Wood Anatomical Features and Chemical Composition of Prosopis Kuntzei from the Paraguayan Chaco

IAWA Journal, Vol. 31 (1), 2010: 39–52

Wood Anatomical features and chemical composition of Prosopis kuntzei from the paraguayan chaco

Gunthard Scholz1, Elisabeth Windeisen2, Falk Liebner3, Ernst Bäucker4 and Claus-Thomas Bues4

SUMMARY Anatomical features for Prosopis kuntzei Harms were studied by light and scanning electron microscopy. The wood is mainly diffuse-porous with indistinct growth ring boundaries. Vessel diameter ranges between 11 to 193 µm. The thick-walled fibres average 1275µ m in length. Paren- chyma bands are 66 to 1066 µm apart. Heartwood extractives were studied in the vessels, rays and part of the fibres by means of scanning UV microspectrophotometry. The pyrolitic lignin content is 30.7%. The percentage of polyphenolic compounds, such as flavonoids, hydrolysable tannins and proanthocyanidins, is comparatively high at 5.8%. Total extract contents were determined after organic solvent extractions (23.2%) and water extractions (24.9%). The FTIR spectroscopy showed nearly identical spectra for the methanol and water extracts, with characteristic absorption bands for aromatic substances at 1615 and 1520 cm-1. The spectrum of the acetone extract differs only due to an additional but distinct absorption in the carbonyl range at 1695 cm-1. GC/MS analyses revealed that in the acetone and methanol extracts, tetrahydroxy-flavan- 3-ols (isomers of catechin and epicatechin) were the main constituents with a ratio of 25.3 and 27.6%. Key words: Prosopis kuntzei, wood anatomy, chemical composition.

INTRODUCTION The genus Prosopis is one of 78 genera belonging to the subfamiliy Mimosoideae (Evans et al. 2006) and comprises approximately 50 species growing worldwide in tropical and subtropical habitats (Begemann 1966). Prosopis kuntzei Harms occurs in the Chaco region covering parts of Paraguay, Bolivia and Argentina (Dimitri et al. 2000; López 2002). The tree species grows solitarily or in groups (Giménez & Moglia 2003) and is able to colonize abandoned pastures (Dimitri et al. 2000). This is important because the Chaco region – including vast parts of Paraguay, Argentina and Bolivia – is a fragile ecosystem, distinguished by prolongued drought periods and latent water

1) Department of Wood Biology and Wood Technology, Georg-August-Universität Göttingen, Büsgen- weg 4, 37077 Göttingen, Germany. — Corresponding author [E-mail: [email protected]]. 2) Holzforschung München, Technische Universität München, Winzererstraße 45, 80797 München, Germany. 3) Department of Organic Chemistry, University of Natural Resources and Applied Life Sciences, Wien, Austria. 4) Institute for Forest Utilization and Forest Technology, Dresden University of Technology, Pienner Straße 19, 01735 Tharandt, Germany.

Downloaded from Brill.com09/24/2021 02:38:59PM via free access 40 IAWA Journal, Vol. 31 (1), 2010 shortage. Intensive land use is characterized by slash and burn, exhaustive cultivation of valuable tree species and adjacent pasture farming. In the Central Paraguayan Chaco alone, 400,000 ha of woodland was legally cleared between the years 2000 and 2003 (Anonymus 2003). The absence of xerophytic forests induces several environmental problems like local salting or devastation of soils. The brownish-violet to dark-blue coloured heartwood possesses good technological properties and is known for its ex- traordinary durability. The tree forms nutritious pods in the dry season which could be used to feed cattle. The tannin content of the pods amounts to approx. 26.6% (Giménez & Moglia 2003) and the protein content to approx. 9% (Scholz et al. 2005). Bees use the tree blossoms to forage for food, making the presence of Prosopis kuntzei important for beekeeping and seed-set in agricultural crops. In literature, the following applica- tions are cited: jewellery, posts (Dimitri et al. 2000), indigenous weapons, charcoal (López 2002), spokes, tool shafts, walking sticks and terrace floors (Giménez & Moglia 2003). According to Scholz et al. (2005), the heartwood is comparable to Bongossi (Lophira alata), especially due to the very high surface hardness. It is suited for ex- terior constructions (e.g., fences, decorative palisades). Furthermore, the timber can be used for tools, musical instruments, sleepers etc. Forestry legislation is insufficient to avoid over-exploitation in these tropical countries. It seems necessary to integrate wood utilization into the agricultural land use. However, for commercial use, extensive information on chemical, structural and physical properties of the wood species must be available. These are the objectives of the present investigation which are, unlike previous literature data, considered for Paraguayan provenances.

MATERIALS AND METHODS Material Eight trees from three distinct sites of the Central Paraguayan Chaco were harvested to obtain the raw material. The investigation sites are located 45 km (North-East), 90 km (North-West) and 55 km (South) from the Mennonite settlement Loma Plata (22° 22' 46.55" S, 59° 49' 59.41" W). The tree heights varied between 6–7 m with an average diameter at breast height (dbh) of 29 cm. It was not possible to determine the tree age from tree ring or other analyses. The analysed material was extracted at about 50 cm from the dbh in both directions.

Microscopy The brief description of the wood anatomy is based on the definitions of the IAWA List of Microscopic Features for Hardwood Identification (1993), translated and commented on by Richter and Trockenbrodt (1999). Tangential, radial and transverse microscope slides (22–40 µm thick, cut with a sliding microtome), macerated cells and SEM samples (sputter-coated with carbon and gold) of sap- and heartwood were used for anatomical study. Histometric measurements were done from digital images using the software MicroImage (Softwarebürp Weirauch). In addition to anatomical investigations, the chemical composition of crystals was determined by means of SEM energy dispersive X-ray microanalysis (EDX-Röntec M5). Specimens of all eight trees were investigated.

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Topochemical analyses To identify the lignin distribution and other phenolic compounds within the cell walls of Prosopis kuntzei, a heartwood sample (1 × 1 × 5 mm) was transferred to quartz microscope slides for cellular UV microspectrophotometry (Spurr 1969; Koch & Kleist 2001). The sample was analyzed using a Zeiss UMSP 80 microspectrophotometer equipped with a scanning stage which enables the determination of image profiles at constant wavelengths of, e.g., 278 mm. The used scanning software APAMOS® was developed by Zeiss. This program digitizes rectangular fields of the tissue with a local geometrical resolution of 0.25 × 0.25 µm and a photometrical resolution of 4096 grey-scale levels, which are converted into 14 basic colours to visualize the absorbance intensities. In addition, thin sections were also studied by means of point measurements with a spot size of 1 µm2 between 240 and 560 nm wavelengths in order to characterize the lignin and accessory compounds of phenolic character (Koch & Grünwald 2004). The LAMWIN® computer program was used (ZEISS). One sample was used for the analysis.

Wood chemistry Analyses to determine the pH value and ash content (T 211 om-93, 1996) were carried out according to TAPPI (Technical Association of Pulp and Paper Industry) standards. The chemical constitution of Prosopis kuntzei wood samples was studied by means of Curiepoint pyrolysis GC/MS (Roschy et al. 2002; Scholz et al. 2007) using a CPP-40 pyrolyzing unit (FISCHER/GSG) coupled with a GC 6890/MSD 5973 benchtop system (Agilent Technologies). Prior to the analysis, all materials were vacuum-dried and finely ground. Approx. 200µ g of each sample were placed in small FecralloyTM (Goodfellow Ltd.) tubes and subsequently pyrolyzed at 600 °C for 10 seconds The mixture of volatile pyrolysis products was carried into the inlet of the gas chromatograph by helium (250 °C, split 1 : 20). Separation of the various gas components was achieved using a fused silica column (Optima-5, 30 m, 0.25 mm, 25 µm) with a column flow of 0.9 ml/min, an oven programme starting at 50 °C (5 min) fol- lowed by 5 °C/min to 280 °C (2 min), and an auxiliary temperature of 250 °C. The mass spectrometer was operated in EI mode at 70 eV, 230 °C, and 1.5·10-5 Torr. The total ion chromatograms (programs) were evaluated by using the mass spectra library NIST 2002 (National Institute of Standards and Technology, USA). The pyrolytic lignin and carbohydrate contents of Prosopis kuntzei were calculated from the total peak area of the obtained pyrograms and the total peak area of lignin (LPP) and carbohydrate pyrolysis products (CPP). Sample material of all eight trees was analysed.

Quantitative and qualitative determination of the heartwood extractives The heartwood extractives were successively extracted with organic solvents (petroleum ether, acetone and methanol) using a Soxhlet apparatus according to T 204 om-88 (1996). Further successive extraction was realized with cold and hot water according to T 207 om-93 (1996). Exudates, resulting from bleeding of sap- and heartwood (comparable with resin exudates) were collected in the field as nuggets of up to a few grams, and analysed. Exudate was not present in all the trees. FTIR spectroscopy was applied

Downloaded from Brill.com09/24/2021 02:38:59PM via free access 42 IAWA Journal, Vol. 31 (1), 2010 for the characterisation of extracts and the resin-like exudates. For this purpose, the samples were embedded in KBr pellets (1 mg/300 mg). The FTIR-analyses were carried out on a FTS-40 (Biorad) with a resolution of 4 cm-1 (16 scans). The spectra were baseline corrected and normalized at the highest band. GC and GC/MS analyses were carried out after silylation (heating 1 mg sample in 500 µl solvent and 500 µl silylation reagent (Fluka 15239-1) for 1 h at 80 °C), separation and identification were performed on a GC 2010 with FID (Shimadzu) and/or a GC/MS QP 5000 (Shimadzu). Tempera- tures Inj.: 320 °C; Det.: 330 °C (GC/MS: 250 °C); program T1 = 130 °C/2 min, R1 = 10 °C/min, T2 = 320 °C/min. Column SGE BP5, 15 m (GC/MS: BPX5, 30 m, 0.25 µm film, 0.25 mm ID). The sample material was derived from a mixture of three trees from each of the three sites.

RESULTS AND DISCUSSION Anatomical features Important anatomical features are illustrated in Figure 1 and Table 1. In general, the observations and measurements of anatomical features are in accordance with Tortorelli (1956), Giménez et al. (1997) and Evans et al. (2006). The wood of Prosopis kuntzei

Figure 1. Selected anatomical features of Prosopis kuntzei. – a: Some vessels contain brownish organic deposits. They are arranged in multiple radial rows and clusters. – b: The cross section show crystals inside the marginal parenchyma bands and the axial parenchyma. On the left image border the exceptionally thick fibre cell walls are visible. – c: Vestured intervessel pits (tangential section). – d: Axial parenchyma in strands with one crystal per chamber (radial section).

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Table 1. Comparative overview of selected anatomical features of Prosopis kuntzei. — 18: arrangement of vessels – 26: tangential vessel diameter [µm] – 28: vessels/mm² – 30: vessel element length [µm] – 56: fibre length [µm] – 66, 71, 73, 75: presence and type of axial parenchyma – 82: ray width in cells – 88: ray composition.

Features This study Evans et al. 2006 Giménez et al. 1997 Tortorelli 1956 18 diffuse-porous diffuse-porous semi-ring-porous diffuse-porous 26 (11–)63(–193) 130 (72-)98(-106) (30-)60(-100) 28 5–18 35 10 – 20 20 – 70 30 (82–)200(–322) — 114 – 190 — 56 (557–)1257(–1775) — — 800 66, 71, marginal banded, paratracheal, vasicentric, marginal banded marginal banded, 73, 75 vasicentric, reticulate, confluent, reticulate, in paratracheal, vasi- vasicentric, con- in strands strands centric, confluent, fluent, aliform aliform 82 (1–)4(–6) 3–4 (2–)4(–5) (1–)3(–4) 88 homocellular homocellular — homocellular is predominantly diffuse-porous, but sometimes also semi-ring-porous (Fig. 1a). Some trees have interlocked grain with tangential rotation angles of up to 82°. The observed vessel size and vessel frequency seem lower than the cited Argentinian provenances (see Tortorelli 1956; Giménez et al. 1997; Evans et al. 2006). The average tangential vessel diameter is about 63 µm. Some solitary vessels reach a diameter of up to 193 µm. The smaller vessels appeared in grouped clusters, in line with Evans et al. (2006). In accordance with Tortorelli (1956), the pits are vestured (Fig. 1c). The vessel pit size ranges between 7–11 µm which is higher than reported by Evans et al. (2006) who reported pit sizes between 4–6 µm. The net vessel area percentage reported by Giménez et al. (1997) ranged between 3.7 and 9.5%. Our values range between 6 and 9%. The partly very low vessel area ratio in some xylem zones can be explained by the semi-ring-porous vessel arrangement found by Giménez et al. (1997). The fibres have very thick walls with a Runkel ratio of approx. 10.2 (Fig. 1b) which is above that of some of the heaviest timbers in the world, like Lophira alata, having a Runkel rate of 6.0 (Wagenführ & Scheiber 1989) or even more. The average fibre length is 1257 µm in comparison to 800 µm measured by Tortorelli (1956). The marginal parenchyma (Fig. 1a & 1b) is mostly in bands of two cells wide. The radial distance between these bands varies between (66–)402(–1066) µm. Crystals are present in chambered parenchyma cells (Fig. 1d). Evans et al. (2006) reported axial parenchyma in strands of 2–4 cells. By using EDXA on ashed samples, exclusively calcium was detected. The results are likely to indicate calcium oxalate crystals which were also reported by Tortorelli (1956) and Giménez et al. (1997), and crystals in the fibres and axial parenchyma tissue (Evans et al. 2006). The rays are homocellular and (1–)3(–6) cells wide. The ray height ranges between 130 and 380 µm with an average value of 244 µm. Their frequency with (4–)6(–8)/mm is comparable to 5/mm found by Evans et al. (2006).

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Figure 2. UV microscopic scanning profile of the fibres of Prosopis kuntzei heartwood (scanning field 48.5 × 52.25 μm²).

Figure 3. UV microscopic scanning profile of a vessel and adjacent axial parenchyma tissue of Prosopis kuntzei heartwood (scanning field 43.5× 48.5 μm²).

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Topochemical investigations Figures 2 and 3 illustrate the representative UV scanning profiles of fibres, vessels and axial parenchyma. The maximum absorbance was detected within a range of 0.67–0.74 inside the cell corners (CC) of the parenchyma cell walls. The absorbance of fibres tends to show lower values between 0.41–0.48. The thick S2 layers of the fibres are characterized by relatively uniform UV absorbance at 278 nm in the range of 0.16–0.22. The compound middle lamella (CML) and cell corners (CC) of the individual fibres can be distinguished by significantly higher UV absorbances in the range of 0.30 (CML) and 0.35–0.48 (CC). The different absorbance values strongly correlate with various lignin contents in the individual cell wall layers. In some fibre lumina, the presence of accessory compounds can be observed (Fig. 2).

0.9 Fibre-S2 0.8 Fibre-CC

0.7 Ray-Par 0.6 Vessel-S2 0.5 0.4 0.3 Absorbance 0.2 0.1 0 -0.1 240 260 280 300 320 340 360 380 400 420 440 460 480 500 Wavelength (nm) Figure 4. Point measurements of distinct cell wall layers, e.g., fibres, parenchyma cells or vessels (S2 = secondary cell wall layer; CC = cell corner). At the range of around 320 nm, slight shoulders are visible for the fibre cell corners and parenchyma cells.

The S2 layer of the vessels is characterized by UV absorbance values in the range of 0.29–0.41 representing higher lignification than in the cell wall layers of fibres and parenchyma cells. In local parts of the vessel cell wall, directly associated with the CML adjacent to the parenchyma cell, spots with very high absorbance values can be detected. These spots with UV absorbance values up to 0.67–0.74 represent the deposition of highly condensed phenolic compounds. According to Fergus and Goring (1970), the maximum absorbance of point measurements at 278 nm is indicated by the presence of highly absorbing guaiacyl and syringyl lignins. At a wavelength of 320 nm, slight shoulders were detected in the cell corners, partly in the secondary vessel walls and the impregnated ray cell walls (Fig. 4). According to Feist and Hon (1990), the detected shoulders at 320 nm suggest the existence of chromophoric groups such as conjugated double bonds (Fig. 4). In general, small amounts of phenolic compounds like tannin or flavonoids could be detected.

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Table 2. Products obtained from Curie-point pyrolysis of Prosopis kuntzei.

peak r.t . Compound peak r.t. Compound no. (min) no. (min)

1 1.79 Carbon dioxide 32 18.08 1,4:3,6-Dihydro-α-D-glucopyranose 2 2.16 Methyl formiate 33 18.21 4-Vinylphenol 3 2.28 Methyl furane 34 18.64 5-Hydroxymethyl-furan-2-carboxaldehyde 4 2.37 Acetic acid 35 19.44 Methoxyhydroquinone 5 2.60 1-Hydroxypropan-2-one 36 19.94 4-Ethylguaiacol 6 2.73 Hydroxyacetaldehyde 37 20.32 3-Methylcatechol 7 3.00 Dimethylfurane 38 20.94 4-Vinylguaiacol 8 3.83 1-Hydroxybutan-2-one 39 21.97 Syringol 9 3.88 Ethandiol monoacetat 40 22.17 4-Hydroxyveratrol 10 4.08 Succinic aldehyde 41 22.35 4-Propylguaiacol 11 4.27 Methylpyruvate 42 23.08 Eugenol 12 5.45 Furfural 43 23.20 Vanillin 13 5.65 Cyclopenten-1,2-dione 44 23.41 cis-Isoeugenol 14 6.18 Furanmethanol 45 24.44 4-Methylsyringol and Vanillic acid 15 6.62 Ethandiol diacetat 46 24.51 trans-Isoeugenol 16 8.13 2(5H)-Furanone 47 24.75 4-Propioguaiacon 17 8.55 Cyclopentan-1,2-dione 48 25.16 Benzofurane 18 8.64 2-Hydroxy-cyclopent-2-en-1-one 49 25.41 4-Acetylguaiacol 19 9.87 5-Methyl-2-carboxaldehyde 50 26.38 4-Ethylsyringol 20 11.26 2,2-Diethyl-3-methyl-oxazolidine 51 26.50 Guaiacylacetone 21 12.17 3-Methylcyclopentan-1,2-dione 52 27.31 4-Vinylsyringol 22 12.66 Methylfuranone 53 28.17 4-(Prop-2-en-1-yl)-syringol 23 13.80 p-Cresol 54 28.33 4-Propylsyringol 24 13.97 Methylfuroate 55 29.28 4-(cis-prop-1-en-1-yl)syringol 25 14.07 Dimethylhydroxyfuranone 56 29.86 Coumarin derivative 26 14.20 Guaiacol 57 30.01 Coumarin derivative 27 14.94 Hydroxybenzoquinone 58 30.38 4-(trans-prop-2-en-1-yl)syringol 28 16.98 2-Methylguaiacol 59 31.08 4-Acetylsyringol 29 17.38 Dihydroxybenzoquinone 60 31.89 Syringylacetone 30 17.44 4-Methylguaiacol 61 33.02 Propiosyringone 31 17.78 Catechol

Chemical properties The average pH value of Prosopis kuntzei heartwood is 4.5. The ash content is 1.76%, based on the dry weight. Figure 5 shows the pyrogram of a Prosopis kuntzei heartwood sample and a pie diagram depicting the relative constitution of the lignin fraction in

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H units 7x106 3.0 %

S units 6x106 52.8 % G units 5x106 44.2 %

4x106

3x106 Absorbance 2x106

1x106

0 5 10 15 20 25 30 Retention time (min) Figure 5. Curie-point pyrogram of a Paraguayan provenance of Prosopis kuntzei. terms of 4-hydroxyphenyl (H units), 4-hydroxy-3-methoxy (guaiacyl = G units), and 4-hydroxy-3,5-dimethoxy (syringyl = S units) derivatives. As the assignment of almost all lignin pyrolysis products in the obtained pyrograms is comparatively reliable, the error range of the calculated percentages of H, G, and S units should be rather low. The lignin structure of Prosopis kuntzei heartwood was found to be composed of approx. 3.0% p-hydroxyphenyl, 44.2% guaiacyl and 52.8% syringyl units. The total pyrolytic lignin content of the examined wood sample was 30.7%. This value is in good agreement with the lignin content reported for other Prosopis species such as Prosopis laevigata (29.8–31.4%, Carrillo et al. 2008) or Prosopis spp. (31%, Rajput & Tewari 1986). In general, lignin contents ranging from 20 to 40% are typical for tropical hardwood species (Harzmann 1988). In addition to marginal contents of nitrogeneous compounds, which most likely originate from protein pyrolysis, there is some evidence from the comparatively high content of catechol and dihydroxyphenyl derivatives (cf. Table 2, compounds 27, 29, 31, 37) that polyphenolic compounds such as hydrolyzable tannins, proanthocyanidins (condensed tannins) or flavonoids seem also to be present in considerable amounts. The latter are typically deposited in the parenchyma tissues and adjacent fibre lumina and contribute to about 5.8% of the total peak area of the pyrogram. Tortorelli (1956) mentioned the presence of tannin in the heartwood, which is now confirmed. Compared to the calculation of the lignin content and its relative composition of H, G, and S units, quantitative evaluation of the carbohydrate fraction is rather dif- ficult as a certain percentage of peaks (13.8% of the cumulated peak area within the retention time range of 3.0 and 33.0 min) could not unambiguously be assigned to a particular chemical structure. However, as the calculation of the lignin content is hardly impaired by non-assignable peaks originating from lignin pyrolysis, and assuming that the vacuum-dried wood samples only contained a very small amount of polar or higher molecular extractives, the carbohydrate content can be roughly calculated

Downloaded from Brill.com09/24/2021 02:38:59PM via free access 48 IAWA Journal, Vol. 31 (1), 2010 from the total peak area and the peak area of the lignin pyrolysis products, catechol and dihydroxyphenyl derivatives. Hence, a carbohydrate content of approx. 63.5% was calculated which is in good accordance with the values published by Carillo et al. (2008) for Prosopis laevigata (61.7 to 64.5%, outer to inner heartwood). The contents of successively extractable organic compounds show high values. A content of 0.2% was extracted for petroleum ether, 12.1% for acetone and 10.9% for methanol fractions. The value for the acetone fraction is higher than the 11.7% of P. africana (Gérardin et al. 2004) and similar to the 11.6–12.8% of P. laevigata (Carrillo et al. 2008). Furthermore, approx. 15.8% (cold fraction) and 9.1% (hot fraction) are soluble in water. These values approximate the 27% water extract content of P. africana (Gérardin et al. 2004).

Methanol Cold water Hot water 1284 1.0 1199 1614 1053 1517 1352 1473 1087 1448 1112 1159

0.5 997 790 Absorbance 979

2000 1500 1000 Wavenumber (cm-1) Figure 6a. Comparison of the FTIR-Spectra of methanol, cold and hot water extract.

Sapwood 1053 Heartwood 1025 1.0 1155 1613 1419 1445 1286 1365 1202

0.5 1519 Absorbance 901 787 835 0

2000 1500 1000 Wavenumber (cm-1) Figure 6b. Comparison of the FTIR-Spectra of sapwood and heartwood exudates.

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Investigations of the extracts by means of FTIR spectroscopy showed nearly identical FTIR-spectra in comparison to acetone, methanol, cold and hot water extracts (Fig. 6a). Characteristic absorptions of tannins or their building blocks, e.g., aromatic absorptions at about 1615, 1520 and 790 cm-1 (Foo 1981; Streit 1993), can be seen. The aromatic band at about 1520 cm-1 can be observed in the spectra of flavanols with two hydroxyl groups in the B ring (Fig. 7) and procyanidins. Furthermore, it is notice- able that only the acetone extract showed an additionally new absorption band at 1695 3' 4' 8 B O 5' 7 A C

OH 5 4 Figure 7. Flavan-3-ol with numbering of the position of hydroxyl groups. cm-1, which is characteristic for the carbonyl (C= O) vibration, e.g., in aryl ketones, conjugated aldehydes and carboxylic acids or quinones. Hydrolysable tannins of gallic acid esters would have an absorption at about 1715 cm-1. Therefore, the spectra can be better assigned to condensed tannins. FTIR spectra (Fig. 6b) of the yellowish (sapwood) and dark violet (heartwood) exudates show absorption bands of C-H, C-O and C-O-C, e.g., at about 900, 1025, 1050 cm-1 and 1155 cm-1, which, in wood extracts, are normally related to polysaccharides. The absorption band at 1613 cm-1 could be due to carboxyl groups, which are probably related to these polysaccharides. Differences between sapwood and heartwood are seen in several absorption bands, e.g., at about 1520, 1365 and 1285 cm1, which can be assigned to an aromatic skeletal vibration and a corresponding C-O-CH3 vibration. Moreover, the lower resolution of the FTIR spectrum of the sapwood exudates sug- gests that it contains higher polymeric constituents in comparison to the heartwood exudates. Such exudates can be observed after cutting or damaging the tree (q.v. Giménez & Moglia 2003). GC/MS of the petroleum ether extract has shown a few fatty acids (e.g., hexa- decanoic acid, octadecanoic acid, tetracosanoic acid and hexacosanoic acid) and as main constituents several steroids (sitosterol, stigmasterol) and two triterpenes with the formula C30H50O, which could not be clearly identified. Due to the low amount of the petroleum ether extract no further quantification by means of GC has been carried out. The GC and GC/MS analyses of the organic solvent extracts of acetone (Fig. 8a) and methanol (Fig. 8b) showed more or less only two main compounds, which are isomeric to catechin and epicatechin (5, 7, 3’, 4’-tetrahydroxyflavan-3-ol). By means of the mass spectrum only, it was not possible to absolutely clarify whether the unknown substance is mollisacacidin (7, 3’, 4’-trihydroxyflavan-3,4-diol) or 3’, 4’, 7, 8-tetrahydroxyflavan-3-ol, which has already been identified inProsopis glandulosa by Jacobs et al. (1983). In addition, small amounts of epicatechin, catechin and taxifolin could be identified. Two substances with retention times of 24.0 and 25.0 min in the

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uV(x 10,000) 10.0 Chromatogram 9.0

8.0

7.0 IS 6.0 Unknown flavanol 5.0

4.0

3.0

2.0

1.0

0.0

-1.0 12.0 13.0 14.0 15.0 16.0 17.0 18.0 19.0 20.0 21.0 22.0 23.0 24.0 min Figure 8a. GC-spectrum of acetone extract with internal standard (IS).

uV(x 10,000) 10.0 Chromatogram 9.0

8.0 IS 7.0 Unknown flavanol 6.0

5.0

4.0

3.0

2.0

1.0

0.0

-1.0 12.0 13.0 14.0 15.0 16.0 17.0 18.0 19.0 20.0 21.0 22.0 23.0 24.0 min

Figure 8b. GC-spectrum of methanol extract with internal standard (IS).

GC could not be identified by means of GC/MS due to the use of a longer column. The exudates of the heartwood show the same substances but with a lower amount. In the exudates of the sapwood no substances were detected by means of GC or GC/ MS. Table 3 shows the determined amounts of the main constituents of the respective

Table 3. Extractives quantitatively determined by means of GC.

Unknown flavanol Unknown flavanol Epicatechin Catechin Taxifolin (%) (%) (%) (%) (%)

Acetone extract 16.60 8.71 0.44 0.83 0.50 Methanol extract 18.63 8.97 0.50 1.39 0.46 Heartwood exudates 6.42 3.09 1.21 1.39 —

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CONCLUSIONS Generally, the observed anatomical features are mostly similar to Paraguayan and Ar- gentinian provenances and to those of other species of the genus Prosopis, e.g., P. spicigera (Iqbal & Ghouse 1983; Rajput et al. 2008); P. flexuosa (Villalba 1985) or P. laevigata (Carrillo et al. 2008) and 7 other species (Evans et al. 2006). Tortorelli (1956) and Dimitri et al. (2000) mention a high natural durability of Prosopis kuntzei heartwood. This could be explained by some of the detected wood components. Flavonoid-like compounds occur partly in the secondary vessel cell walls, the parenchyma cells and some fibre lumina within the heartwood. The total amount of 5.8% tannins and flavonoids contains tetrahydroxy-flavan-3-ols, e.g., epicatechin, catechin and taxifolin. Furthermore, higher amounts of two unknown flavanols were detected, which have approx. 6% mass ratio in the heartwood. The exudates of the sap- and heartwood mainly contain polysaccharides; the heartwood exudate also includes catechin and epicatechin. The results indicate a high natural durability. However, these assumptions are only hypothetical and require further experimental research.

ACKNOWLEDGEMENTS

We gratefully acknowledge Dr. Gerald Koch (Federal Research Institute for Rural Areas, Forestry and Fisheries – Johann Heinrich von Thünen Institute, Germany) for his support by using UMSP equip- ment and for valuable comments by evaluating the results.

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