena (11) 1551, 174 STANIS£AWSKA-GLUBIAK Ewa (3) 445, 50 PYZA Agnieszka (8) 1059, 118 STANKOWSKI S³awomir (12) 1715, 192 R STAWARZ Robert (5–6) 673, 73 ADKOWSKA Iwona (3) 413, 46; (8) 1111, 126; STEINDOR Karolina (3) 453, 51 (11) 1521, 169 STEPANIUK Sylwia (8) 1093, 123 RADKOWSKI Adam (3) 413, 46; (8) 1111, 126; STOLARSKA Anna (7) 959, 107; (12) 1705, 190 (11) 1521, 169; (12) 1737, 195 SUCHORAB Zbigniew (7) 865, 97; (7) 877, 98 RADZIEJEWSKA-KUBZDELA El¿bieta (1) 129, SZA£ATA Rafa³ (5–6) 737, 82 16 SZCZEPANIAK Stanis³awa (1) 51, 6 RADZIEMSKA Maja (11) 1497, 166 SZCZYKOWSKA Joanna (9–10) 1363, 154 RAFA£OWSKA Ma³gorzata (11) 1441, 161 SZEJNIUK Bo¿ena (12) 1691, 189 RAJKOWSKA Monika (3) 419, 47; (5–6) 805, 92 SZEWCZYK Wojciech (8) 1041, 115 RAJMUND Agnieszka (4) 515, 56 SZOPIÑSKA Anna (1) 113, 14 RAUCKYTE-¯AK Teresa (12) 1667, 186; (12) 1691, 189 ŒCIGALSKA Barbara (3) 461, 52 ROCHWERGER Andrzej (12) 1633, 182 ŒWIDERSKI Adam (1) 83, 10 ROLKA El¿bieta (12) 1641, 183 ŒWIERCZYÑSKA Anna (7) 895, 100 ROPEK Dariusz (3) 419, 47; (5–6) 685, 75; (5–6) 757, 85; (5–6) 813, 93; (8) 1117, 127 ŒWIÊCI£O Agata (7) 967, 108 RYCHTER Piotr (8) 1025, 113 T RYDLEWSKA Magdalena (8) 1087, 122; (9–10) ABAK Monika (9–10) 1355, 153; (11) 1533, 171 1287, 145 TARGOWSKI Adam (2) 301, 32 TARNAWSKI Marek (7) 941, 105; (11) 1505, 167 SACA£A El¿bieta (12) 1615, 180 TREDER Waldemar (1) 35, 4 SADY W³odzimierz (1) 97, 12; (1) 105, 13 TRYNGIEL-GAÆ Anna (1) 35, 4 SAJDAK Aleksandra (8) 1071, 120 TRZEWIK Aleksandra (11) 1551, 174 SAS-GOLAK Iwona (1) 89, 11 TSAKOVSKI Stefan (2) 275, 30 S¥DEJ Wiera (8) 1123, 128; (9–10) 1327, 150 TUMIALIS Dorota (3) 397, 43; (3) 407, 45; (5–6) SENZE Magdalena (3) 371, 39; (5–6) 737, 82; (8) 749, 84; (5–6) 801, 91; (8) 1065, 119; (8) 1139, 1047, 116; (11) 1545, 173 130 SIDORUK Marcin (12) 1633, 182 SIEMIENIUK Anna (9–10) 1363, 154 VAN GRIEKEN Rene (7) 933, 104 SIMEONOV Vasil (2) 275, 30 VETROVCOVÁ Martina (2) 265, 29 SIWEK Piotr (1) 139, 17 SIWULSKI Marek (1) 89, 11 WADAS Wanda (1) 123, 15 Indexes 1791

WALCZAK Maciej (3) 469, 53; (5–6) 827, 95 WOROBIEC Anna (7) 933, 104 WALKOWIAK-TOMCZAK Dorota (1) 129, 16 WÓJCIK Jerzy (9–10) 1271, 143 WARDZYÑSKA Regina (7) 911, 102 WRAGA Krzysztof (1)147, 18 WIATER Józefa (9–10) 1363, 154 WRÓBEL Jacek (5–6) 763, 86 WIATKOWSKI Miros³aw (2) 289, 31 WYSZKOWSKI Miros³aw (9–10) 1373, 155; (11) WIDOMSKI Marcin (7) 853, 96; (7) 865, 97; (11) 1485, 165; (11) 1497, 166; (12) 1641, 183 1467, 163 WILKOWSKI Przemys³aw (3) 359, 37 ZADRO¯NY Pawe³ (4) 577, 64; (5–6) 771, 87 WIŒNIOWSKA-KIELIAN Barbara (2) 235, 26; ZA£ÊSKA-CHRÓST Beata (7) 911, 102 (9–10) 1175, 132 ZAMACHOWSKI W³adys³aw (5–6) 673, 73 WITKOWICZ Robert (5–6) 793, 90 ZAWADZKI Jaros³aw (2) 301, 32 W£ODARCZYK Ma³gorzata (8) 1147, 131 ZAWIEJA Iwona (7) 923, 103; (12) 1659, 185 WODNICKA Alicja (7) 991, 111 ZIMOCH Aldona (9–10) 1313, 148 WOJCIECHOWSKA Renata (1) 139, 17 ZIÓ£KOWSKA Agnieszka (9–10) 1373, 155 WOJTAS Alojzy (4) 637, 71 ZWODZIAK Anna (7) 933, 104 WOLNY Lidia (7) 887, 99; (7) 923, 103; (12) ZWODZIAK Jerzy (7) 933, 104 1659, 185 WOLSKA-SOBCZAK Aneta (8) 1059, 118 ¯AK S³awomir (12) 1667, 186 WOLSKI Pawe³ (7) 923, 103; (12) 1659, 185 ¯ELAZNA Agnieszka (7) 877, 98

SUBJECT INDEX

Meaning of the digits in the index entries – (no. of issue) first page, no. of the article (in the volume contents). abiotic factors (5–6) 665, 72 ash (7) 973, 109 Acer platanoides L. (3) 377, 40 assessment methods (9–10) 1211, 136 Acidithiobacillus ferrooxidans (3) 391, 42 assimilation dyes (5–6) 763, 86; (8) 1093, 123 acidity (11) 1497, 166 assimilation pigments (3) 377, 40; (5–6) 777, 88; actinomycetes (12) 1715, 192 (8) 1105, 125 activated sludge (7) 895, 100 atmospheric precipitation (4) 515, 56 advection-dispersion equation (7) 865, 97 Auricularia auricula-judae (1) 89, 11 aerosol fraction (2) 275, 30 Auricularia polytricha (1) 89, 11 Aesculus hippocastanum (3) 453, 51 auxin (1) 27, 3 agricultural catchment area (5–6) 723, 80 Bacillus cereus (12) 1727, 194 AgroHydrogel (5–6) 777, 88 Bacillus mycoides (12) 1727, 194 agro-utilization of wastewaters from vegetable fat bacteria (2) 183, 20; (5–6) 685, 75; (12) 1715, 192 production (12) 1667, 186 bacterial preparations (4) 619, 69 alfalfa (9–10) 1313, 148 bacterioneuston (5–6) 827, 95 alginate (8) 1147, 131 Baycidal WP 25 (5–6) 749, 84 alkalisation (9–10) 1245, 140 beneficial insects (3) 425, 48 allelopathy (12) 1615, 180 bentonite (9–10) 1373, 155; (12) 1641, 183 alluvial soils (9–10) 1279, 144 Betula pendula Roth. (5–6) 785, 89 Alphitobius diaperinus (5–6) 749, 84 Bieszczady National Park (5–6) 709, 78 aluminium (11) 1545, 173 bioaccumulation (11) 1545, 173 aluminium content (12) 1641, 183 biodegradation (11) 1401, 156 amino acids (4) 523, 57; (4) 545, 60 biodegradation of pollutants in sewer system (7) ammonia (4) 585, 65; (4) 637, 71 865, 97 ammonium ions (1) 139, 17 biodiesel (11) 1401, 156 amphibian (5–6) 673, 73 biogas (7) 923, 103 antagonism of elements (3) 413, 46 biogenic (9–10) 1203, 135 anthocyanins (1) 67, 8 biogenic compounds (9–10) 1363, 154 anthropogenic pollution (3) 397, 43 biological SBR (7) 895, 100 anthropogenic water bodies (2) 225, 25 biomass (3) 353, 36; (7) 973, 109; (12) 1721, 193 anthurium (1) 43, 5 Biskupin (2) 183, 20 antioxidant activity (1) 59, 7 black and green teas (9–10) 1185, 133 antioxidative status (7) 967, 108 black locust (3) 385, 41; (5–6) 679, 74 aquatic plants (11) 1545, 173 boron (1) 19, 2 aquatic vascular plants (12) 1763, 198 Botrytis fabae (11) 1515, 168 arable layer (3) 461, 52 bottom deposits (12) 1633, 182 arable soils (3) 385, 41 bottom sediments (2) 219, 24; (5–6) 805, 92; (7) arsenic contamination (11) 1485, 165 941, 105; (9–10) 1175, 132; (9–10) 1293, 146; Ascochyta fabae (11) 1515, 168 (11) 1457, 162; (11) 1505, 167 ascorbic acid (8) 1025, 113 brew (9–10) 1185, 133 1794 Indexes brine shrimp (3) 371, 39 content (9–10) 1175, 132 broiler chickens (3) 335, 33 content of macroelements (3) 413, 46 brown coal (12) 1641, 183 content of nitrates(V) (8) 1123, 128 Bruchus rufimanus Boh. (11) 1515, 168; (12) 1651, content of organic compounds (11) 1521, 169 184 conventional cultivation (1) 113, 14 building barriers (7) 877, 98 copper (3) 445, 50; (5–6) 673, 73; (9–10) 1211, building envelopes (7) 877, 98 136; (9–10) 1327, 150; (11) 1533, 171 building materials (7) 877, 98 Corsican hellebore (1) 51, 6 cultivars (1) 9, 1; (8) 1019, 112; (12) 1675, 187 C/N ratio, (4) 553, 61; (11) 1457, 162 cultivation (1) 147, 18 C4 photosynthesis (12) 1615, 180 cadmium (2) 265, 29; (3) 365, 38; (5–6) 673, 73; dairy industries (9–10) 1193, 134 (5–6) 763, 86; (5–6) 771, 87; (8) 1059, 118; dairy wastewater (2) 175, 19 (9–10) 1211, 136; (11) 1425, 159 Dar 2.5GR (9–10) 1263, 142 calcium (8) 1059, 118; (9–10) 1175, 132 dehydrogenase activity (3) 385, 41 calcium oxide (9–10) 1373, 155; (11) 1497, 166 density (11) 1527, 170 calibration of sewer system hydraulic model (7) detection (11) 1551, 174 853, 96 dielectric methods (7) 877, 98 carbon nanotubes (5–6) 757, 85 diesel oil (9–10) 1373, 155 carp (8) 1047, 116 3,4-dimethylpyrazol phosphate (DMPP) (1) 97, 12 carrot (1) 19, 2; (1) 113, 14 Dimilin 25WP (9–10) 1263, 142; (11) 1557, 175 catchment area (12) 1633, 182 Diptera (11) 1557, 175 cationic surfactant (9–10) 1301, 147 distillery spent wash (4) 537, 59 celery (8) 1025, 113 diversity (11) 1527, 170 cement dust (9–10) 1245, 140 DNA synthesis (3) 469, 53 centrifuging (7) 887, 99 dug wells (2) 207, 23 cereals (4) 507, 55; (5–6) 665, 72 dusts (9–10) 1287, 145 chemical composition (1) 9, 1; (1) 89, 11; (5–6) 793, 90; (7) 973, 109; (12) 1737, 195 E. fetida (11) 1557, 175 chemical properties (9–10) 1271, 143 earthworm Dendrobena veneta (9–10) 1263, 142 chemometrics (2) 275, 30 earthworm ecology box (11) 1557, 175 chilling tolerance (12) 1615, 180 ecological box (9–10) 1263, 142 chlorine (5–6) 723, 80 ecological risk (2) 301, 32; (12) 1763, 198 chlorophyll (1) 35, 4; (1) 83, 10; (8) 1025, 113 eggplant (1) 73, 9 chromium (11) 1533, 171 Eisenia fetida (5–6) 703, 77 chromium(III) (11) 1497, 166 electrocoagulation (7) 911, 102 chromium(VI) (11) 1497, 166 elemental composition (7) 933, 104 classification (2) 275, 30 emigration activity (5–6) 717, 79 climatic condition (4) 553, 61 energy crops (7) 973, 109 CO2 assimilation (5–6) 763, 86 entomopathogenic fungi (3) 359, 37; (11) 1571, coke-making wastewater treatment (12) 1751, 197 177; (12) 1711, 191 Col. Carabidae (5–6) 813, 93 entomopathogenic nematodes (3) 359, 37; (3) 397, Col. Staphylinidae (5–6) 813, 93 43; (3) 407, 45; (5–6) 749, 84; (5–6) 757, 85; coli bacteria (12) 1715, 192 (5–6) 801, 91; (8) 1065, 119; (8) 1101, 124; commodity evaluation (7) 991, 111 (11) 1575, 178; (12) 1711, 191 common dandelion (11) 1425, 159 environmental pollution (3) 347, 35 compost (1) 147, 18; (3) 347, 35; (4) 545, 60; enzymatic activity (5–6) 827, 95 (9–10) 1193, 134; (9–10) 1219, 137; (9–10) enzymatic preparation (3) 335, 33 1227, 138; (9–10) 1355, 153; (9–10) 1373, 155; EPN (entomopathogenic nematodes) (5–6) 749, 84 (11) 1497, 166; (12) 1641, 183 ethephon (1) 27, 3 composting of municipal solid wastes (4) 497, 54 eutrophication (11) 1433, 160 concentrated phosphoric acid (7) 983, 110 exchangeable complex (9–10) 1287, 145 conditioning (12) 1659, 185 constructed wetlands (2) 175, 19; (9–10) 1203, 135 F. culmorum (11) 1477, 164; (12) 1625, 181 contamination (5–6) 743, 83; (7) 959, 107; (9–10) faba bean (8) 1071, 120 1373, 155; (11) 1497, 166 Fagus sylvatica L. (5–6) 709, 78 Indexes 1795 farmstead (2) 207, 23 heavy metals (1) 97, 12; (1) 105, 13; (2) 191, 21; farmyard manure (4) 545, 60 (2) 235, 26; (2) 301, 32; (3) 347, 35; (3) 419, fenamidon (11) 1551, 174 47; (3) 453, 51; (3) 461, 52; (4) 563, 62; (5–6) fertilization (1) 123, 15; (3) 359, 37; (4) 507, 55; 697, 76; (5–6) 709, 78; (5–6) 785, 89; (7) 941, (4) 601, 67; (4) 611, 68; (8) 1019, 112; (8) 105; (8) 1101, 124; (9–10) 1219, 137; (9–10) 1033, 114; (8) 1041, 115; (8) 1117, 127; (8) 1237, 139; (9–10) 1279, 144; (9–10) 1293, 146; 1123, 128; (11) 1407, 157; (11) 1539, 172; (12) (11) 1515, 168; (11) 1527, 170; (11) 1565, 176; 1675, 187 (12) 1651, 184; (12) 1667, 186 fertilization with microelements (12) 1737, 195 herbicides (7) 959, 107 films with different oxygen permeability (1) 129, herbs (4) 531, 58 16 Heterorhabditis bacteriophora (5–6) 749, 84; (8) final hydration (7) 887, 99 1065, 119 flax (2) 265, 29 Heterorhabditis megidis (8) 1139, 130; (11) 1575, floodplain water bodies (11) 1457, 162 178 fluazifop-p-butyl (7) 967, 108 hilling (12) 1599, 179 fluidal ash (12) 1715, 192 hop wastes (11) 1407, 157 fodder additive (7) 983, 110 horses (8) 1053, 117 fodder phosphate (7) 983, 110 humic acids (9–10) 1347, 152 foliar feeding (4) 523, 57 humic substances (3) 469, 53; (9–10) 1347, 152 foliar fertilization (1) 35, 4; (3) 353, 36; (12) 1721, humus acids (11) 1407, 157 193 husked oats (5–6) 793, 90 forest soils (5–6) 821, 94 hydrogel microcapsules (8) 1147, 131 forms of Pb (8) 1131, 129 hypertrophy (11) 1441, 161 fractal dimension (7) 911, 102 Fraxinus excelsior (3) 453, 51 IAA (indoleacetic acid) (7) 967, 108 fresh alluvium (9–10) 1279, 144 incineration wastes (9–10) 1271, 143 fruiting bodies (1) 89, 11 indoor air quality (7) 991, 111 fulvic acids (9–10) 1347, 152 industrial areas (8) 1131, 129 fungi (7) 959, 107; (12) 1705, 190; (12) 1715, 192 inhibition (11) 1551, 174 Fusarium (3) 353, 36; (12) 1721, 193 inland water (11) 1433, 160 fusibility (7) 973, 109 insecticide (5–6) 749, 84 integrated cultivation (1) 113, 14 Galleria mellonella (3) 365, 38; (8) 1101, 124; integrated system (12) 1751, 197 (11) 1575, 178 interstitial water (2) 201, 22 garden sorrel (11) 1425, 159 ionizing radiation (3) 407, 45; (5–6) 801, 91 gas exchange (1) 35, 4 iron(II) (3) 391, 42 geochemical background (9–10) 1211, 136 geostatistics (2) 301, 32 kitchen organic waste (11) 1557, 175 globose scale (3) 397, 43 Krakow (5–6) 697, 76 grain (5–6) 665, 72 Kwacza River (2) 241, 27 grain yield (5–6) 793, 90 granulometry (11) 1457, 162 Lactuca sativa var. capitata (8) 1105, 125 grapes (1) 59, 7 lakes (5–6) 805, 92; (12) 1633, 182 grasslands (4) 531, 58 land use (4) 585, 65 gravitational sewer system (11) 1467, 163 landfill leachates (7) 895, 100 gravity thickening (7) 887, 99 landscape (4) 563, 62 greater plantain (11) 1425, 159 lead (1) 19, 2; (2) 265, 29; (3) 365, 38; (9–10) green waste compost (9–10) 1327, 150 1211, 136; (9–10) 1227, 138; (11) 1417, 158; groundwater (2) 255, 28; (4) 637, 71 (12) 1633, 182 lead ions (8) 1101, 124; (11) 1575, 178 habitat conditions (2) 255, 28 leaf area (5–6) 777, 88 hair (8) 1053, 117 leaf colour (1) 83, 10 halophytes (2) 225, 25 leaves (9–10) 1185, 133 harvest date (12) 1599, 179 legumes (4) 531, 58 hay (3) 413, 46 Lentinula edodes (1) 89, 11 health risk (2) 207, 23 lettuce (1) 19, 2; (4) 571, 63 1796 Indexes light soil (11) 1505, 167 micronutrients (4) 523, 57 light soils (8) 1131, 129 microorganisms (3) 341, 34; (4) 563, 62; (7) 895, lignite (11) 1407, 157 100; (11) 1401, 156 lime (8) 1041, 115; (9–10) 1287, 145; (12) 1641, microscopic fungi (2) 183, 20 183 military training ground (5–6) 743, 83 limnic processes (2) 225, 25 mine (5–6) 737, 82 linear growth (3) 353, 36; (12) 1625, 181; (12) mineral components (9–10) 1339, 151 1721, 193 mineral composition (5–6) 673, 73 linseed (2) 265, 29 mineral nitrogen (4) 585, 65 Linum usitatissimum L. (2) 265, 29 mineralization (2) 255, 28 Lipases (12) 1727, 194 Miscanthus (12) 1599, 179; (12) 1615, 180 liquid in vitro cultures (8) 1139, 130 mixed crops (4) 619, 69 liquid manure (12) 1743, 196 modified atmosphere (1) 129, 16 lithium (9–10) 1279, 144 moisture (7) 877, 98 load provided (11) 1539, 172 monocalcium phosphate (7) 983, 110 load reduction (3) 435, 49 motorization (9–10) 1321, 149 lobelia lakes (9–10) 1293, 146 mountain meadow (8) 1041, 115 Lolium perenne (12) 1675, 187 mountains soils (4) 553, 61 long-term experiment (4) 601, 67; (4) 611, 68; (7) multinutrient complex fertilizers (1) 123, 15 951, 106 municipal landfill sites (3) 425, 48; (5–6) 685, 75; long-term fertilizer experiment (9–10) 1251, 141 (5–6) 813, 93 low cost methods (9–10) 1193, 134 municipal solid waste compost (9–10) 1327, 150 lowland river (2) 201, 22 mycoestrogens (7) 903, 101 low-retention reservoirs (9–10) 1363, 154 mycotoxins (5–6) 665, 72 macroelements (1) 43, 5; (3) 419, 47; (8) 1033, NAA (neutron activation analysis) (1) 27, 3 114; (8) 1079, 121; (9–10) 1175, 132; (11) nail polish removers (7) 991, 111 1505, 167; (11) 1539, 172; (12) 1683, 188 naked oats (5–6) 793, 90 macrofauna (8) 1117, 127 nanofiltration (7) 903, 101; (12) 1743, 196 magnesium (5–6) 673, 73; (8) 1059, 118; (9–10) Nanogro (12) 1625, 181 1175, 132; (11) 1485, 165 nanosilver (8) 1065, 119 maintenance works (12) 1763, 198 b-naphthoxyacetic acid (7) 967, 108 maize (4) 545, 60; (9–10) 1355, 153 narrow-leaved lupine (4) 619, 69 manganese (8) 1059, 118; (9–10) 1279, 144; Nematop (5–6) 749, 84; (8) 1065, 119 (9–10) 1327, 150 neutralizing substances (11) 1485, 165 manning formula (7) 853, 96 nickel (3) 445, 50 meadow (9–10) 1251, 141 N-NO3 (1) 123, 15 meadow sward (8) 1111, 126; (11) 1521, 169; (12) nitrates(III) (1) 129, 16; (4) 571, 63 1737, 195 nitrates(V) (1) 129, 16; (2) 207, 23; (4) 571, 63 melioration systems (9–10) 1339, 151 nitrates (1) 139, 17; (4) 515, 56; (4) 585, 65; (4) membrane processes (12) 1743, 196 637, 71; (9–10) 1339, 151 membranes pressure techniques (12) 1751, 197 nitrate reductase (1) 139, 17 mercury (8) 1087, 122; (9–10) 1245, 140; (9–10) nitrification inhibitor (1) 97, 12 1251, 141 nitrogen (3) 401, 44; (4) 523, 57; (4) 531, 58; (4) mesofauna (8) 1117, 127 553, 61; (4) 563, 62; (4) 577, 64; (4) 637, 71; metalaxyl (11) 1551, 174 (9–10) 1175, 132 metals (5–6) 805, 92 nitrogen bioconversion (4) 537, 59 metals speciation forms (12) 1667, 186 nitrogen content reduction (4) 537, 59 metazachlor (8) 1147, 131 nitrogen fertilization (1) 51, 6; (4) 593, 66; (4) 619, methane (7) 923, 103 69; (12) 1599, 179 microbial biomass (4) 577, 64 nitrogen forms (1) 73, 9; (4) 507, 55 microbial transformation modelling (7) 865, 97 nitrogen leaching (1) 97, 12; (1) 105, 13 microbiological research (12) 1691, 189 nitrogen transformations (4) 497, 54 microelement fertilization (8) 1111, 126 Nomolt 150SC (9–10) 1263, 142 microelements (1) 43, 5; (3) 419, 47; (8) 1079, 121 NPK (nitrogen, phosphorus, potassium) (4) 545, 60 microelements dose and form (4) 571, 63 nutrient ratio (1) 43, 5 Indexes 1797 nutrients (2) 175, 19 polysulfone ultrafiltration membranes (12) 1751, nutrients and organic substances (9–10) 1203, 135 197 nutritional value (11) 1521, 169 pond (3) 435, 49 population dynamics (5–6) 717, 79 oat (4) 619, 69 pot cultivation (1) 51, 6 Ojcow National Park (5–6) 771, 87 pot experiment (3) 401, 44 one-crop system (3) 461, 52 potassium (8) 1059, 118; (9–10) 1175, 132 onion (1) 19, 2 potassium deficit (5–6) 731, 81 optimisation of parameters (8) 1139, 130 potato (1) 123, 15; (1) 147, 18 organic cultivation (1) 113, 14 potato tubers (8) 1123, 128 organic kitchen waste (12) 1683, 188 Poznan (9–10) 1211, 136 organic materials (9–10) 1355, 153 pre-dam (2) 289, 31 organic matter (11) 1457, 162 pre-dam reservoir efficiency (2) 289, 31 organic substances (11) 1407, 157 primary production (11) 1441, 161 Owinema (8) 1065, 119 production field (4) 629, 70 oxygen-free stabilization (7) 923, 103 proline (12) 1705, 190 p protected areas (5–6) 821, 94 arent rock (4) 553, 61 protected zone (8) 1131, 129 pea varieties (5–6) 731, 81 protein (8) 1025, 113 peas (1) 83, 10 protein synthesis (3) 469, 53 peat (4) 571, 63 PTWI (permissible temporary weekly intake) pedofauna (11) 1527, 170 (9–10) 1185, 133 Pelcznica River (5–6) 737, 82 pulp and paper wastewater (7) 911, 102 Pentas lanceolata (1) 147, 18 pulsed magnetic field (11) 1477, 164 pesticide (2) 191, 21 pumpkin species (1) 9, 1 pesticide graveyard (2) 191, 21 pesticides (5–6) 703, 77 quality and nutritional value (8) 1111, 126 pests (3) 425, 48 petrol (9–10) 1373, 155 radish (1) 19, 2; (9–10) 1237, 139 petroleum-derivative substances (9–10) 1321, 149 rainfall (8) 1071, 120 pH (8) 1041, 115; (12) 1683, 188 rainwaters (2) 235, 26 phenolic compounds (1) 59, 7 raised bog (2) 255, 28 phosphorus (2) 219, 24; (9–10) 1175, 132 rape cakes (3) 335, 33 phosphorus farming (5–6) 731, 81 reclamation (9–10) 1271, 143 Photorhabdus luminescens (8) 1139, 130 recreational parks (9–10) 1211, 136 Phrynohyas resinifictrix larvae (5–6) 673, 73 recultivated land (4) 593, 66 physical and chemical parameters (2) 183, 20 red beet (1) 19, 2 physico-chemical sewage parameters (7) 865, 97 reflectometric moisture measurement methods (7) Phytophthora spp. (11) 1551, 174 877, 98 phytoremediation (3) 445, 50; (12) 1615, 180 reject water (9–10) 1203, 135 plant analysis (1) 43, 5 release (8) 1147, 131 plant coverage of the soil (8) 1033, 114; (11) 1539, removal micropollutants (7) 903, 101 172 renaturization (2) 241, 27 plant growth regulators (4) 619, 69 renewable energy sources (RES) (7) 923, 103 plant vigor (1) 35, 4 resistance to As(III) and Sb(III) (3) 391, 42 plant yield (4) 593, 66 respiratory activity (5–6) 827, 95 plants (11) 1485, 165; (12) 1641, 183 restoration (2) 219, 24 Pleurotus eryngii (1) 89, 11 retention reservoir (5–6) 723, 80; (2) 235, 26 PM1.0 (7) 933, 104 rhododendron baits (11) 1551, 174 PM2.5 (7) 933, 104 river macrophytes (2) 241, 27 PM10 indoor-outdoor particles (7) 933, 104 river-lake system (11) 1441, 161 Poa pratensis (8) 1019, 112 rivers (11) 1545, 173 pollution (5–6) 697, 76; (5–6) 771, 87; (5–6) 785, Robinia pseudoacacia (3) 341, 34 89 root quality (1) 113, 14 polyelectrolyte (7) 887, 99 roughness coefficient (7) 853, 96 polymictic lakes (11) 1441, 161 Roztocze National Park (8) 1131, 129 1798 Indexes runoff (2) 235, 26 soil carbon (4) 601, 67 rye (8) 1059, 118 soil management method (4) 577, 64 soil microorganisms (5–6) 821, 94 Saccharomyces cerevisiae (7) 967, 108 soil nitrogen (4) 601, 67; (4) 611, 68 Saint-Venant equation (7) 865, 97 soil organic matter (4) 601, 67 saline waters (2) 225, 25 soil pollution (3) 445, 50; (11) 1515, 168; (11) salinity (8) 1025, 113; (12) 1683, 188; (12) 1705, 1565, 176; (12) 1651, 184 190 soil properties (9–10) 1373, 155 Salix viminalis (5–6) 763, 86 soil reaction (9–10) 1347, 152 sand (4) 571, 63 soil type (9–10) 1313, 148 sanitary state (9–10) 1237, 139 soils (2) 301, 32; (9–10) 1245, 140 sanitary status indicator (12) 1691, 189 Sola cascade dam reservoirs (11) 1433, 160 sawdust (1) 89, 11 soluble forms (7) 941, 105 scale insects (3) 397, 43 sorption (9–10) 1301, 147 Scardinius erythrophthalmus (3) 419, 47 sorption isotherm (2) 191, 21 season of year (11) 1571, 177 sorption properties (11) 1497, 166 seasonal changes (1) 43, 5 sowing density (5–6) 793, 90 seasonal sampling (2) 275, 30 soybean (8) 1079, 121 Secale cereale L. (8) 1059, 118 spatial variability (4) 629, 70; (8) 1087, 122 secondary sampling designs (2) 301, 32 spinach (1) 19, 2 seed yield (8) 1019, 112; (12) 1675, 187 sporulation (3) 353, 36; (12) 1625, 181; (12) 1721, selenium (5–6) 743, 83; (8) 1047, 116; (8) 1053, 193 117; (9–10) 1313, 148 spring wheat (4) 619, 69 self-organizing maps (2) 275, 30 SRP (soluble reactive phosphorus) (2) 201, 22 self-purification (2) 235, 26 Steinernema feltiae (3) 365, 38; (3) 407, 45; (5–6) separation mechanism (7) 903, 101 801, 91; (8) 1065, 119; (8) 1101, 124 sequential analysis (11) 1417, 158 storage (1) 9, 1 sequential extraction (5–6) 805, 92; (9–10) 1227, strawberry (5–6) 777, 88 138; (11) 1533, 171 stream (3) 435, 49 sewage sludge (7) 887, 99; (9–10) 1193, 134; structure change (7) 887, 99 (9–10) 1219, 137; (9–10) 1227, 138; (9–10) sugars (1) 73, 9; (8) 1025, 113 1355, 153; (12) 1659, 185 sulfur (3) 401, 44; (8) 1093, 123 sewage sludges (4) 545, 60 surface horizon (4) 629, 70; (8) 1087, 122 sewer (7) 865, 97 surface microlayer (3) 469, 53; (5–6) 827, 95 sex ratio (5–6) 717, 79 surface TDR probe (7) 877, 98 Sielianinov coefficient (8) 1071, 120 surface water (9–10) 1321, 149 Silesian and Ciezkowickie Foothills (4) 577, 64 survivability (3) 371, 39 silver (3) 371, 39 silver nanoparticles (8) 1065, 119 Taraxacum officinale (8) 1093, 123 Sinapsis alba L. (3) 445, 50 Taxus baccata (3) 453, 51 Sitona sp. (11) 1565, 176 TDR (Time Domain Reflectometry) (7) 877, 98 Sitophilus oryzae L. (5–6) 717, 79 temperature (8) 1071, 120; (12) 1659, 185 sludge classification (11) 1467, 163 1,3,4-thiadiazole derivatives (12) 1691, 189 sludge in sewer system (11) 1467, 163 thrips (1) 83, 10 sludge transport in sewer system (11) 1467, 163 tissues (8) 1047, 116 small radish (1) 129, 16 total content (7) 941, 105 sodium (3) 435, 49; (9–10) 1175, 132 total copper (7) 951, 106 sodium chloride (8) 1105, 125 total heavy metals (9–10) 1347, 152 soil (2) 191, 21; (3) 341, 34; (3) 359, 37; (4) 507, total lead (5–6) 821, 94 55; (4) 531, 58; (4) 563, 62; (4) 571, 63; (5–6) total manganese (7) 951, 106 685, 75; (5–6) 703, 77; (5–6) 771, 87; (8) 1087, total nitrogen (1) 123, 15; (4) 515, 56; (4) 577, 64; 122; (9–10) 1211, 136; (9–10) 1251, 141; (4) 629, 70 (9–10) 1287, 145; (9–10) 1301, 147; (9–10) total organic carbon content (9–10) 1347, 152 1327, 150; (9–10) 1373, 155; (11) 1401, 156; total phosphorus (2) 201, 22 (11) 1417, 158; (11) 1497, 166; (11) 1527, 170; total zinc (7) 951, 106 (11) 1533, 171; (11) 1571, 177; (12) 1711, 191 trace elements (9–10) 1185, 133; (12) 1633, 182 Indexes 1799 traffic pollution (11) 1533, 171 water (2) 191, 21; (8) 1033, 114; (11) 1545, 173 training method (1) 73, 9 water balance (5–6) 763, 86; (5–6) 777, 88; (8) transparent, white and black film (1) 139, 17 1093, 123; (8) 1105, 125 transpiration (5–6) 763, 86 water environment (8) 1147, 131 trees (3) 347, 35 water hydrochemistry (2) 241, 27 tributary of Slupia River (2) 241, 27 water plants (5–6) 743, 83 tributyrin (12) 1727, 194 water pollution (9–10) 1363, 154 Trichoderma spp. (5–6) 679, 74 water protection (2) 289, 31 triticale (3) 461, 52 water quality (2) 235, 26; (2) 289, 31; (4) 637, 71; trophic index (11) 1433, 160 (5–6) 737, 82; (9–10) 1339, 151; (9–10) 1363, Tween (12) 1727, 194 154; (11) 1433, 160 water reservoir (2) 289, 31; (12) 1743, 196 ultrafiltration (12) 1743, 196 water treatment (7) 903, 101 urban soils (5–6) 697, 76 watercourses regulation (12) 1763, 198 water-soluble heavy metals (9–10) 1347, 152 urea (4) 523, 57 well waters (2) 207, 23 UV radiation (3) 469, 53; (5–6) 827, 95 winter wheat (3) 401, 44; (4) 523, 57 wood colonization (2) 183, 20 vegetal cover (8) 1117, 127 vegetation (2) 241, 27; (4) 553, 61 yield (1) 73, 9; (8) 1071, 120; (8) 1079, 121; (11) vermicompost (9–10) 1193, 134; (9–10) 1263, 142; 1505, 167 (12) 1683, 188 yield structure (1) 67, 8; (3) 401, 44 vermiculture (9–10) 1193, 134 yielding (1) 113, 14; (8) 1041, 115 viscosity (12) 1659, 185 vitamin C (1) 67, 8; (1) 73, 9 zeolite (11) 1497, 166 volatile organic compounds (7) 991, 111 zinc (2) 265, 29; (3) 445, 50; (5–6) 673, 73; (8) 1059, 118; (9–10) 1211, 136; (9–10) 1327, 150; waste dump (7) 923, 103 (11) 1533, 171; (12) 1633, 182

INDEKS RZECZOWY

Sposób zapisu odnoœników hase³ – (nr zeszytu) pierwsza strona artyku³u, nr artyku³u (w spisie treœci rocznika).

Acer platanoides L. (3) 377, 40 azotany(III) (1) 129, 16; (4) 571, 63 Acidithiobacillus ferrooxidans (3) 391, 42 azotany(V) (1) 129, 16; (2) 207, 23; (4) 571, 63 Aesculus hippocastanum (3) 453, 51 babka lancetowata (11) 1425, 159 AgroHydrogel (5–6) 777, 88 Bacillus cereus (12) 1727, 194 agroutylizacja œcieków z produkcji t³uszczów ro- Bacillus mycoides (12) 1727, 194 œlinnych (12) 1667, 186 badania mikrobiologiczne (12) 1691, 189 aktywnoœæ antyoksydacyjna (1) 59, 7 bakterie (2) 183, 20; (5–6) 685, 75; (12) 1715, 192 aktywnoœæ dehydrogenazy (3) 385, 41 bakterie z grupy coli (12) 1715, 192 aktywnoœæ emigracyjna (5–6) 717, 79 bakterioneuston (5–6) 827, 95 aktywnoœæ enzymatyczna (5–6) 827, 95 bakteryjne preparaty (4) 619, 69 aktywnoœæ oddechowa (5–6) 827, 95 barwa liœci (1) 83, 10 alginian sodu (8) 1147, 131 barwniki asymilacyjne (3) 377, 40; (5–6) 763, 86; alkalizacja (9–10) 1245, 140 (5–6) 777, 88; (8) 1093, 123; (8) 1105, 125 allelopatia (12) 1615, 180 Baycidal WP 25 (5–6) 749, 84 Alphitobius diaperinus (5–6) 749, 84 bentonit (9–10) 1373, 155; (12) 1641, 183 aluminium (11) 1545, 173 benzyna (9–10) 1373, 155 aminokwasy (4) 523, 57; (4) 545, 60 Betula pendula (5–6) 785, 89 amoniak (4) 637, 71 bia³ko (8) 1025, 113 analiza sekwencyjna (11) 1417, 158 biegaczowate (5–6) 813, 93 analizy roœlin (1) 43, 5 Bieszczadzki Park Narodowy (5–6) 709, 78 antagonizm pierwiastków (3) 413, 46 bilans wodny (5–6) 763, 86; (5–6) 777, 88; (8) antocyjany (1) 67, 8 1093, 123; (8) 1105, 125 anturium (1) 43, 5 bioakumulacja (11) 1545, 173 artemia (3) 371, 39 biodegradacja (11) 1401, 156 Ascochyta fabae (11) 1515, 168 biodegradacja œcieków w kanalizacji (7) 865, 97 asymilacja CO2 (5–6) 763, 86 biodiesel (11) 1401, 156 atmosfera modyfikowana (1) 129, 16 biogaz (7) 923, 103 auksyna (1) 27, 3; (7) 967, 108 biogeny (2) 175, 19; (9–10) 1203, 135 Auricularia auricula-judae (1) 89, 11 biokonwersja azotu (4) 537, 59 Auricularia polytricha (1) 89, 11 biomasa (3) 353, 36; (7) 973, 109; (12) 1721, 193 azot (3) 401, 44; (4) 523, 57; (4) 531, 58; (4) 553, bioreaktor SBR (7) 895, 100 61; (4) 563, 62; (9–10) 1175, 132 Biskupin (2) 183, 20 azot biomasy mikrobiologicznej (4) 577, 64 b³ona biologiczna (7) 865, 97 azot mineralny (4) 585, 65 bobik (8) 1071, 120 azot ogólny (1) 123, 15; (4) 515, 56; (4) 577, 64; bor (1) 19, 2 (4) 629, 70 Botrytis fabae (11) 1515, 168 azot w glebie (4) 601, 67; (4) 611, 68 Bruchus rufimanus Boh. (11) 1515, 168; (12) 1651, azotany (1) 139, 17; (4) 515, 56; (4) 637, 71; 184 (9–10) 1339, 151 burak æwik³owy (1) 19, 2 1802 Indexes

C4 fotosynteza (12) 1615, 180 fitoremediacja (3) 445, 50; (12) 1615, 180 ca³kowite zawartoœci (7) 951, 106 fluazyfop-p-butylowy (7) 967, 108 cebula (1) 19, 2 folia bezbarwna, bia³a i czarna (1) 139, 17 chemometria (2) 275, 30 folie o ró¿nej przepuszczalnoœci dla tlenu (1) 129, chlor (5–6) 723, 80 16 chlorek sodu (8) 1105, 125 formy azotu (1) 73, 9; (4) 507, 55 chlorofil (8) 1025, 113 formy o³owiu (8) 1131, 129 chrom (11) 1533, 171 formy rozpuszczalne (7) 941, 105 chrom(III) (11) 1497, 166 formy specjacyjne metali (12) 1667, 186 chrom(VI) (11) 1497, 166 fosfor (2) 219, 24 ciek (3) 435, 49 fosfor ca³kowity (2) 201, 22 ciemiernik korsykañski (1) 51, 6 fosforan jednowapniowy (7) 983, 110 ciœnieniowe techniki membranowe (12) 1751, 197 fosforan paszowy (7) 983, 110 cukry (1) 73, 9; (8) 1025, 113 frakcja aerozolowa (2) 275, 30 cynk (2) 265, 29; (3) 445, 50; (5–6) 673, 73; (7) Fraxinus excelsior (3) 453, 51 951, 106; (8) 1059, 118; (9–10) 1211, 136; Fusarium (3) 353, 36; (12) 1721, 193 (9–10) 1327, 150; (11) 1533, 171; (12) 1633, 182 Galleria mellonella (3) 365, 38; (8) 1101, 124; czerwce (3) 397, 43 (11) 1575, 178 czynniki abiotyczne (5–6) 665, 72 gatunek gleby (9–10) 1313, 148 czynniki glebotwórcze (4) 553, 61 geostatystyka (2) 301, 32 gêstoœæ siewu (5–6) 793, 90 Dar 2,5GR (9–10) 1263, 142 gleba (2) 191, 21; (3) 341, 34; (3) 359, 37; (4) 507, dawka polielektrolitu (7) 887, 99 55; (4) 531, 58; (4) 563, 62; (4) 571, 63; (5–6) dawki azotu (1) 73, 9 685, 75; (5–6) 703, 77; (5–6) 771, 87; (8) 1087, dawki i formy mikroelementów (4) 571, 63 122; (9–10) 1211, 136; (9–10) 1245, 140; deszczowanie (4) 507, 55 (9–10) 1251, 141; (9–10) 1287, 145; (9–10) 3,4-dimetylopyrazolofosfat (DMPP) (1) 97, 12 1301, 147; (9–10) 1327, 150; (9–10) 1373, 155; Dimilin 25WP (9–10) 1263, 142; (11) 1557, 175 (11) 1401, 156; (11) 1417, 158; (11) 1527, 170; Diptera (11) 1557, 175 (11) 1533, 171; (11) 1571, 177; (12) 1711, 191 d³ugotrwa³e doœwiadczenie nawozowe (9–10) gleby aluwialne (9–10) 1279, 144 1251, 141 gleby górskie (4) 553, 61 dodatek paszowy (7) 983, 110 gleby lekkie (8) 1131, 129; (11) 1505, 167 dolistne dokarmianie (4) 523, 57 gleby leœne (5–6) 821, 94 dorzecze S³upi (2) 241, 27 gleby miejskie (5–6) 697, 76 doœwiadczenia wieloletnie (4) 601, 67; (4) 611, 68 gleby uprawne (3) 385, 41 doœwiadczenie wazonowe (3) 401, 44 gnojowica (12) 1743, 196 drobnoustroje glebowe (5–6) 821, 94 gorczyca bia³a (3) 445, 50 drzewa (3) 347, 35 gospodarka fosforowa (5–6) 731, 81 dynia (1) 9, 1 granulometria (11) 1457, 162 d¿d¿ownice D. veneta (9–10) 1263, 142 groch (1) 83, 10 d¿d¿ownicowa skrzynka ekologiczna (11) 1557, grzyby (7) 959, 107; (12) 1705, 190; (12) 1715, 175 192 grzyby entomopatogenne (3) 359, 37; (11) 1571, E. fetida (5–6) 703, 77; (11) 1557, 175 177; (12) 1711, 191 efektywnoœæ (4) 619, 69 grzyby mikroskopowe (2) 183, 20 ekstrakcja sekwencyjna (5–6) 805, 92; (9–10) 1227, 138; (11) 1533, 171 halofity (2) 225, 25 elektrokoagulacja (7) 911, 102 hamowanie (11) 1551, 174 etefon (1) 27, 3 herbaty czarne (9–10) 1185, 133 eutrofizacja (11) 1433, 160 herbaty zielone (9–10) 1185, 133 herbicydy (7) 959, 107 F. culmorum (11) 1477, 164; (12) 1625, 181 Heterorhabditis bacteriophora (5–6) 749, 84; (8) Fagus sylvatica L. (5–6) 709, 78 1065, 119 fauna glebowa (11) 1527, 170 Heterorhabditis megidis (8) 1139, 130; (11) 1575, fenamidon (11) 1551, 174 178 Indexes 1803 hipertrofia (11) 1441, 161 kwasy humusowe (11) 1407, 157 hydrochemia wód (2) 241, 27 hydro¿elowe mikrokapsu³ki (8) 1147, 131 Lactuca sativa var. Capitata (8) 1105, 125 larwy Phrynohyas resinifictrix (5–6) 673, 73 impulsowe pole magnetyczne (11) 1477, 164 len oleisty (2) 265, 29 inhibitor nitryfikacji (1) 97, 12 len w³óknisty (2) 265, 29 insektycyd (5–6) 749, 84 Lentinula edodes (1) 89, 11 izotermy adsorpcji (2) 191, 21 lepkoœæ (12) 1659, 185 j liczebnoœæ populacji (5–6) 717, 79 akoœæ i wartoœæ pokarmowa (8) 1111, 126 Linum usitatissimum L. (2) 265, 29 jakoœæ korzeni (1) 113, 14 lipazy (12) 1727, 194 jakoœæ powietrza wewnêtrznego (7) 991, 111 liœcie (9–10) 1185, 133 jakoœæ wód (9–10) 1339, 151; (9–10) 1363, 154; lit (9–10) 1279, 144 (11) 1433, 160 Lolium perenne (12) 1675, 187 jakoœæ wody (2) 235, 26; (2) 289, 31; (4) 637, 71; lotne zwi¹zki organiczne (VOCs) (7) 991, 111 (5–6) 737, 82 lucerna (9–10) 1313, 148 jeziora (5–6) 805, 92; (12) 1633, 182 jeziora lobeliowe (9–10) 1293, 146 ³adunek wyniesiony (11) 1539, 172 jeziora polimiktyczne (11) 1441, 161 ³¹ka górska (8) 1041, 115 jony amonowe (1) 139, 17 ³ubin w¹skolistny (4) 619, 69 jony o³owiu (8) 1101, 124; (11) 1575, 178 magnez (5–6) 673, 73; (8) 1059, 118; (9–10) 1175, kadm (2) 265, 29; (3) 365, 38; (5–6) 673, 73; (5–6) 132 763, 86; (5–6) 771, 87; (8) 1059, 118; (9–10) makro- i mikroelementy (3) 419, 47 1211, 136; (11) 1425, 159 kalibracja modelu hydraulicznego sieci kanalizacy- makroelementy (1) 43, 5; (8) 1079, 121; (9–10) jnej (7) 853, 96 1175, 132; (11) 1505, 167 kanalizacja grawitacyjna (7) 865, 97; (11) 1467, makrofauna (8) 1117, 127 163 makrofity rzeczne (2) 241, 27 karpie (8) 1047, 116 makropierwiastki (12) 1683, 188 kaskada So³y (11) 1433, 160 makrosk³adniki (8) 1033, 114; (11) 1539, 172 klasyfikacja (2) 275, 30 makuch rzepakowy (3) 335, 33 klasyfikacja osadów (11) 1467, 163 mangan (7) 951, 106; (8) 1059, 118; (9–10) 1279, kolonizacja drewna (2) 183, 20 144; (9–10) 1327, 150 kompleks sorpcyjny (9–10) 1287, 145 mapy samoorganizuj¹ce siê (2) 275, 30 kompost (1) 147, 18; (3) 347, 35; (4) 545, 60; marchew (1) 19, 2; (1) 113, 14 (9–10) 1193, 134; (9–10) 1219, 137; (9–10) materia organiczna gleby (4) 601, 67; (11) 1457, 1227, 138; (9–10) 1355, 153; (9–10) 1373, 155; 162 (11) 1497, 166; (12) 1641, 183 materia³y budowlane (7) 877, 98 kompost z odpadów komunalnych (9–10) 1327, materia³y organiczne (9–10) 1355, 153 150 mechanizm separacji (7) 903, 101 kompost z zieleni miejskiej (9–10) 1327, 150 metalaksyl (11) 1551, 174 kompostowanie odpadów miejskich (4) 497, 54 metale (5–6) 805, 92 kondycjonowanie (12) 1659, 185 metale ciê¿kie (1) 97, 12; (1) 105, 13; (2) 191, 21; konie (8) 1053, 117 (2) 235, 26; (2) 301, 32; (3) 347, 35; (3) 419, kopalnia (5–6) 737, 82 47; (3) 453, 51; (3) 461, 52; (4) 563, 62; (5–6) krajobraz (4) 563, 62 697, 76; (5–6) 709, 78 ; (5–6) 785, 89; (7) 941, Kraków (5–6) 697, 76 105; (8) 1101, 124; (9–10) 1219, 137; (9–10) kuchenne odpady organiczne (11) 1557, 175 1237, 139; (9–10) 1279, 144; (9–10) 1293, 146; kukurydza (4) 545, 60; (9–10) 1355, 153 (11) 1515, 168; (11) 1527, 170; (11) 1565, 176; kurczêta brojlery (3) 335, 33 (12) 1651, 184; (12) 1667, 186 kwas askorbinowy (8) 1025, 113 metan (7) 923, 103 kwas b-naftoksyoctowy (7) 967, 108 metody dielektryczne (7) 877, 98 kwasowoœæ (11) 1497, 166 metody oceny (9–10) 1211, 136 kwasy fulwowe (9–10) 1347, 152 metody prowadzenia (1) 73, 9 kwasy huminowe (9–10) 1347, 152 mezofauna (8) 1117, 127 1804 Indexes miedŸ (3) 445, 50; (5–6) 673, 73; (7) 951, 106; obornik (4) 545, 60 (9–10) 1211, 136; (9–10) 1327, 150; (11) 1533, obredlanie (12) 1599, 179 171 ocena towaroznawcza (7) 991, 111 mieszane agrofitocenozy (4) 619, 69 ochrona wód (2) 289, 31 mikotoksyny (5–6) 665, 72 oczyszczanie œcieków koksowniczych (12) 1751, mikroelementy (1) 43, 5; (4) 523, 57; (8) 1079, 121 197 mikroorganizmy (3) 341, 34; (4) 563, 62; (7) 895, oczyszczanie wody (7) 903, 101 100; (11) 1401, 156 odcieki (9–10) 1203, 135 mikrowarstwa powierzchniowa (3) 469, 53; (5–6) odcieki sk³adowiskowe (7) 895, 100 827, 95 odczyn gleby (9–10) 1347, 152 mineralizacja (2) 255, 28 odmiany (1) 9, 1; (8) 1019, 112; (12) 1675, 187 mineralny sk³ad (5–6) 673, 73 odmiany grochu (5–6) 731, 81 misecznik tarniowy (3) 397, 43 odnawialne Ÿród³a energii (OZE) (7) 923, 103 miskant olbrzymi (12) 1599, 179 odpady paleniskowe (9–10) 1271, 143 miskantus (12) 1615, 180 odzysk wody (12) 1743, 196 mniszek pospolity (11) 1425, 159 Ojcowski Park Narodowy (5–6) 771, 87 mocznik (4) 523, 57 okresowe zmiany (1) 43, 5 mogilnik (2) 191, 21 okrywa roœlinna (8) 1033, 114; (8) 1117, 127; (11) monokultura (3) 461, 52 1539, 172 motylkowate (4) 531, 58 olej napêdowy (9–10) 1373, 155 mykoestrogeny (7) 903, 101 o³ów (1) 19, 2; (2) 265, 29; (3) 365, 38; (5–6) 821, 94; (9–10) 1211, 136; (9–10) 1227, 138; (11) NAA (neutronowa analiza aktywacyjna) (1) 27, 3 1417, 158; (12) 1633, 182 naczyniowe roœliny wodne (12) 1763, 198 opad atmosferyczny (4) 515, 56 nanocz¹stki srebra (8) 1065, 119 opady (8) 1071, 120 nanofiltracja (7) 903, 101; (12) 1743, 196 opornoœæ na As(III) i Sb(III) (3) 391, 42 Nanogro (12) 1625, 181 opróbkowanie dodatkowe (2) 301, 32 nanorurki wêglowe (5–6) 757, 85 optymalizacja parametrów (8) 1139, 130 napary (9–10) 1185, 133 organiczne odpady kuchenne (12) 1683, 188 nawo¿enie (1) 123, 15; (3) 359, 37; (4) 507, 55; (4) osad czynny (7) 895, 100 601, 67; (4) 611, 68; (8) 1019, 112; (8) 1033, osady denne (2) 219, 24; (5–6) 805, 92; (7) 941, 114; (8) 1041, 115; (8) 1117, 127; (8) 1123, 105; (9–10) 1175, 132; (9–10) 1293, 146; (11) 128; (11) 1407, 157; (11) 1539, 172; (12) 1675, 1457, 162; (11) 1505, 167; (12) 1633, 182 187 osady œciekowe (4) 545, 60; (7) 887, 99; (9–10) nawo¿enie azotem (1) 51, 6; (4) 593, 66; (12) 1599, 1193, 134; (9–10) 1219, 137; (9–10) 1227, 138; 179 (9–10) 1355, 153; (12) 1659, 185 nawo¿enie dolistne (1) 35, 4 osady w kanalizacji (11) 1467, 163 nawo¿enie mikroelementami (8) 1111, 126; (12) otulina parku (8) 1131, 129 1737, 195 owady po¿yteczne (3) 425, 48 nawozy azotowe (4) 619, 69 owies (4) 619, 69; (5–6) 793, 90 nawozy dolistne (3) 353, 36; (12) 1721, 193 Owinema (8) 1065, 119 Nematop (5–6) 749, 84; (8) 1065, 119 owocniki (1) 89, 11 nicienie entomopatogenne (3) 359, 37; (3) 397, 43; (3) 407, 45; (5–6) 749, 84; (5–6) 801, 91; (8) parametry fizykochemiczne (2) 183, 20 1065, 119; (8) 1101, 124; (11) 1575, 178; (12) parki rekreacyjne (9–10) 1211, 136 1711, 191 Pentas lanceolata (1) 147, 18 nicienie owadobójcze (5–6) 757, 85 pestycydy (2) 191, 21; (5–6) 703, 77 niedobór potasu (5–6) 731, 81 pH (8) 1041, 115; (12) 1683, 188 nikiel (3) 445, 50 Photorhabdus luminescens (8) 1139, 130 niskonak³adowe metody (9–10) 1193, 134 Phytophthora spp. (11) 1551, 174 N-NO3 (1) 123, 15 piasek (4) 571, 63 Nomolt 150SC (9–10) 1263, 142 pierwiastki œladowe (9–10) 1185, 133; (12) 1633, NPK (4) 545, 60 182 p³az (5–6) 673, 73 ober¿yna (1) 73, 9 Pleurotus eryngii (1) 89, 11 obni¿enie zawartoœci azotu (4) 537, 59 p³odozmian zbo¿owy (4) 507, 55 Indexes 1805 plon (1) 73, 9; (8) 1079, 121; (11) 1505, 167 roboty konserwacyjne (12) 1763, 198 plon nasion (8) 1019, 112; (12) 1675, 187 roœlinnoœæ (2) 241, 27 plon roœlin (4) 593, 66 roœliny (11) 1485, 165; (12) 1641, 183 plon ziarna (5–6) 793, 90 roœliny energetyczne (7) 973, 109 plonowanie (1) 113, 14; (8) 1041, 115 roœliny wodne (11) 1545, 173; (5–6) 743, 83 p³ynne hodowle in vitro (8) 1139, 130 rozmiar fraktalny (7) 911, 102 PM1.0 (7) 933, 104 ró¿norodnoœæ (11) 1527, 170 PM2.5 (7) 933, 104 Roztoczañski Park Narodowy (8) 1131, 129 PM10 (7) 933, 104 rtêæ (8) 1087, 122; (9–10) 1245, 140; (9–10) 1251, Poa pratensis (8) 1019, 112 141 pochodne 1,3,4-tiadiazolu (12) 1691, 189 ruñ (8) 1111, 126; (9–10) 1251, 141; (11) 1521, Pogórze Œl¹skie i Ciê¿kowickie (4) 577, 64 169; (12) 1737, 195 pole produkcyjne (4) 629, 70 ryzyko ekologiczne (2) 301, 32 poligon wojskowy (5–6) 743, 83 ryzyko zdrowotne (2) 207, 23 polisulfonowe membrany ultrafiltracyjne (12) rzeka Kwacza (2) 241, 27 1751, 197 rzeka Pe³cznica (5–6) 737, 82 popió³ (7) 973, 109 rzeki (11) 1545, 173 popió³ fluidalny (12) 1715, 192 rzeki nizinne (2) 201, 22 potas (8) 1059, 118; (9–10) 1175, 132 rzodkiewka (1) 19, 2; (1) 129, 16 powierzchnia liœci (5–6) 777, 88 powierzchniowa sonda TDR (7) 877, 98 Saccharomyces cerevisiae (7) 967, 108 powietrze wewnêtrzne i zewnêtrzne (7) 933, 104 sa³ata (1) 19, 2; (4) 571, 63 poziom chlorofili (1) 83, 10 Salix viminalis (5–6) 763, 86 poziom powierzchniowy (4) 629, 70; (8) 1087, 122 samooczyszczanie (2) 235, 26 Poznañ (9–10) 1211, 136 Scardinius erythrophthalmus (3) 419, 47 preparat enzymatyczny (3) 335, 33 Secale cereale L. (8) 1059, 118 próbkowanie sezonowe (2) 275, 30 selen (5–6) 743, 83; (8) 1047, 116; (8) 1053, 117; procesy limniczne (2) 225, 25 (9–10) 1313, 148 procesy membranowe (12) 1743, 196 seler (8) 1025, 113 produkcja pierwotna (11) 1441, 161 sezon fenologiczny (11) 1571, 177 prolina (12) 1705, 190 siano (3) 413, 46 promieniowanie jonizuj¹ce (3) 407, 45; (5–6) 801, siarka (3) 401, 44; (8) 1093, 123 91 Sitona sp. (11) 1565, 176 promieniowanie UV (3) 469, 53; (5–6) 827, 95 sk³ad chemiczny (1) 9, 1; (1) 89, 11; (5–6) 793, 90; promieniowce (12) 1715, 192 (7) 933, 104; (7) 973, 109; (12) 1737, 195 przechowanie (1) 9, 1 sk³adniki mineralne (9–10) 1339, 151 przemys³ mleczarski (9–10) 1193, 134 sk³adniki organiczne (11) 1521, 169 prze¿ywalnoœæ (3) 371, 39 sk³adowisko odpadów (3) 425, 48; (5–6) 685, 75; pszenica jara (4) 619, 69 (5–6) 813, 93 (7) 923, 103 pszenica ozima (3) 401, 44; (4) 523, 57 skrzynka ekologiczna (9–10) 1263, 142 pszen¿yto (3) 461, 52 skutecznoœæ zbiornika wstêpnego (2) 289, 31 pu³apki ró¿anecznikowe (11) 1551, 174 sód (3) 435, 49; (9–10) 1175, 132 py³y cementowe (9–10) 1245, 140 soja (8) 1079, 121 py³y wapienne (9–10) 1287, 145 sorpcja (9–10) 1301, 147 sposób u¿ytkowania gleb (4) 577, 64 redukcja ³adunku (3) 435, 49 sposób u¿ytkowania terenu (4) 585, 65 reduktaza azotanowa (1) 139, 17 srebro (3) 371, 39 reflektometryczne metody pomiaru wilgotnoœci (7) SRP (rozpuszczalny fosfor reaktywny) (2) 201, 22 877, 98 stabilizacja beztlenowa (7) 923, 103 regulacja cieków (12) 1763, 198 stan sanitarny (9–10) 1237, 139 regulatory wzrostu (4) 619, 69 staw (3) 435, 49 rekultywacja (2) 219, 2; (9–10) 1271, 1434 Steinernema feltiae (3) 365, 38; (3) 407, 45; (8) relacje miêdzy sk³adnikami (1) 43, 5 1065, 119; (8) 1101, 124 renaturyzacja (2) 241, 27 stosunek C/N (4) 553, 61; (11) 1457, 162 robinia akacjowa (3) 385, 41; (5–6) 679, 74 struktura plonu (1) 67, 8; (3) 401, 44 Robinia pseudoacacia (3) 341, 34 studnie kopane (2) 207, 23 1806 Indexes substancja organiczna (11) 1407, 157 uwalnianie metazachloru (8) 1147, 131, substancje humusowe (3) 469, 53; (9–10) 1347, uwodnienie koñcowe (7) 887, 99 152 u¿ytki zielone (4) 531, 58 substancje neutralizuj¹ce (11) 1485, 165 substancje ropopochodne (9–10) 1321, 149 wapñ (8) 1059, 118; (9–10) 1175, 132 surfaktant kationowy (9–10) 1301, 147 wapno i wapnowanie (8) 1041, 115; (12) 1641, 183 synteza bia³ek (3) 469, 53 warstwa orna (3) 461, 52 synteza DNA (3) 469, 53 wartoœæ pokarmowa (11) 1521, 169 system antyoksydacyjny (7) 967, 108 warunki siedliskowe (2) 255, 28 system rzeczno-jeziorny (11) 1441, 161 wciornastki (1) 83, 10 systemy melioracyjne (9–10) 1339, 151 wermikompost (9–10) 1193, 134; (9–10) 1263, szczaw zwyczajny (11) 1425, 159 142; (12) 1683, 188 szkodniki (3) 425, 48 wermikultura (9–10) 1193, 134 szpinak (1) 19, 2 wêgiel brunatny (11) 1407, 157; (12) 1641, 183 wêgiel w glebie (4) 601, 67 œcieki celulozowo-papiernicze (7) 911, 102 wieloletnie doœwiadczenie (7) 951, 106 œcieki mleczarskie (2) 175, 19 wielosk³adnikowe nawozy kompleksowe (1) 123, œrodowisko wodne (8) 1147, 131 15 œwie¿e namu³y (9–10) 1279, 144 wilgotnoœæ przegród budowlanych (7) 877, 98 winogrona (1) 59, 7 Taraxacum officinale (8) 1093, 123 wirowanie (7) 887, 99 Taxus baccata (3) 453, 51 witamina C (1) 67, 8; (1) 73, 9 TDR (Time Domain Reflectometry) (7) 877, 98 w³aœciwoœci chemiczne (9–10) 1271, 143 TDTP (9–10) 1185, 133 w³aœciwoœci gleby (9–10) 1373, 155 temperatura (8) 1071, 120; (12) 1659, 185 w³aœciwoœci sorpcyjne (11) 1497, 166 tereny chronione (5–6) 821, 94 w³osy (8) 1053, 117 tereny produkcyjne (8) 1131, 129 woda (2) 191, 21; (8) 1033, 114; (11) 1539, 172; tereny rekultywowane (4) 593, 66 (11) 1545, 173 terminy zbioru (12) 1599, 179 woda gruntowa (2) 255, 28 tkanki (8) 1047, 116 woda powierzchniowa (9–10) 1321, 149 tlenek wapnia (9–10) 1373, 155; (11) 1497, 166 wodnorozpuszczalne formy metali ciê¿kich (9–10) t³o geochemiczne (9–10) 1211, 136 1347, 152 tolerancja na ch³ód (12) 1615, 180 wody interstycjalne (2) 201, 22 topliwoœæ (7) 973, 109 wody opadowe (2) 235, 26 torf (4) 571, 63 wody podziemne (4) 637, 71 torfowisko wysokie (2) 255, 28 wody s³one (2) 225, 25 transformacja azotu (4) 497, 54 wody œródl¹dowe (11) 1433, 160 transpiracja (5–6) 763, 86 wody studzienne (2) 207, 23 transport (9–10) 1321, 149 wo³ek ry¿owy (5–6) 717, 79 transport osadów w kanalizacji (11) 1467, 163 wskaŸnik p³ci (5–6) 717, 79 trasa szybkiego ruchu (11) 1425, 159 wskaŸnik sanitarny œrodowiska (12) 1691, 189 tributyryna (12) 1727, 194 wskaŸnik trofii (11) 1433, 160 Trichoderma spp. (5–6) 679, 74 wspó³czynnik Sielianinowa (8) 1071, 120 trociny (1) 89, 11 wspó³czynnik szorstkoœci (7) 853, 96 truskawka (5–6) 777, 88 wychmieliny (11) 1407, 157 Tween (12) 1727, 194 wycierka ziemniaczana (1) 147, 18 wykrywanie (11) 1551, 174 uk³ady zintegrowane (12) 1751, 197 wymiana gazowa (1) 35, 4 ultrafiltracja (12) 1743, 196 wymywanie azotu (1) 97, 12; (1) 105, 13 uprawa (1) 147, 18 wywar gorzelniczy (4) 537, 59 uprawa ekologiczna (1) 113, 14 wzór Manninga (7) 853, 96 uprawa integrowana (1) 113, 14 wzrost liniowy (3) 353, 36; (12) 1625, 18; (12) uprawa konwencjonalna (1) 113, 14 1721, 193 uprawa w doniczkach (1) 51, 6 wzrost roœlin (1) 35, 4 usuwanie mikrozanieczyszczeñ (7) 903, 101 utlenianie Fe(II) (3) 391, 42 zagêszczanie grawitacyjne (7) 887, 99 Indexes 1807 zagêszczenie (11) 1527, 170 zawartoœæ magnezu (11) 1485, 165 zagroda wiejska (2) 207, 23 zawartoœæ makroelementów (3) 413, 46 zanieczyszczenia (9–10) 1363, 154 zbiornik ma³ej retencji (9–10) 1363, 154 zanieczyszczenia antropogenne (3) 397, 43 zbiornik wodny (2) 289, 31 zanieczyszczenia komunikacyjne (11) 1533, 171 zbiornik wstêpny (2) 289, 31 zanieczyszczenie (5–6) 697, 76; (5–6) 743, 83; zbiorniki antropogenne (2) 225, 25 (5–6) 771, 87; (5–6) 785, 89; (7) 959, 107; zbiorniki retencyjne (2) 235, 26; (5–6) 723, 80 (9–10) 1373, 155; (11) 1497, 166 zbiorniki wodne terenów zalewowych (11) 1457, zanieczyszczenie arsenem (11) 1485, 165 162 zanieczyszczenie gleby (3) 445, 50; (11) 1515, zbo¿a (5–6) 665, 72 168; (11) 1565, 176; (12) 1651, 184 zeolit (11) 1497, 166 zanieczyszczenie kadmem (12) 1641, 183 ziarno (5–6) 665, 72 zanieczyszczenie œrodowiska (3) 347, 35 ziemniak (1) 123, 15; (8) 1123, 128 zarodnikowanie (3) 353, 36; (12) 1625, 181; (12) zio³a (4) 531, 58 1721, 193 zasolenie (8) 1025, 113; (12) 1683, 188; (12) 1705, zlewnia (12) 1633, 182 190 zlewnia rolnicza (5–6) 723, 80 zatê¿ony kwas fosforowy (7) 983, 110 z³o¿a hydrofilowe (9–10) 1203, 135; (2) 175, 19 zawartoœæ (9–10) 1175, 132 zmiana struktury (7) 887, 99 zawartoœæ azotanów(V) (8) 1123, 128 zmiennoœæ przestrzenna (4) 629, 70; (8) 1087, 122 zawartoœæ ca³kowita (7) 941, 105 zmywacze do paznokci (7) 991, 111 zawartoœæ ca³kowita metali ciê¿kich (9–10) 1347, zwi¹zki azotu (4) 637, 71 152 zwi¹zki biogenne (9–10) 1363, 154 zawartoœæ ca³kowita wêgla organicznego (9–10) zwi¹zki fenolowe (1) 59, 7 1347, 152 zwi¹zki organiczne (9–10) 1203, 135 zawartoœæ chlorofilu (1) 35, 4 zawartoœæ glinu (12) 1641, 183 ¿yto (8) 1059, 118

INDEX OF LATIN, POLISH AND ENGLISH SPECIES NAMES OF MICROORGANISMS, PLANTS AND ANIMALS AND THEIR ANATOMICAL PARTS

WYKAZ £ACIÑSKICH, POLSKICH I ANGIELSKICH NAZW MIKROORGANIZMÓW, ROŒLIN I ZWIERZ¥T I ICH CZÊŒCI ANATOMICZNYCH

Meaning of the digits in the index entries – (no. of issue) first page, no. of the article (in the volume contents). Sposób zapisu odnoœników hase³ – (nr zeszytu) pierwsza strona artyku³u, nr artyku³u (w spisie treœci rocznika).

Bruchus rufimanus Boh. (11) 1515, 168; (12) 1651, cer platanoides L. (3) 377, 40 A 184 Acidithiobacillus ferrooxidans (3) 391, 42 burak æwik³owy (1) 19, 2 actinomycetes (12) 1715, 192 Aesculus hippocastanum (3) 453, 51 carp (8) 1047, 116 alfalfa (9–10) 1313, 148 carrot (1) 19, 2; (1) 113, 14 Alphitobius diaperinus (5–6) 749, 84 cebula (1) 19, 2 amphibian (5–6) 673, 73 celery (8) 1025, 113 anthurium (1) 43, 5 cereals (5–6) 665, 72 anturium (1) 43, 5 ciemiernik korsykañski (1) 51, 6 artemia (3) 371, 39 Ascochyta fabae (11) 1515, 168 Col. Carabidae (5–6) 813, 93 Auricularia auricula-judae (1) 89, 11 Col. Staphylinidae (5–6) 813, 93 Auricularia polytricha (1) 89, 11 coli bacteria (12) 1715, 192 common dandelion (11) 1425, 159 babka lancetowata (11) 1425, 159 corsican hellebore (1) 51, 6 Bacillus cereus (12) 1727, 194 czerwce (3) 397, 43 Bacillus mycoides (12) 1727, 194 bacteria (2) 183, 20; (5–6) 685, 75; (12) 1715, 192 Diptera (11) 1557, 175 bakterie (2) 183, 20; (5–6) 685, 75; (12) 1715, 192 drobnoustroje glebowe (5–6) 821, 94 bakterie z grupy coli (12) 1715, 192 drzewa (3) 347, 35 Betula pendula (5–6) 785, 89 d¿d¿ownice D. Reneta (9–10) 1263, 142 biegaczowate (5–6) 813, 93 black and green teas (9–10) 1185, 133 E. fetida (11) 1557, 175 black locust (3) 385, 41; (5–6) 679, 74 earthworm Dendrobena veneta (9–10) 1263, 142 bobik (8) 1071, 120 eggplant (1) 73, 9 Botrytis fabae (11) 1515, 168 Eisenia fetida (5–6) 703, 77 brine shrimp (3) 371, 39 entomopathogenic fungi (3) 359, 37; (11) 1571, broiler chickens (3) 335, 33 177; (12) 1711, 191 1810 Indexes entomopathogenic nematodes (3) 359, 37; (3) 397, Linum usitatissimum L. (2) 265, 29 43; (3) 407, 45; (5–6) 749, 84; (5–6) 757, 85; liœcie (9–10) 1185, 133 (5–6) 801, 91; (8) 1065, 119; (8) 1101, 124; Lolium perenne (12) 1675, 187 (11) 1575, 178; (12) 1711, 191 lucerna (9–10) 1313, 148

F. culmorum (11) 1477, 164; (12) 1625, 181 ³ubin w¹skolistny (4) 619, 69 Fagus sylvatica L. (5–6) 709, 78 fauna glebowa (11) 1527, 170 macrofauna (8) 1117, 127 flax (2) 265, 29 maize (4) 545, 60; (9–10) 1355, 153 Fraxinus excelsior (3) 453, 51 makrofauna (8) 1117, 127 fruiting bodies (1) 89, 11 makrofity rzeczne (2) 241, 27 fungi (7) 959, 107; (12) 1705, 190; (12) 1715, 192 marchew (1) 19, 2; (1) 113, 14 Fusarium (3) 353, 36; (12) 1721, 193 meadow (9–10) 1251, 141 meadow sward (8) 1111, 126; (11) 1521, 169; (12) Galleria mellonella (3) 365, 38; (8) 1101, 124; 1737, 195 (11) 1575, 178 mesofauna (8) 1117, 127 garden sorrel (11) 1425, 159 mezofauna (8) 1117, 127 globose scale (3) 397, 43 microscopic fungi (2) 183, 20 gorczyca bia³a (3) 445, 50 Miscanthus (12) 1599, 179; (12) 1615, 180 grain (5–6) 665, 72 misecznik tarniowy (3) 397, 43 grapes (1) 59, 7 miskant olbrzymi (12) 1599, 179 greater plantain (11) 1425, 159 miskantus (12) 1615, 180 groch (1) 83, 10 mniszek pospolity (11) 1425, 159 grzyby (7) 959, 107; (12) 1705, 190; (12) 1715, motylkowate (4) 531, 58 192 naked oats (5–6) 793, 90 grzyby entomopatogenne (3) 359, 37; (11) 1571, narrow-leaved lupine (4) 619, 69 177; (12) 1711, 191 nicienie entomopatogenne (3) 359, 37; (3) 397, 43; grzyby mikroskopowe (2) 183, 20 (3) 407, 45; (5–6) 749, 84; (5–6) 801, 91; (8) 1065, 119; (8) 1101, 124; (12) 1711, 191 hair (8) 1053, 117 nicienie owadobójcze (5–6) 757, 85 hay (3) 413, 46 herbaty czarne (9–10) 1185, 133 oat (4) 619, 69 herbaty zielone (9–10) 1185, 133 ober¿yna (1) 73, 9 herbs (4) 531, 58 onion (1) 19, 2 Heterorhabditis bacteriophora (5–6) 749, 84; (8) owies (4) 619, 69 1065, 119 owies nieoplewiony (5–6) 793, 90 Heterorhabditis megidis (8) 1139, 130; (11) 1575, owies oplewiony (5–6) 793, 90 178 owocniki (1) 89, 11 horses (8) 1053, 117 husked oats (5–6) 793, 90 peas (1) 83, 10 pedofauna (11) 1527, 170 karpie (8) 1047, 116 Pentas lanceolata (1) 147, 18 konie (8) 1053, 117 Photorhabdus luminescens (8) 1139, 130 kukurydza (4) 545, 60; (9–10) 1355, 153 Phrynohyas resinifictrix larvae (5–6) 673, 73 kurczêta brojlery (3) 335, 33 Phytophthora spp. (11) 1551, 174 kusakowate (5–6) 813, 93 p³az (5–6) 673, 73 Pleurotus eryngii (1) 89, 11 Lactuca sativa var. Capitata (8) 1105, 125 Poa pratensis (8) 1019, 112 larwy Phrynohyas resinifictrix (5–6) 673, 73 potato (1) 123, 15 leaves (9–10) 1185, 133 potato tubers (8) 1123, 128 legumes (4) 531, 58 promieniowce (12) 1715, 192 len oleisty (2) 265, 29 pszenica jara (4) 619, 69 len w³óknisty (2) 265, 29 pszenica ozima (3) 401, 44; (4) 523, 57 Lentinula edodes (1) 89, 11 pszen¿yto (3) 461, 52 lettuce (1) 19, 2; (4) 571, 63 linseed (2) 265, 29 radish (1) 19, 2; (9–10) 1237, 139 Indexes 1811 red beet (1) 19, 2 Steinernema feltiae (3) 365, 38; (3) 407, 45; (5–6) river macrophytes (2) 241, 27 801, 91; (8) 1065, 119; (8) 1101, 124 robinia akacjowa (3) 385, 41; (5–6) 679, 74 strawberry (5–6) 777, 88 Robinia pseudoacacia (3) 341, 34 szczaw zwyczajny (11) 1425, 159 roœlinnoœæ (2) 241, 27 szpinak (1) 19, 2 roœliny wodne (5–6) 743, 83 araxacum officinale (8) 1093, 123 ruñ (8) 1111, 126; (9–10) 1251, 141; (11) 1521, T Taxus baccata (3) 453, 51 169; (12) 1737, 195 thrips (1) 83, 10 rye (8) 1059, 118 trees (3) 347, 35 rzodkiewka (1) 19, 2; (1) 129, 16; (9–10) 1237, 139 Trichoderma spp. (5–6) 679, 74 triticale (3) 461, 52 accharomyces cerevisiae (7) 967, 108 S truskawka (5–6) 777, 88 sa³ata (1) 19, 2; (4) 571, 63 Salix viminalis (5–6) 763, 86 vegetation (2) 241, 27 scale insects (3) 397, 43 Scardinius erythrophthalmus (3) 419, 47 water plants (5–6) 743, 83 Secale cereale L. (8) 1059, 118 wciornastki (1) 83, 10 seler (8) 1025, 113 winogrona (1) 59, 7 siano (3) 413, 46 winter wheat (3) 401, 44; (4) 523, 57 Sinapsis alba L. (3) 445, 50 w³osy (8) 1053, 117 Sitona sp. (11) 1565, 176 wo³ek ry¿owy (5–6) 717, 79 Sitophilus oryzae L. (5–6) 717, 79 yield (8) 1071, 120 small radish (1) 129, 16 soil microorganisms (5–6) 821, 94 zbo¿a (5–6) 665, 72 soja (8) 1079, 121 ziarno (5–6) 665, 72 soybean (8) 1079, 121 ziemniak (1) 123, 15; (8) 1123, 128 spinach (1) 19, 2 zio³a (4) 531, 58 spring wheat (4) 619, 69 ¿yto (8) 1059, 118

INDEX OF ACRONYMS

Meaning of the digits in the index entries – (no. of issue) first page, no. of the article (in the volume contents).

DMPP – 3,4-dimethylpyrazol phosphate (1) 97, 12 PTWI – permissible temporary weekly intake DNA – deoxyribonucleic acid (3) 469, 53 (9–10) 1185, 133 EPN – entomopathogenic nematodes (5–6) 749, 84 RES – renewable energy sources (7) 923, 103 IAA – indoleacetic acid (7) 967, 108 SBR – sequential biological reactor (7) 895, 100 NAA – neutron activation analysis (1) 27, 3 SRP – soluble reactive phosphorus (2) 201, 22 NPK – nitrogen, phosphorus, potassium (4) 545, 60 PM1.0 – (7) 933, 104 TDR – time domain reflectometry (7) 877, 98 PM2.5 – (7) 933, 104 UV – ultraviolet (3) 469, 53; (5–6) 827, 95 PM10 – (7) 933, 104 VOCs – volatile organic compounds (7) 991, 111

WYKAZ AKRONIMÓW

Sposób zapisu odnoœników hase³ – (nr zeszytu) pierwsza strona artyku³u, nr artyku³u (w spisie treœci rocznika).

DMPP – 3,4-dimetylopyrazolofosfat (1) 97, 12 PM2.5 – (7) 933, 104 DNA – kwas dezoksyrybonukleinowy (3) 469, 53 PM10 – (7) 933, 104 EPN – nicienie entomopatogenne (5–6) 749, 84 SBR – biologiczny reaktor sekwencyjny (7) 895, 100 IAA – kwas indolilo-3-octowy (7) 967, 108 SRP – rozpuszczalny fosfor reaktywny (2) 201, 22 NAA – neutronowa analiza aktywacyjna(1) 27, 3 TDTP – tymczasowe dopuszczalne tygodniowe po- NPK – azot, fosfor, potas (4) 545, 60 brania (9–10) 1185, 133 OZE – odnawialne Ÿród³a energii (7) 923, 103 UV – ultrafiolet (3) 469, 53; (5–6) 827, 95 PM1.0 – (7) 933, 104 VOCs – lotne zwi¹zki organiczne (7) 991, 111

Varia

CENTRAL EUROPEAN CONFERENCE ECOpole ’11

Short Conference Report

The Conference ECOpole ’11 was held in 13–15 X 2011 at the Conference Center “Rzemieslnik” in Zakopane, PL. It was the jubilee – the twentieth ecological conference of the series of meetings organised by the Society of Ecological Chemistry and Engineering, Opole, PL. 178 participants, including delegates representing 10 countries (Bulgaria, Czech Republic, Germany, Latvia, Lithuania, Poland, Romania, Russian Federation, Slovakia and Ukraine) took part in the event and presented 30 oral contributions and 177 posters. The Abstracts of the Conference contributions were available on the Conference website before and during the Conference. The Conference issue of the quarterly Ecological Chemistry and Engineering S 2011, 18(3) containing among others keynote speakers papers, was distributed at the Conference Reception desk together with a PenDrive (containing short info on Keynote Speakers, Abstracts of the Conference presentations as well as ECOpole ’11 Conference Programme). On Wednesday (12th October 2011) at 20.30 the participants were invited for Get-Together Party. The Conference Agenda was divided into 5 sections: – SI Ecological Chemistry and Engineering – SII Environment Friendly Production and Use of Energy – SIII Risk, Crisis and Security Management – SIV Forum of Young Scientists and Environmental Education – SV Health, Ecology and Agriculture. The Conference was opened by prof. Maria Waclawek, Chairperson of the Organising Committee and prof. Witold Waclawek, Chairman of the Scientific Board and President of the Society of Ecological Chemistry and Engineering. Prof. Paul Jozef CRUTZEN (Max-Planck-Institute for Chemistry, Mainz, DE), the Nobel Prize Winner, initiated the First Plenary Session with the invited lecture: “Atmospheric chemistry and climate in the Anthropocene”. This contribution was met with great interest of the audience. During the conference the plenary lectures were also delivered by other invited lecturers: prof. Marina V. FRONTASYEVA (Joint Institute for Nuclear Research, 1820 Varia

Dubna, RU): “NAA for Life Sciences at Frank Laboratory of Neutron Physics, Joint Institute for Nuclear Research in Dubna”, prof. Hartmut FRANK (University of Bayreuth, Bayreuth, DE): “Fresh water pearl mussels, vanishing witnesses of water quality”, prof. Jozef LEHOTAY (Slovak Technical University, Bratislava and University of SS Cyril and Methodius in Trnava, SK): “Trace analysis of some toxic compounds by HPLC” and prof. Bogdan ZYGMUNT (Gdansk University of Technology, Gdañsk, PL): “Increasingly green approaches to determine selected trace organics in complex matrices”. There were also presented very interesting lectures, eg: S. Ledakowicz and L. Bilinska (Lodz University of Technology, £ódŸ, PL): “Application of the Fenton reagent in the textile wastewater treatment under industrial conditions”, A. Nowak (West Pomeranian University of Technology, Szczecin, PL): “Are the bacteria necessary in the environment?”, M. Bratychak (Lviv Polytechnic National University, Lviv, UA): “Utilization ways of by-products of hydrocarbon raw material pyrolysis”, S. Bocian and B. Buszewski (Nicolaus Copernicus University, Toruñ, PL): “The new approach to the retention mechanism in reversed-phase liquid chromatography”, D. Panasiuk, A. Glodek and J.M. Pacyna (NILU Polska Ltd., Norwegian Institute for Air Research Branch Poland, Katowice, PL): “Scenarios of mercury emission to air, water and soil in Poland to year 2020”, G. Boczkaj, M. Jaszczolt and M. Kaminski (Gdansk University of Technology, Gdañsk, PL): “New methods for process control of the thermal cracking rate of vacuum distillates and products from crude oil vacuum distillation”, E. Kraleva, V. Karamfilov and G. Hiebaum (Institute of Biodiversity and Ecosystem Research – BAS, Sofia, BG): “Determination of polycyclic aromatic hydrocarbons in the Black Sea water by GC/MS following preconcentration with solid-phase extraction”, K. Pio- trowska, M. Imbierowicz and A. Chacuk (Lodz University of Technology, £ódŸ, PL): “Wet oxidation of dairy sewage: the kinetic study of intermediate products decomposi- tion”, M. Siemieniec, H. Kierzkowska-Pawlak and A. Chacuk (Lodz University of Technology, £ódŸ, PL): “Reaction kinetics of CO2 in aqueous ethylethanolamine and methyldiethanolamine solutions using the stopped-flow technique”, R. Slavik, M. Juli- nova, and M. Labudikova (Tomas Bata University, Zlin, CR): “Screening of the spatial distribution of risk metals in topsoil from an industrial complex” and A.I. Stoica and A.A. Ciucu (University of Bucharest, Bucharest, RO): “Kinetic approach for heavy metals detection by glucose-oxidase inhibition studies”. Thursday, a day of hard work, was finished with the Poster Session of the Section The Quality of Environment and its Monitoring. Many of the discussions started at the posters, lasted until the evening hours. The next point of Thursday Conference Programme was the 45 minutes-long Musical Soirée. Songs and dances from Rocky Podhale were presented by highlander team “M³ode Klimki”. The audience applauded the players and claimed for encore. At 20.00 the Conference participants were invited for a Conference Dinner. As usually during the ECOpole Conferences, the second day included the Session of the Young Scientists (a forum of young scientists that present and discuss local ecological problems of their countries). During the Young Scientists’, Environmental Education and Renewable Energy Poster Session 57 posters were presented. Varia 1821

The Scientific Board (prof. Stanis³aw Ledakowicz (Lodz University of Technology, £ódŸ, PL), prof. Mikhail Bratychak (Lviv Polytechnic National University, Lviv, UA), prof. Bohumil Vybiral (University of Hradec Kralove, Hradec Králové, CZ) and prof. Witold Waclawek (Society of Ecological Chemistry and Engineering, Opole, PL)) granted awards (sponsored by the Society of Ecological Chemistry and Engineering) for the best presentations. The award for oral presentation was given to Anna Kwiecinska, MSc (Silesian University of Technology, Gliwice, PL) for the oral A. Kwiecinska and K. Konieczny: “Application of membrane processes in treatment of slurry from high-density pig farming”. The awards for poster presentations were given to Lucie Trnkova, MSc (University of Hradec Kralove, Hradec Kralove, CZ) for the poster: L. Trnkova, I. Bousova, F. Altieri and J. Drsata: “Interaction of proteins with low-molecular substances occurring in environment: Structure-activity relationships”, to Agnieszka Baran, MSc (University of Agriculture, Kraków, PL) for the poster: A. Baran and M. Tarnawski: “Content of heavy metals in leachate from bottom sediments as a potential source of contamination of water and ground environment” and to Aleksander Zaremba, MSc (Czestochowa University of Technology, Czêstochowa, PL) for the poster: T. Rodziewicz, J. Nakata, K. Taira, I. Inagawa, A. Zaremba and M. Waclawek: “Performance of flat module made of Sphelar cells in higher latitude areas”. On Saturday morning an excursion was organised to Kuznice. Participants visited Manor-Palace in Kuznice, Memorial Chamber of Count Wladyslaw Zamoyski and an exhibition of works by Antoni Kocjan “Animals of the Tatras”. Closing the conference, prof., prof. Maria and Witold Waclawek made short recapitulation. In general, ECOpole ’11 was focused on monitoring of the quality of natural environment, its effects on human life, environmental education as well as application of renewable sources of energy. They expressed gratitude to all participants for coming and taking active part in the Conference and thanked all Chairpersons of Sessions. They asked the participants to publish electronic version of presented during ECOpole ’11 Conference contributions (oral presentations as well as posters) at the conference website ecopole.uni.opole.pl – some persons have already took advantage of this call. They announced, that full texts of the presented papers will be published (after obtaining reviewers’ positive opinions) in the successive issues of the journals Ecological Chemistry and Engineering A and S and they will be distributed to all participants. The Extended Abstracts of the presentations will be published in two subsequent issues of semi-annual Proceedings of the ECOpole. Thanks to the financial support of the Polish Ministry of Scientific Research and Higher Education, it was possible, among others to publish Abstracts and full Conference contributions on the Conference website. At the end they invited all Colleagues to attend the ECOpole ’12 Conference, which will be held at the Conference Center “Rzemieslnik” in Zakopane, PL in the next October.

Maria Wac³awek 1822 Varia

HONORARY COMMITTEE Prof. dr hab. Barbara KUDRYCKA – Chairperson Ministry of Scientific Research and Higher Education Prof. dr hab. in¿. Andrzej KRASZEWSKI Ministry of the Environment Józef SEBESTA Marshall of the Opole Province Prof. dr hab. Bogus³aw BUSZEWSKI President of the Polish Chemical Society Prof. dr hab. in¿. Krystyna CZAJA Rector of the Opole University

SCIENTIFIC BOARD

Witold WACLAWEK – Opole University, Opole – Chairman Jerzy BARTNICKI – Meteorological Institute – DNMI, Oslo-Blindern, NO Mykhaylo BRATYCHAK – National University of Technology, Lviv, UA Boguslaw BUSZEWSKI – Nicolaus Copernicus University, Toruñ Andrzej GAWDZIK – Opole University, Opole Milan KRAITR – University, Plzeò, CZ Andrzej KULIG – Warsaw University of Technology, Warszawa Bernd MARKERT – International Graduate School [IHI], Zittau, DE Jacek NAMIESNIK – Gdansk University of Technology, Gdañsk Lucjan PAWLOWSKI – Lublin University of Technology, Lublin Vasil SIMEONOV – University of Sofia, Sofia, BG Bohumil VYBIRAL – University of Hradec Kralove, Hradec Králové, CZ Wies³aw WASIAK – Adam Mickiewicz University, Poznañ Roman ZARZYCKI – Technical University of Lodz, £ódŸ

ORGANIZING COMMITTEE

Maria WACLAWEK – Opole University, Opole – Chairperson Zbigniew FIGAS – Provincial Fund of Environmental Protection and Water Management, Opole Agnieszka DOLHANCZUK-SRODKA – Opole University, Opole Andrzej KLOS – Opole University, Opole Karel KOLAR – University of Hradec Kralove, Hradec Králové, CZ Jan KRIZ – University of Hradec Kralove, Hradec Králové, CZ Tadeusz MAJCHERCZYK – Opole University, Opole Malgorzata RAJFUR – Opole University, Opole Aleksander ZAREMBA – Czestochowa University of Technology, Czêstochowa Zbigniew ZIEMBIK – Opole University, Opole

SPONSORS

Ministry of Scientific Research and Higher Education, Warszawa Private Travel Agency SINDBAD Varia 1823

Invited Lectures

Jozef Crutzen – Atmospheric chemistry and climate in the Anthropocene

Marina V. Frontasyeva – NAA for Life Sciences at Frank Laboratory of Neutron Physics, Joint Institute for Nuclear Research in Dubna

Hartmut Frank – Fresh water pearl mussels, vanishing witnesses of water quality

Jozef Lehotay – Trace analysis of some toxic compounds by HPLC

Bogdan Zygmunt – Increasingly green approaches to determine selected trace organics in complex matrices 1824 Varia

Professor Paul Jozef Crutzen, the Nobel Prize Winner, delivering his lecture (photo by Andrzej Nowak)

Professor Marina V. Frontasyeva as a Chairperson (photo by Tomasz Ciesielczuk) Invitation for ECOpole ’12 Conference CHEMICAL SUBSTANCES IN ENVIRONMENT

We have the honour to invite you to take part in the 21st annual Central European Conference ECOpole ’12, which will be held in 11–13 X 2012 (Thursday–Saturday). The Conference Programme includes oral presentations and posters and will be divided into five sections: – SI Chemical Pollution of Natural Environment and Its Monitoring – SII Environment Friendly Production and Use of Energy – SIII Risk, Crisis and Security Management – SIV Forum of Young Scientists and Environmental Education in Chemistry – SV Impact of Environment Pollution on Food and Human Health The Conference language is English. Contributions to the Conference will be published as: – abstracts on the CD-ROM (0.5 page of A4 paper sheet format), – extended Abstracts (4–6 pages) in the semi-annual journal Proceedings of ECOpole, – full papers will be published in successive issues of the Ecological Chemistry and Engineering/Chemia i In¿ynieria Ekologiczna (Ecol. Chem. Eng.) ser. A or S.

Additional information one could find on the Conference website:

ecopole.uni.opole.pl

th The deadline for sending the Abstracts is 15 July 2012 and for the Extended st Abstracts: 1 October 2012. The actualised list (and the Abstracts) of the Conference contributions accepted for presentation by the Scientific Board, one can find (starting st from 31 July 2012) on the Conference website. The papers must be prepared according to the Guide for Authors on Submission of Manuscripts to the Journals. 1826 Varia

At the Reception Desk each participant will obtain abstracts of the Conference contributions as well as the Conference Programme recorded on electronic media (the Programme will be also published on the ECOpole ’12 website).

Further information is available from: Prof. dr hab. Maria Wac³awek Chairperson of the Organising Committee of ECOpole ’12 Conference University of Opole email: [email protected] and [email protected] phone +48 77 455 91 49 and +48 77 401 60 42 fax +48 77 401 60 51

Conference series

1. 1992 Monitoring ’92 Opole 2. 1993 Monitoring ’93 Turawa 3. 1994 Monitoring ’94 Pokrzywna 4. 1995 EKO-Opole ’95 Turawa 5. 1996 EKO-Opole ’96 Kêdzierzyn KoŸle 6. 1997 EKO-Opole ’97 Duszniki Zdrój 7. 1998 CEC ECOpole ’98 Kêdzierzyn-KoŸle 8. 1999 CEC ECOpole ’99 Duszniki Zdrój 9. 2000 CEC ECOpole 2000 Duszniki Zdrój 10. 2001 CEC ECOpole ’01 Duszniki Zdrój 11. 2002 CEC ECOpole ’02 Duszniki Zdrój 12. 2003 CEC ECOpole ’03 Duszniki Zdrój 13. 2004 CEC ECOpole ’04 Duszniki Zdrój 14. 2005 CEC ECOpole ’05 Duszniki Zdrój 15. 2006 CEC ECOpole ’06 Duszniki Zdrój 16. 2007 CEC ECOpole ’07 Duszniki Zdrój 17. 2008 CEC ECOpole ’08 Piechowice 18. 2009 CEC ECOpole ’09 Piechowice 19. 2010 CEC ECOpole ’10 Piechowice 20. 2011 CEC ECOpole ’11 Zakopane Zapraszamy do udzia³u w Œrodkowoeuropejskiej Konferencji ECOpole ’12 w dniach 11–13 X 2012 SUBSTANCJE CHEMICZNE W ŒRODOWISKU PRZYRODNICZYM

Bêdzie to dwudziesta pierwsza z rzêdu konferencja poœwiêcona badaniom podstawo- wym oraz dzia³aniom praktycznym dotycz¹ca ró¿nych aspektów ochrony œrodowiska przyrodniczego. Doroczne konferencje ECOpole maj¹ charakter miêdzynarodowy i za takie s¹ uznane przez Ministerstwo Nauki i Szkolnictwa Wy¿szego. Obrady konferencji ECOpole ’12 bêd¹ zgrupowane w piêciu sekcjach: – SI Chemiczne substancje w œrodowisku przyrodniczym oraz ich monitoring – SII Odnawialne Ÿród³a energii i jej oszczêdne pozyskiwanie oraz u¿ytkowanie – SIII Zarz¹dzanie œrodowiskiem w warunkach kryzysowych – SIV Forum M³odych (FM) i Edukacja proœrodowiskowa – SV Wp³yw zanieczyszczeñ œrodowiska oraz ¿ywnoœci na zdrowie ludzi. Materia³y konferencyjne bêd¹ opublikowane w postaci: – abstraktów (0,5 strony formatu A4) na CD-ROM-ie; – rozszerzonych streszczeñ o objêtoœci 4–6 stron w pó³roczniku Proceedings of ECOpole; – artyku³ów: w abstraktowanych czasopismach: Ecological Chemistry and Engineer- ing/Chemia i In¿ynieria Ekologiczna (Ecol. Chem. Eng.)ser.AiSoraz niektórych w pó³roczniku Chemia – Dydaktyka – Ekologia – Metrologia. Termin nadsy³ania angielskiego i polskiego streszczenia o objêtoœci 0,5–1,0 strony (wersja cyfrowa + wydruk) planowanych wyst¹pieñ up³ywa w dniu 15 lipca 2012 r. Lista prac zakwalifikowanych przez Radê Naukow¹ Konferencji do prezentacji bêdzie sukcesywnie publikowana od 15 lipca 2012 r. na stronie webowej

ecopole.uni.opole.pl 1828 Varia

Aby praca (dotyczy to tak¿e streszczenia, które powinno mieæ tytu³ w jêzyku polskim i angielskim, s³owa kluczowe w obydwu jêzykach) przedstawiona w czasie konferencji mog³a byæ opublikowana, jej tekst winien byæ przygotowany zgodnie z wymagania- mi stawianymi artyku³om drukowanym w czasopismach. Zalecenia te s¹ umieszczone na stronie webowej Towarzystwa.

tchnie.uni.opole.pl

Po konferencji zostan¹ wydane 4–6-stronicowe rozszerzone streszczenia wyst¹pieñ w pó³roczniku Proceedings of ECOpole. Artyku³y te winny byæ przes³ane do 1 paŸdzier- nika 2012 r. Wszystkie nadsy³ane prace podlegaj¹ zwyk³ej procedurze recenzyjnej. Wszystkie streszczenia oraz program Konferencji zostan¹ wydane na CD-ROM-ie, który otrzyma ka¿dy z uczestników podczas rejestracji. Program bêdzie tak¿e umieszczony na stronie webowej Konferencji.

Prof. dr hab. Maria Wac³awek Przewodnicz¹ca Komitetu Organizacyjnego Konferencji ECOpole ’12

Wszelkie uwagi i zapytania mo¿na kierowaæ na adres: [email protected] lub [email protected] tel. 77 401 60 42 i 77 455 91 49 fax 77 401 60 51 ACKNOWLEDGEMENT OF REVIEWERS

PODZIÊKOWANIA DLA RECENZENTÓW

The Editors would like to express their gratitude to the following Reviewers who hel- ped in the peer-review process of the papers considered for publication in the journal Ecological Chemistry and Engineering A:

Editorial Board of Ecological Chemistry and Engineering A Society of Ecological Chemistry and Engineering ul. kard. B. Kominka 6, 45–032 OPOLE Phone +48 77 4 559 149; +48 77 401 60 42; http://tchie.uni.opole.pl/

Jacek ANTONKIEWICZ Florian GAMBUŒ Piotr BANASZUK Andrzej GAWDZIK Jerzy BARTNICKI Maria GAWÊDA Boles³aw BIENIEK Dimitrios A. GEORGAKELLOS El¿bieta BOLIG£OWA Anna GOLCZ Teresa BOWSZYS Krzysztof GONDEK Michael BRATYCHAK Andrzej GÓRNIAK W³odzimierz BREŒ Gyorgy HELTAI Witold BROSTOW Lidmila HYSPLEROVA Zdzis³aw CIEÆKO Ewa JADCZUK-TOBIASZ Jacek CZEKA£A Czes³awa JASIEWICZ Jean Bernard DIATTA Magdalena JAWORSKA Dragan DJORDJEVIC Marek JÓWIAK Agnieszka DO£HAÑCZUK-ŒRÓDKA Adam KACZOR Danuta DOMSKA W³adys³aw KAMIÑSKI Agata FARGASOVA Stanis³aw KALEMBASA Tadeusz FILIPEK Aleksander KIRYLUK Barbara FILIPEK-MAZUR Bo¿ena KIZIEWICZ Stefan FRAENZLE Andrzej K£OS Hartmut FRANK Barbara KO£WZAN Marina V. FRONTASYEVA Miros³aw KONOPIÑSKI Stanis³aw GAJDA Micha³ KOPEÆ 1830 Varia

Jan KOPER Petr SKARPA Jolanta KORZENIOWSKA Jerzy SKRZYPSKI Joanna KOSTECKA Roman SLAVIK Milan KRAITR Lech SMOCZYÑKI Katarina KRALOVA Andrzej SOLECKI Jan KØͦ Ewa STANIS£AWSKA-GLUBIAK Jan KUCHARSKI Eiliv STEINNES Edward KUNICKI ) Anca STOICA Jozef LEHOTAY Jerzy SZWEJDA Andrzej £ACHACZ Piotr TOMASIK Elena MAESTRI Elwira TOMCZAK Elena MASAROVICOVA Jan TYWOÑCZUK Stanis³aw MAZUR Magnuss VIRCAVS Teofil MAZUR Bohumil VYBIRAL Jacek MIÊDZOBRODZKI Maria WAC£AWEK Marcin NIEMIEC Joanna NIEMYSKA-£UKASZUK Witold WAC£AWEK Ma³gorzata RAJFUR Józefa WIATER Leszek ROGALSKI Miros³aw WIATKOWSKI Czes³awa ROSIK-DULEWSKA Barbara WIŒNIOWSKA-KIELIAN Krzysztof J. RUDZIÑSKI Czes³aw WO£OSZYK Manfred SAGER Leszek WONIAK Wiera S¥DEJ Stanis³aw WRÓBEL František ŠERŠEÒ Miros³aw WYSZKOWSKI Jerzy SIEPAK Tomasz ZALESKI Vasil SIMEONOV Roman ZARZYCKI Pavlina SIMEONOVA Volodymyr G. ZINKOVSKYY GUIDE FOR AUTHORS ON SUBMISSION OF MANUSCRIPTS

A digital version of the Manuscript addressed: Professor Witold Wac³awek Editorial Office of monthly Ecological Chemistry and Engineering A (Ecol. Chem. Eng. A) Uniwersytet Opolski ul. kard. B. Kominka 6, 45–032 Opole, Poland Phone: +48 77 401 60 42, fax +48 77 401 60 51, email: [email protected] should be sent by email to the Editorial Office Secretariat – [email protected] The Editor assumes, that an author submitting a paper for publication has been autho- rised to do that. It is understood the paper submitted to be original and unpublished work, and is not being considered for publication by another journal. After printing, the copyright of the paper is transferred to Towarzystwo Chemii i In¿ynierii Ekologicznej (Society for Ecological Chemistry and Engineering). “Ghostwriting” and “guest authorship” are a sign of scientific misconduct. To counteract them, please provide information, for the Editor, on the percentage contribution of individual Authors in the creation of publications (including the information, who is the author of concepts, principles, methods, etc.). Editorial Board believes that the main responsibility for those statements bears the Author sending the manuscript. In preparation of the manuscript please follow the general outline of papers published in the most recent issues of Ecol. Chem. Eng. A, a sample copy can be sent, if requested. Papers submitted are supposed to be written in English language and should include a summary and keywords, if possible also in Polish language. If not then the Polish summary and keywords will be provided by the Editorial Office. All authors are re- quested to inform of their current addresses, phone and fax numbers and their email addresses. It is urged to follow the units recommended by the Systéme Internationale d’Unites (SI). Graph axis labels and table captions must include the quantity units. The use of the following commonly applied expressions is recommended: mass – m/kg, time – t/s or t/min, current intensity – I/A; thermodynamic temperature – T/K, Celsius scale tempera- ture – t/°C or q/°C (if both time and Celsius scale units need to be used, the symbol q/°C for temperature is to be taken) etc. 1832 Varia

Symbols recommended by the International Union of Pure and Applied Chemistry (Pure and Appl. Chem., 1979, 51, 1–41) are to be followed. Graphics (drawings, plots) should also be supplied in the form of digital vector – type files, eg Corel-Draw, Grapher for Windows or at least in a bitmap format (TIF, PCK, BMP). In the case of any query please feel free to contact with the Editorial Office. Footnotes, tables and graphs should be prepared as separate files. References cited chronologically should follow the examples given below: [l] Kowalski J. and Malinowski A.: Polish J. Chem. 1990, 40(3), 2080–2085. [2] Nowak S: Chemia nieorganiczna, WNT, Warszawa 1990. [3] Bruns I., Sutter K., Neumann D. and Krauss G.-J.: Glutathione accumulation – a specific response of mosses to heavy metal stress, [in:] Sulfur Nutrition and Sulfur Assimilation in Higher Plants, P. Haupt (ed.), Bern, Switzerland 2000, 389–391. Journal titles should preferably follow the Chem. Abst. Service recommended abbre- viations. Each publication is evaluated by at least two independent reviewers from outside of the unit. In the case of paper written in a foreign language, at least one of reviewers is affiliated to a foreign institution other than the author’s work. Sometimes so-called “double-blind review process” occurs (the author(s) and reviewers do not know their identities). In other cases Editor must be sure that no conflict of interest (direct personal relationships, professional relationships, or direct scientific cooperation in the past two years) occurs between the reviewer and the author. Reviewer has to fill in the Reviewers report. On its end must be an explicit request to the approval of the article for publication or its rejection. Receipt of a paper submitted for publication will be acknowledged by email. If no acknowledgement has been received, please check it with the Editorial Office by email, fax, letter or phone. ZALECENIA DOTYCZ¥CE PRZYGOTOWANIA MANUSKRYPTÓW

Praca przeznaczona do druku w miesiêczniku Ecological Chemistry and Engineering A/Chemia i In¿ynieria Ekologiczna A powinna byæ przes³ana na adres Redakcji: Profesor Witold Wac³awek Redakcja Ecological Chemistry and Engineering A Uniwersytet Opolski ul. kard. B. Kominka 6, 45–032 Opole tel. 77 401 60 42, fax 77 401 60 51 email: [email protected] w postaci cyfrowej w formacie Microsoft Word (ver. 7.0 dla Windows) emailem ([email protected]). Redakcja przyjmuje, ¿e przesy³aj¹c artyku³ do druku autor w ten sposób oœwiadcza, ¿e jest upowa¿niony do tego oraz zapewnia, i¿ artyku³ ten jest oryginalny i nie by³ wczeœniej drukowany gdzie indziej i nie zosta³ wys³any do druku gdzie indziej oraz ¿e po jego wydrukowaniu copyright do tego artyku³u uzyskuje Towarzystwo Chemii i In- ¿ynierii Ekologicznej. “Ghostwriting” i “guest authorship” s¹ przejawem nierzetelnoœci naukowej. Aby im przeciwdzia³aæ, prosimy o podanie informacji, do wiadomoœci Redakcji, o wk³adzie procentowym poszczególnych autorów w powstanie publikacji (wraz z informacj¹, kto jest autorem koncepcji, za³o¿eñ, metod itp.). Redakcja uwa¿a, ¿e g³ówn¹ odpowiedzial- noœæ za te stwierdzenia ponosi Autor zg³aszaj¹cy manuskrypt. W przygotowaniu manuskryptu nale¿y przede wszystkim wzorowaæ siê na postaci najnowszych artyku³ów opublikowanych w Ecological Chemistry and Engineering A, na przyk³ad zamieszczanych na stronie webowej Towarzystwa: http://tchie.uni.opole.pl/ Prace przesy³ane do publikacji winny byæ napisane w jêzyku angielskim oraz zaopa- trzone w streszczenia oraz s³owa kluczowe w jêzykach angielskim oraz polskim. Zalecamy, a¿eby artyku³ zawiera³ adresy i emaile oraz numery telefonów i faksów wszystkich autorów danej pracy, szczególnie g³ównego autora, którego nazwisko wy- ró¿niamy gwiazdk¹. Usilnie prosimy o stosowanie uk³adu jednostek SI. Zwracamy uwagê, ¿e osie wykre- sów oraz g³ówki tabel powinny bezwzglêdnie zawieraæ jednostki stosownej wielkoœci. Polecamy symbolikê zalecan¹ przez PTChem (Symbole i terminologia wielkoœci i jed- 1834 Varia nostek stosowanych w chemii fizycznej, Ossolineum, Wroc³aw 1989; Pure Appl. Chem. 1979, 51, 1–41). Materia³ graficzny (rysunki, wykresy), obok wersji na papierze, powi- nien równie¿ byæ dostarczony w postaci cyfrowych plików wektorowych, np. za po- moc¹ programu: CorelDraw wersja 3.0–8.0, Grafer dla Windows lub przynajmniej bito- we (TIF, PCX, BMP). W przypadku trudnoœci z wype³nieniem tego warunku Redakcja zapewnia odp³atne wykonanie materia³u graficznego na podstawie dostarczonego szki- cu, bli¿sze informacje mo¿na uzyskaæ telefonicznie 077 401 60 42. Przypisy i tabele podobnie jak rysunki zapisujemy jako osobne pliki. Literaturê prosimy zamieszczaæ wg poni¿szych przyk³adów: [1] Kowalski J. and Malinowski A.: Polish J. Chem. 1990, 40, 2080–2085. [2] Nowak S.: Chemia nieorganiczna, WNT, Warszawa 1990. [3] Bruns I., Sutter K., Neumann D. and Krauss G.-J.: Glutathione accumulation – a specific response of mosses to heavy metal stress, [in:] Sulfur Nutrition and Sulfur Assimilation in Higher Plants, P. Haupt (ed.), Bern, Switzerland 2000, 389–391. Tytu³y czasopism nale¿y skracaæ zgodnie z zasadami przyjêtymi przez amerykañsk¹ Chemical Abstracts Service. Autor mo¿e, je¿eli uwa¿a to za wskazane, podawaæ te¿ tytu³ cytowanych artyku³ów z czasopism, który bêdzie sk³adany kursyw¹ oraz numer zeszytu danego woluminu (w nawiasie, po numerze woluminu). Ka¿da publikacja jest opiniowana przez dwóch niezale¿nych recenzentów spoza jednostki, w której pracuje Autor przesy³aj¹cy artyku³. W przypadku tekstów z za- granicy co najmniej jeden z recenzentów jest afiliowany w instytucji zagranicznej innej ni¿ autor pracy. Niekiedy recenzenci nie znaj¹ nazwisk autorów publikacji (tzw. double-blind review proces). W pozosta³ych przypadkach Redakcja musi mieæ pew- noœæ, ¿e nie wystêpuje konflikt interesów (bezpoœrednie relacje osobiste, relacje podleg³oœci zawodowej, bezpoœrednia wspó³praca naukowa w ci¹gu ostatnich dwóch lat) pomiêdzy recenzentem i autorami. Recenzent wype³nia formularz recenzji. Koñczy siê on jednoznacznym wnioskiem o dopuszczeniu artyku³u do publikacji lub jego odrzuceniu. Redakcja potwierdza emailem otrzymanie artyku³u do druku. W przypadku braku potwierdzenia prosimy o interwencjê: emailem, faksem, listem lub telefonicznie. REDAKTOR TECHNICZNY Halina Szczegot

SK£AD I £AMANIE Jolanta Brodziak

PROJEKT OK£ADKI Marian Wojewoda

Druk: „Drukarnia Smolarski”, Józef Smolarski, 45–326 Opole, ul. Sandomierska 1. Objêtoœæ: ark. wyd. 17,75, ark. druk. 15,25 Nak³ad: 350 egz. + 5 nadb. aut.