Phenotypic Variation Among Mentha Spp

Home , Mentha aquatica, Mentha arvensis, Mentha longifolia, Mentha suaveolens

Pakistan J. Agric. Res. Vol. 25 No. 1, 2012

PHENOTYPIC VARIATION AMONG MENTHA SPP.

Shazia Erum*, Muhammad Naeemullah and Shahid Masood**

ABSTRACT:-The aim of the present study was to investigate the morphological diversity and relationship within 17 genotypes of mentha species. Different species of Mentha were collected from different ecological zones of the world. Great variation was observed in stem length with the coefficient of variation of 198.7% and standard deviation of 14.1. Among the 17 mint collection, nine special aromas was smelled including mint gum, strong camphoraceous, mint like, spicy, lemon, pungent, musky and acrid due to the presence of diverse bioactive compounds and essential oils. The present study may help to patronize the essential oil industry on latest scientific techniques. Like other herbs Mentha have diverse aromatic properties that can be explored for utilization in the industry and this information will be helpful for exploitation and development of natural agricultural resources on scientific grounds.

Key Words: Morphology; Mentha; Diversity; Aroma; Pakistan. INTRODUCTION years. Furthermore, the hybridization, that occurs frequently when the Mints are perennial aromatic species of Mentha are in contact, herbs that are used both for contributes to the complex variation medicinal and aromatic purposes. patterns characterizing most wild Mentha is distributed worldwide and populations.Linneaus (1767) is native from north temperate described the species of the genus regions and occur in all five based on inflorescence morphology. continents. Genus Mentha, a Menthol is the main commercial member of the family Lamiaceae and product obtained from mint. It is the tribe Mentheae, is divided into produced in specialized glands five sections and consists of present in the leaves and flowers of approximately 25 species (Harley and the plant. Moreover, Mentha Brighton, 1977). Within Mentha, it produces secondary metabolites has been suggested that the five basic such as alkaloids, flavanoids, species Mentha arvensis L., Mentha phenols, gummy polysaccharides, aquatica L., Mentha spicata L., terpenes and quinines that are used Mentha longifolia (L.) Huds, and in food, pharmaceutical, cosmetics Mentha suaveolens exist. Most and pesticide industries (Khanuja et Mentha species are characterized by al., 2000). Mint oils are mainly a great morphological variation produced in Argentina, Angola, which is reflected on different Australia, Brazil, Bulgaria, China, taxonomic rank names attributed to Czechoslovakia, France, Hungary, mint plants during the past 200 India, Italy, Paraguay, Switzerland,

* International Islamic University, Islamabad, Pakistan. **Institute of Agricultural Biotechnology and Genetic Resource, National Agricultural Research Centre, Islamabad, Pakistan. Corresponding author: [email protected]

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Thailand and USA. In India, mint numbers (24-120) and much of the species are mainly cultivated in Uttar taxonomy of Mentha has been Pradesh, Haryana and Punjab over complicated by hybridization, by a 162800ha of agricultural land. The high morphological polymorphism, annual world production of Mentha as well as polyploidy and vegetative arvensis L. and M. piperita L. oils are propagation. The best known hybrids 22,000mt and 7500 mt, while India is are M. piperita (peppermint) and M. producing 16000mt and 100 mt per spicata L. (native spearmint), which year, respectively (Khanuja, et al., are intensively cultivated for their 2000; Patra et al., 2002). Brazil was essential oils. Mentha piperita results once an important world producer from a cross between M. aquatica and and exporter of mint essential oils M. spicata. Later being the hybrid (Clark, 1998). between M. suaveolens and M. Several features have been longifolia (Harley and Brighton, 1977). used in the past to examine the It was noted that mints are diversity of Mentha using mainly used to cure gastrointestinal morphological (Malinvaud, 1880), disorders, but the spectrum of cytological (Ruttle, 1931; Heimans, medical activities is broader (McKay 1938; Morton, 1956; Sharma and and Blumberg, 2006). Currently, Bhattacharyya, 1959; Harley, 1967; emphasis is, on conserving plant 1972; Harley and Brighton, 1977; germplasm as valuable bio-resources Singh and Sharma, 1986) and because obscure genes from these chemical (Lawrence, 1978) markers. plants may provide solutions to new Cluster analysis has been diseases, insects, environmental or used to assess similarities among crop production problems (Khanuja landraces in plant breeding et al., 2000; Barbara, 1999). programmes where genotypic and Selection between and within phenotypic repetitions of several accessions for a high level of essential characters were found among oil content, herbage yield and other populations, lines, or accessions characters require an effective tool to from which parents were selected for be employed by mint breeders hybridization (Wilson et al., 1990; (Mirzaie-Nodoushan et al., 2000). Zeinali et al., 2004). Mentha must be considered an Harley and Brighton (1977) important species for the conserva- published a critical review of the tion of genetic resources of medicinal chromosome numbers in relation to and aromatic plants. The collection the taxonomy of the genus. Natural and introduction of new genetic interspecific hybridization occurs materials of Mentha is fundamental with high frequency in Mentha, both to obtain genetic pool adequate for in wild and cultivated populations. further research in plant breeding for Most hybrids are sterile or subfertile, the selection of genotypes with but vegetative propagation enables superior quality. The objective of this them to persist. Complex hybrid work was to evaluate the populations may arise, and if they are morphological differences and subfertile, may cross with parental or relatedness among the mint, nonparental species. This leads to a collected from various ecological large diversity of chromosome zones of the world.

56 PHENOTYPIC VARIATION AMONG MENTHA SPP

MATERIALS AND METHOD

Plant Material In the present study, 17 genotypes collected by the scientists at Plant Genetics Research Institute, National Agricultural Research Centre, Islamabad were evaluated for Figure 1. Morphological variation various morphological and agrono- in leaf among mentha mical traits in an augmented design. genotypes These were planted under green ed by using Mstat-C and Excel house and field for multiplication and software. Analysis of Variance and evaluation. The passport data of mint Duncan Multiple Range Test (DMRT) germplasm showed that collection were applied for comparison among was made from local as well as from different collections of Mentha foreign sources. Local varieties with species. variety codes include: Mentha local purple flower (1), Mentha local white RESULTS AND DISCUSSION flower (2), Peppermint / Mentha piperita Local (6), Lavender mint (7), In the present study 17 Lemon mint sp. (8), Mentha aquatica genotypes of Mentha species were (9), European Pennyroyal (10), White evaluated for both qualitative and mint (12-Rawalakot), Mint Camphor quantitative traits at National (13-Rawalakot), Spear mint (16), Agricultural Research Centre, MintBattal (17-Mansehra), Mentha Islamabad, Pakistan. Plant material rolyana/Pahari pudina (19). under study was collected from Whereas the foreign collection diverse region of the world including consist of Cool mint (11) from Canada, Saudi Arabia, Japan, Canada, China Mentha arvensis (18) from China, and Pakistan. The quantitative Nana asavi (3) and Nana maghrabi (4) traits which are mainly considered as belongs to Saudi Arabia. the economic/ agronomic traits were leaf area, leaf thickness, stem Morphological Traits thickness, stem length and petiole The data/plant were recorded length for 17 genotypes of Mentha. in triplicate. Observations documen- Analysis of variance (ANOVA) and ted for leaf area, thickness of leaf, LSD at 5% probability level showed thickness of stem, stem length and significant interaction (Table 1). Leaf petiole length under quantitative area ranged from 0.76 to 7.8 cm2 category. Whereas, qualitative traits with 2.8% C. V. High variation were shape of leaf blade, leaf margin, ranging from 8.6 to 57.6 cm was leaf apex, leaf base, leaf color, leaf observed for stem length with high arrangement, leaf venation (Figure 1), degree of S.D. (14.11), indicating a leaf odor, stem color and flower color good scope of selection for this trait (Ghafoor et al., 2005). (Table 2). however, a positive correlation was observed among Statistical Analysis thickness of leaf and stem with leaf Statistically data were analyz- area (Table 3).

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Table 1 . Mean square values of quantitative traits

Thickness Thickness Stem Petiole Leaf Variety Code of leaf of length length area(cm2) (mm) stem(mm) (cm) (cm) Mentha local purple flower 1 2.500bcd 0.1560de 2.7030ab 46.6700b 0.23def Mentha local white flower 2 6.083a 0.2230bcd 2.3467b 13efg 0.23def Nana asavi 3 3.967bc 0.2060bcde 1.3900cdef 37.67c 0.0f Nana maghrabi 4 3.133bc 0.1760cde 1.4900cde 57.67a 0.2def 5 Peppermint - Japan 3.667bc 0.1530de 0.9667f 13.33efg 0.2def Mentha piperita - Local 6 4.070b 0.2200bcd 1.6870c 14.33defg 0.6cd Lavender mint 7 1.85cd 0.1860bcde 1.7430c 17.67def 0.4def Mentha aquatica 9 2.267bcd 0.200bcde 1.0770ef 16.67def 0.2def European Pennyroyal 10 0.763d 0.2467bc 1.7330c 16defg 0.233def Cool mint 11 3.800bc 0.1967bcde 1.660c 18def 0.133ef White mint 12 1.833cd 0.1200e 1.050ef 13.33efg 0.83c Mint Camphor 13 3.4bc 0.1660cde 2.830a 8.66g 0.5cde Field mint 15 3.817bc 0.1233e 1.680c 13efg 0.433de Spear mint 16 3.817bc 0.2733b 0.960f 20de 1.83b Mint -Battal, Mansehra 17 3.033bc 0.2367bcd 1.4960cde 38.67c 0.166ef Mentha arvensis 18 7.833a 0.3667a 1.570cd 21.33d 1.8b Mentha rolyana/Pahari pudina 19 2.967bc 0.1530de 1.1300def 40.33bc 0.266def Mean ± SE 3.5 ± 0.39 0.20 ± 0.01 1.6 ± 0.12 22.6 ± 3.23 0.51±0.13 S.D. 1.70 0.06 0.53 14.10 0.58 CV (%) 2.88 0.00 0.29 198.74 0.34

Means followed by same letter (s) do not differ significantly at 0.5% probability level.

Table 2. Coefficient of Correlations among 17 genotypes for quantitative traits

Leaf area(cm2 ) Thickness of Thickness of Stem length Petiole length leaf (mm) stem(mm) (cm) (cm) L eaf area(cm2) 1 T hickness of leaf (mm) 0.588878 1 T hickness of stem(mm) 0.119067 -0.03704 1 S tem length (cm) 0-.142410 -0.05380 0.009302 1 P etiole length (cm) 0.650775 0.534084 -0.04109 -0.30716 1

Table 3. Analysis of variance (Mean Square) for quantitative traits Degree of Thickness of Thickness of Stem length Petiole length S ources Leaf area(cm 2 ) freed om leaf ( mm) stem( mm) (cm ) (cm )

P lots 2 0.922 0.00 0 0.98 0 22.33 0 0.021 C ultivars 18 8.704 ** 0.01 0** 0.87 3** 596.220 ** 1.265 ** E rror 36 1.221 0.00 2 0.06 8 15.61 1 0.046 C V (%) 30.940 20.38 0 16.32 0 17.43 0 37.610

58 PHENOTYPIC VARIATION AMONG MENTHA SPP

Table 4. Qualitative traits of Mentha spp.

Shape of Leaf Code Variety Shape of Shape of Shape of Leaf Leaf Leaves Flower leaf arrange Stem color leaf blade leaf apex leaf base color venation Odor color margin ment Mentha local Cordate Light Cross Dark reddish 1 Elliptic Undulate Acute Opposite Mint gum Lilac (purple flower) base green venulate purple Mentha local Cordate Light purplish 2 Oval Serrate Obtuse Green Opposite Pinnate Mint gum Lilac white flower base green Slightly Light Strong 3 Nana asavi Linear Acute Auricled Opposite Pinnate Light green White serrate green camphoraceous Cordate Dark Cross 4 Nana maghrabi Ovate Serrate Obtuse Opposite Mint gum Dark purple Lilac base green venulate Peppermint Cross Dark reddish Dark 5 Ovate Serrate Acute Obtuse Green Opposite Mint gum Japan venulate purple purple Mentha piperita 6 Ovate Serrate Acute Obtuse Green Opposite Pinnate Mint gum Dark purple Lilac Local Purplish Slightly 7 Lavender mint Orbicular Obtuse Truncate dark Rosulate Arcuate Spicy Reddish purple Lilac serrate green Mentha Long Dark Light purplish 9 Serrate Acute Obtuse Opposite Pinnate Pungent Lilac Aquatica elliptic green green European Slightly Light 10 Oval Obtuse Obtuse Whorled Pinnate Musky Light purple Blue Pennyroyal serrate green Cross 11 Cool mint Elliptic Aerrate Acute Obtuse Green Opposite Resh mint gum Dark Purple Lilac venulate Ordate Light Purplish 12 White mint Orbicular Serrate Obtuse Green Opposite Pinnate Camphoraceous Lilac base green Light 13 Mint Camphor Ovate Serrate Acute Obtuse purplish Opposite Arcuate Camphoraceous Light green Lilac green Cordate Dark Cross Light reddish 15 Field mint Orbicular Serrate Obtuse Opposite Mint gum Lilac base green venulate purple Long Light Cross 16 Spear mint Crenate Obtuse Truncate Opposite Acrid Light green Lilac elliptic green venulate Mint -Battal, Cross 17 Ovate Crenate Obtuse Truncate Green Whorled Mint Brown Lilac Mansehra venulate Cordate Cross Blue 18 Mentha arvensis Orbicular Crenate Obtuse Green Opposite Lemon Greenish base venulate lilac purple Mentha Slightly Light 19 rolyana/Pahari Linear Acute Obtuse Opposite Pinnate Pungent Light green Lilac serrate green pudina

The data depicted that leaf showed dwarf/short stem length (8.6 area ranged from 0.7 to 7.8 cm2. cm). Lemon mint officinais (Lemon However greater leaf area exhibited mint sp.) exhibited maximal petiole by Mentha arvensis (7.83 cm2) from length (2.2 cm) and Mansehra mint China. In contrast lowest leaf area with reduced length (0.16 cm). was acquired by European Exception to that sessile leaves were pennyroyal (0.76 cm2). Mentha apparent through Mint Maghrabi. arvensis from China again boost up These results partially agree to the with maximal leaf thickness earlier findings of Tucakov and (0.36mm) whereas White mint and Savin (1967) that revealed the Field mint showed lowest thickness morphology of mint (Mentha piperita of leaf. Difference among the 17 L.) of different origin experimentally genotypes, regarding stem thickness cultivated in Belgrade. Silva et al. ranged from 0.9 to 2.8 mm. But (2006) evaluated and conserved mint Camphor mint emerged out as a (Mentha spp) germplasm included 14 strength full stem with optimal stem species and 67 accessions, which are thickness. Among the 17 genotypes, maintained at field and greenhouse Mint Maghrabi from Saudi Arabia conditions in Brazil. attained greatest stem length (57.6 In the present study, great cm). In contrast, Camphor mint diversity was expressed within 17

59 SHAZIA ERUM ET AL. genotypes for qualitative traits types), leaf margin (>3 types), leaf including shape of leaf blade, leaf apex (3 types) and four types of leaf margin, leaf apex, leaf base, leaf bases were observed in the mint color, leaf arrangement, leaf germplasm. Moreover, similar venation, leaf odor, stem color and observations was reported by flower color (Table 4). Flower color of Shinwari et al. (2011) on the diversity mint genotypes was found lilac in of Mentha species that showed taxa most of the cases except to Mint Asavi maintained high levels of genetic and Mentha arvensis with white and polymorphism among species but blue lilac colors respectively. not among populations. The However, stem color showed great polymorphism within populations variation as visible in seven different depicted genotype richness, shades among the genotypes under recombination and gene flow. Higher study (dark reddish purple, light levels of diversity support the reddish purple, light purplish green, concept that mint have a long history light green, brown, dark purple and of independent evolution. light purple). Similar observations Much phenotypic diversity were recorded for leaf color, that are among the 17 Mentha genotypes in five shades (green, light green, collected from different agro- dark green, purplish dark green and ecological zones, and cultivated light purplish green). under field conditions were noticed. Generally, arrangement of leaf Relating to phenotypic diversity to was opposite except Lemon mint spp. origin/collecting sites of the and Lavender mint which have germplasm indicated the potential rosulate arrangement. Likewise, for future exploration mission with cross venulate, pinnate, arcuate and maximum genetic distance to reticulate types of leaf venation was assemble broad based genetic observed in Mentha spp. resources of mint for future use and The odor of Mentha leaves is a selection. As it can be a good source characteristic of each genotype due for perfumery, essential oils and to the presence of unique bioactive condiments for flavoring and compounds, polyphenols (caffeic garnishing the food items as well as acid, rosmarinic acid, ferrulic acid, with great medicinal value, the eugenol etc.) and essential oil present diversity within the genotypes could in them. Consequently, nine special facilitate the selection of the genotype aroma was smelled even in the 17 for distinct traits. samples i.e., mint gum, strong camphoraceous, mint like, spicy, LITERATURE CITED lemon, pungent, musky, rotten egg and acrid. Mentha species are Barbara, M.R. 1999. In vitro storage characterized by different aroma due conditions for mint germplasm, to the presence of bioactive HortSci. 34(2): 350. compounds and essential oils evident Clark, G.S. 1998. Menthol.Perfumer from the literature (Zviniene et al., & Flavorist, 23(5): 33 46. 1996). Moreover, shape of leaf also Ghafoor A. Ahmad, Z. and Afzal, M. exemplify great variation with diverse 2005. Use of SDS-PAGE markers patterns including leaf blade (> 5 for determining quantitative

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traits loci in blackgram [Vigna related characters in Mentha spp. mungo (L.) Hepper] germplasm. Flavour Fragr. J. 16: 340. Pakistan J. Bot. 37(2): 263-269. Morton, J. K. 1956. The chromosome Harley, R.M. and Brighton, C.A. numbers of the British Menthae. 1977. Chromosome numbers in Watsonia, 3: 244-252. the genus Mentha L. Bot. J. Linn Patra, N.K. Kumar, B. Shukla, K. Soc. 74:71-96. Ram, P. and Srivastava, H.K. Harley, R. M. 1967. The spicate 2002. Problems and issues of mints. Proc. Bot. Soc. British Agrotechnology transfer in Isles, 6: 369372. menthol mint: A case study with Harley, R.M. 1972. Notes on the variety Kosi. Proc. First National genus Mentha (Labiatae). In: Interactive Moot on Medicinal & Heywood, V.H.( ed.) Flora Aromatic Plants CIMAP, Europaea. Notulae systmaticae Lucknow. p. 440-443. ad floram Europaeam spectantes, Ruttle, M. L. 1931. Cytological and No. 12; J. Linn. Soc. Bot. 65: 223- embryological studies of the 269. genus Mentha. Gartenbauw- Heimans, J. 1938. Chromosomes in issenschaft, 44: 428468. the genus Mentha. Chronica Sharma, A. K. and Bhattacharyya, N. Botanica, 4: 389-390. K. 1959. Cytological studies on Khanuja, S. P. S. Shasany, A. K. different species of Mentha with Srivastava, A. and Kumar, S. special reference to the 2000. Assessment of genetic occurrence of chromosomal relationships in Mentha species. biotypes. Cytologia, 24: 198212. Euphytica, 111:121-125. Lawrence, B. M. 1978. A study of the Shinwari, Z. K. Sultan, S. and monoterpene interrelationships Mahmood, T. 2011. Molecular in the genus Mentha with special and morphological reference to the origin of pulegone characterization of selected and menthofuran. Ph.D. mentha species. Pakistan J. Bot. dissertation. Groningen Univer- 43(3): 1433-1436. sity, Groningen, Netherlands. Silva, D. B. Vieira, R. F. Alves, R. B. N. Linnaeus, 1767. Systema Naturae. Mendes, R. A. Cardoso, L. D. Stockholm. Tomus II: p.391-392. Queiroz, L. and Santos, I. R. I. Malinvaud, E. 1880. Simple aperc¸u 2006. Mint (Mentha spp) des hybrids dans le genre germplasm conservation in Mentha. Bulletin de la Socie´te´ Brazil. Rev. Bras. Pl. Med., Botanique de France, 27: 332- Botucatu, 8:27-31. 347. Singh, T. P. and. Sharma, A. K. 1986. McKay, D.L. and Blumberg, J.B. Mentha taxonomic status as 2006. A review of the bioactivity interpreted through cytology, and potential health benefits of genetics and phytochemistry. peppermint tea (Mentha piperita Indian J. Genetics, 46 L.). Phytother Res. 20: 619-633. (Supplement): 198-208. Mirzaie-Nodoushan, H. Rezaie, M. B. Tucakov, J. and Savin, K. 1967. and Jaimand, K. 2000. Path Morphology of mint (Mentha analysis of the essential oil piperita L.) of different origin

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