Evaluation of Kasuri Methi Trigonella Foenum- Graecum L.Var

Home , Kasur, Kasur District

Pakistan J. Agric. Res. Vol 24 No. 1-4, 2011. EVALUATION OF KASURI METHI TRIGONELLA FOENUM- GRAECUM L.VAR. TO ESTABLISH GI RIGHT OF PAKISTAN

Shazia Erum, Rashid Anwar and Shahid Masood*

ABSTRACT:- Geographical indicator (GI) of crops create a positive impression of the product quality, the environmental virtue and human skill of the area. In the present study, comparative analysis of total seed proteins of Kasuri methi (GI of Kasur, Pakistan) was evaluated with other Trigonella genotypes by SDS-PAGE. Results showed that at protein level Kasuri methi acquired a unique status as a G.I of Kasur. Cluster analysis (UPGMA) of 28 genotypes including both methi and methray from various agro ecological zones of Pakistan were interlinked to some extent however Kasuri methi make their identity by standing alone among tested genotypes on the dendrogram, due to the geographical environment of Kasur in Punjab. In this connection ‘Benefit sharing’ mechanism can be accelerated by escalating farmer’s participation for more cultivation in the interest of country.

Key Words: Trigonella foeum-graecum; Seeds; Diversity; Geographical Indicator; Phenotype; Pakistan. INTRODUCTION industries, etc. Pharmaceutically medici- Fenugreek, Trigonella foenum-graecum nal species of the genus Trigonella are T. L. is an ancient and annual legume crop foenum-graecum L., T. balansae, T. mainly grown for multiple uses in many corniculata, T. maritima, T. spicata, T. parts of the world. Trigonella, comes from occulta,T. polycerata, T. calliceras, T. cretica, Latin meaning ‘little triangle’, refers to the T. caerulea, T. lilacina, T. radiata, T. spinosa triangular shape of its small yellowish- (Petropoulos, 2002; Basu, 2006). The bio- white flowers. The species epithet foenum- logical and pharmacological actions of graecum means ‘Greek hay’ (Rosengarten, fenugreek are attributed to the variety of 1969). It is also called ‘ox horn’ or ‘goat horn’ its constituent, namely, steroids, N-com- because of the two seed pods projecting in pounds, polyphenolic substances, volatile opposite directions usually from the nodes constituents, amino acids, etc. of the stem base that resemble ox or goat (Mehrafarin et al., 2010). horns (Petropoulos, 2002). Landraces and Fenugreek seed contains 45-60% car- species of Trigonella have been found in bohydrates, mainly mucilaginous fiber parts of Europe, northern Africa, west and (galactomannans), 20-30% proteins high in south Asia, north and south America and lysine and tryptophan, 5 - 10% fixed oils Australia (Basu, 2006; Acharya et al., (lipids), pyridine alkaloids, mainly trigonel- 2006). line (0.2 - 0.38%), choline (0.5%), Fenugreek leaves and seeds are con- gentianine and carpaine, the flavonoids sumed in different countries around the apigenin, luteolin, orientin, quercetin, world for different purposes such as medi- vitexin and isovitexin, free amino acids, cine (anti-diabetic, lowering blood sugar and such as 4-hydroxyisoleucine (0.09%), argi- cholesterol level, anti-cancer, anti-micro- nine, histidine and lysine, calcium and bial, etc.), making food (stew with rice in iron, saponins (0.6 - 1.7%), glycosides yield- Iran, flavor cheese in Switzerland, syrup, ing steroidal sapogenins on hydrolysis (di- mixed seed powder with flour for making osgenin, yamogenin, tigogenin, flat bread in Egypt, curries, dyes, young neotigogenin), cholesterol and sitosterol, seedlings eaten as a vegetable, etc.), roasted vitamins A, B1, C and nicotinic acid and grain as coffee-substitute (in Africa), con- 0.015% volatile oils ‘n-alkanes’ and ‘ses- trolling insects in grain storages, perfume quiterpenes’ (Budavari, 1996; Newall et al., *Institute of Agricultural Biotechnology and Genetic Resources Institute, National Agricultural Research Cen- tre, Islamabad, Pakistan.. 25 SHAZIA ERUM ET AL. 1996; Mehrafarin et al., 2010). attributed as a major reason for failure to Most renowned Pakistani fenugreek consider the possibility that farmers have with remarkable aroma cultivated in the an intellectual investment in their folk Kasur district of Punjab province well varieties (Cleveland and Stephen, 1997; known as ‘Kasuri methi’. Kasur (total area Nagarajan, 2005). 3,995 km2, elevation 218 m.a.s.l). Methi The objective of the study is to evalu- (fenugreek) from Kasur is very famous for ate the Trigonella species collected from its fragrance throughout the country and various agro ecological zones of Pakistan became geographical indicator of Kasur as and to distinguish Kasuri methi at protein Kasuri Methi, so it can be claimed/patent level by SDS- PAGE. for GI or AO. ‘Appellation of Origin’ (AO) means that a product originates in a spe- MATERIALS AND METHODS cific geographic region and the character- The research was conducted at Na- istic qualities of the product are due to the tional Agricultural Research Centre, geographical environment, including natu- Islamabad, Pakistan. Seeds of 28 genotypes ral and human factors (Nagarajan, 2005). of Trigonella (Trigonella foenum graecum- The farming community contributions Methi and Trigonella corniculata-Methray) have to be recognized and rewarded and were collected from various agroecological benefit sharing should become mandatory. zones of Pakistan including Mianwali, Relationship between farmers’ variety (FV) Pakpattan, Bahawalpur, Faisalabad, and GI provides certain rights (Plant Pro- Shorkot, Narowal, Karachi, Okara, tection Variety and Farmers Rights Act Talwandi, Bahawalnagar, Daska, Sargodha, 2001 in India) to farmers, such as to save, Quetta and Kasur according to the quad- use, sow, re-sow, exchange, share or sell rate method. The current study was initi- his farm produce including that of the reg- ated to investigate the potential electro- istered variety. In Pakistan, farmers who phoresis for intra specific characterization develop new plant varieties like any other of Trigonella on the basis of their total seed plant breeder can apply their material for protein and explore distinctiveness of the conduct of Distinctness, Uniformity and kasuri methi (landrace) with other geno- Stability (DUS) testing and registration. In types of Trigonella. the last hundred years there has been a For judgment of Kasuri methi as a G.I., drive for improved agriculture and that has seeds samples were divided into two groups. replaced farmers’ variety in several crops Group I consists of 22 seed samples of with new varieties developed by the plant Trigonella foenum graecum-Methi and Group breeders. Yet farmer’s variety is still domi- II comprises seed samples of Trigonella nant in pulses, vegetables, melons, etc. corniculata-Methray, along with Kasuri There are lots of controversial patent cases methi seed sample in both the groups. involving traditional Knowledge and Ge- Genetic diversity of Trigonella species netic Resources including hoodia cactus, evaluated by sodium dodecylsulphate poly- turmeric, neem, ayahuasca and basmati rice acrylamide gel electrophoresis (SDS-PAGE) case, facing problems for their patent rights according to the standard method as geographical indicators. The GI for agri- (Leammli, 1970; Rabbani et al., 2001; cultural goods like Basmati rice, coffee, tea, Ahmed et al., 2009 and 2010; Turi et al., wine, etc. revolve around consumer pref- 2010; Cheema et al., 2010). The subunit erences for the palate feeling, aroma and molecular weight of the protein bands was physical appearance that enhances the determined according to Weber and Osborne appetite. An ideal mixture of all these at- (1969). Each band was considered as a tributes raises the value of the product due character for which the presence or ab- to reasons of consumer preference sence was coded 1 or 0 respectively in a (Nagarajan, 2005). Lack of documentation data matrix for numerical analysis. Den- of indigenous farmers’ plant breeding is sity of the protein band was not considered. 26 EVALUATION OF KASURI METHI Unweighted pair group method with an major groups were observed on the dendro- arthimetic average (UPGMA) cluster analy- gram (Figure1), where first and third group sis was used to infer genetic relationships showed 100% similarity among themselves and phylogeny among the genotypes of (including 1,2,3,4,5,19,18,17,16,15,14,6,7,8 Trigonella. All computations were done by and 20,21) while 23% deviation was ob- using the NTSYS-pc, Version 2.2 package served in the second group (9,13,11,10 and (Rohlf, 2005; Rabbani et al., 2001 and 12) whereas the genotype No. 22, popularly 2008). For an effective breeding programme, known as “Kasuri Methi” from Kasur (22) information regarding the extent and na- stand alone and expressed itself as a G.I ture of genetic diversity within a crop spe- even at protein level. cies is essential. It is particularly very use- The variable banding pattern shown by ful for characterization of individual acces- Kasuri methi made it different from other sions and as a guide in the selection of par- genotypes. As proteins are the translational ents for hybridization programme. products of genes therefore, the differences in proteins could be related with such dif- RESULTS AND DISCUSSION ferences in genes responsible for synthe- Seed protein analysis by SDS-PAGE has proved to be an effective way of reveal- Table 1. Passport data of Trigonella ing the differences and relationship be- germplasm tween taxa. The high stability of the seed Accession Town/city protein profile and its additive nature make Group I seed protein electrophoresis a powerful tool PAK 020978 Mainwali in elucidating the origin and the evolution PAK 021703 Pakpattan of cultivated plants. The present investi- PAK 022254 Bahawalpur gation revealed variation in the evaluated PAK 021131 Faisalabad 28 accessions of Trigonella species collected PAK 021156 Faisalabad from various ecological zones of Pakistan PAK 021330 Shorkot (Table 1, Figure 1 and 2) and to make a dis- PAK 021675 Narowal tinction of Kasuri methi from Kasur with PAK 021986 Karachi other genotypes through slab type SDS- PAK 021748 Karachi PAGE using 12.25% polyacrylamide gel. PAK 021711 Narowal Initially, 28 seeds samples were di- PAK 021696 Okara vided into two groups. Group I, (22 samples PAK 021117 Faisalabad of methi i.e., Trigonella foenum graecum) and PAK 022260 Talwandi group II (six samples of methray i.e., PAK 022257 Bahawalnagar Trigonella corniculata) including seed PAK 022219 Daska sample of Kasur in both groups. The data PAK 021908 Sargodha were scored for the presence (1) and ab- PAK 021900 Sargodha sence (0) of the bands and entered in a bi- PAK 021889 Sargodha nary data matrix. Based on the results of PAK 021882 Faisalabad electrophoretic band spectra, similarity Donated Kasur index was calculated for all possible pair of Donated Kasur electrophoregrams. The similarity matrix Native/indigenous Kasur thus generated was converted to a dissimi- Group II larity matrix and used to construct the den- 21879 Faisalabad drogram. Studying genetic diversity and 20595 Faisalabad taxonomic relationships of plants previously 21947 Karachi scored for many plants (Bult and Kiang, 21857 Quetta 1992; Zviniene and Pank, 1996). Results of 20593 Faisalabad Group I showed that limited interspecific 11122 Native/indig- diversity was found at protein level. Three enous of Kasur 27 SHAZIA ERUM ET AL. sis of these proteins. The molecular tech- 2006). Inter and Intra- specific variation in niques including SDS-PAGE are very strong SDS-PAGE electrophoregrams of total seed to determine among the biological organ- proteins in wheat, barley and their wild isms based on such molecular markers. relatives was reported by Masood et al. Hence are very common to investigate the (1994). Moreover, electrophoresis of seed genetic diversity among the species, vari- proteins was previously used for cultivar eties and genotypes (Irfan, 2000; Ahmad identification for many other plants as Vicia and Kamal, 2002). The SDS-PAGE tech- faba (Stegmann et al., 1980) and Linum niques have more advantages in the clas- usitatissimum (Sammour, 1988). The tech- sification of genotypes (Abd-El-Zaher et al., nique had been utilized by Masood et al., (2005) for differentiation in different genotypes of wheat and mustard (Rabbani et al., 2001), Irfan (2000) in Adhatoda vesica, Ahmad and Kamal (2002) in Hyppophae rhamnides, Vigna mungo (Ghafoor et al., 2002), Mentha (Badr et al., 2003), Raphanus sativus (Jatoi et al., 2003), Pyrus (Ahmed et al., 2009),Triticum aestivum (Ahmed et al., 2010), Castor bean (Cheema et al., 2010), Brassica species (Turi et al., 2010). Results of Group II explained by a similarity matrix based on the proportion of shared peptide fragments is used to establish the level of relatedness between Methi and Methray. Paired wise estimates of similarity ranged from 0.40 to 1. Total three Figure 1. UPGMA cluster analysis showing the groups emerged on the dendrogram diversity among Trigonella landraces (Figure 2). Group I showed 100% similarity of Pakistan based on seed proteins with unique position of methi be- among themselves. Group II is also the rep- longs to Kasur No.22 resentative of accessions of Methray. Third independent group represent the accession collected from Kasur commonly known as Kasuri Methi. Comparative analysis of pro- 21879 tein profile of Kasuri Methi with Methray (Group I and II) showed that it is 60% dis- 20595 similar (Figure 2). Accession 21879, 20595 Group I belongs to Faisalabad, 21947 was from 21856 Karachi and 21857 from Quetta. The accession 20593 from Faisalabad depicted 21947 some distinctness as compared to other accessions from Faisalabad indicating in- 20593 Group II tra species divergence. The similarity among the accessions from Faisalabad, 11122 Group III Karachi and Quetta may be due to their common origin. Karachi and Quetta are not 0.35 0.350.51 0.68 0.84 1.00 major agricultural production areas but due to high demand for Methi in local market Figure 2. Comparative analysis of methray the farmers may have obtained seeds from (Trigonella corniculata) with Kasuri Faisalabad which has been a centre of ag- Methi (11122) ricultural research and production of ma-

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