Essential oil constituents and RAPD markers to establish species relationship in Cymbopogon Spreng. (Poaceae)

https://doi.org/10.1016/j.bse.2004.06.011Get rights and content

Abstract

Nineteen Cymbopogon taxa belonging to 11 species, two varieties, one hybrid taxon and four unidentified species were analysed for their essential oil constituents and RAPD profiles to determine the extent of genetic similarity and thereby the phylogenetic relationships among them. Remarkable variation was observed in the essential oil yield ranging from 0.3% in Cymbopogon travancorensis Bor. to 1.2% in Cymbopogon martinii (Roxb.) Wats var. motia. Citral, a major essential oil constituent, was employed as the base marker for chemotypic clustering. Based on genetic analysis, elevation of Cymbopogon flexuosus var. microstachys (Hook. F.) Soenarko to species status and separate species status for C. travancorensis Bor., which has been merged under C. flexuosus (Steud.) Wats were suggested towards resolving some of the taxonomic complexes in Cymbopogon. The separate species status for the earlier proposed varieties of C. martinii (motia and sofia) is further substantiated by these analyses. The unidentified species of Cymbopogon have been observed as intermediate forms in the development of new taxa.

Introduction

The genus Cymbopogon (Poaceae) is known to include about 140 species. Among these, more than 52 have been reported to occur in Africa, 45 in India, six each in Australia and South America, four in Europe, two in North America and the remaining are distributed in South Asia (Jagadish Chandra, 1975b). Most of these species produce characteristic aromatic essential oils that have commercial importance in perfumery, cosmetics and pharmaceutical applications. The Cymbopogon essential oils are characterised by monoterpene constituents like citral, citronellol, citronellal, linalool, elemol, 1,8-cineole, limonene, geraniol, β-carophyllene, methyl heptenone, geranyl acetate and geranyl formate. Citral is one of the important components of the oil present in several species of Cymbopogon with wide industrial uses such as raw material for perfumery, confectionery and vitamin A.

After Sprengel named this genus in 1815, a number of taxonomists have attempted to classify the species of Cymbopogon. Hackel, 1887, Hooker, 1897 have treated this genus as a subgenus of Andropogon. However, Stapf (1906) raised Cymbopogon to its original rank of a genus and this has been accepted by all the later taxonomists. Taxonomically, the species of Cymbopogon have been divided into three series viz. ‘Shoenanthi’, ‘Rusae’ and ‘Citrati’ (Stapf, 1906). The leaves of the species in ‘Schoenathi’ series are thin, in ‘Rusae’ subcordate and in ‘citrati’ lanceolate. The identification and classification of Cymbopogon species have become difficult because of the occurrence of numerous transitional forms which are supposed to have arisen by hybridisation (Bor, 1960) and the existence of many varieties and races (Jagadish Chandra, 1975a). Some species, such as Cymbopogon citratus, are not known to flower, others like Cymbopogon nardus and Cymbopogon winterianus flower only rarely and the polyploid forms of Cymbopogon flexuosus and Cymbopogon coloratus flower, but do not set seed.

The morphological variation and oil characteristics of various species and varieties of Cymbopogon have been reported (Husain, 1994), but such information is not sufficient to define precisely the relatedness among the morphotypes and chemotypes. For instance, C. martinii var. sofia and C. martinii var. motia, are morphologically almost indistinguishable, but show distinct chemotypic characteristics in terms of oil constituents (Guenther, 1950). Conversely, phenotypically and taxonomically well distinguishable species produce oils of almost identical chemical compositions such as lemongrass oils from C. citratus and C. flexuosus (Anonymous, 1988). Such phenotypic traits, whether morphological or chemotypic are basically the phenotypic expression of the genotype, while DNA markers are independent of environment, age and tissue and expected to reveal the genetic variation more conclusively in assessing such variations. Introgression of various traits, intermittent mutations and selection through human intervention may lead to variation in chemotypic characters across geographical distributions (Kuriakose, 1995). While natural hybridisation may lead to the formation of morphological or chemotypic intermediates, defining taxa purely on this basis may not be appropriate. The earlier works on phytochemical (Patra et al., 1990, Dhar et al., 1993), biochemical and physiological parameters (Nandi and Chatterjee, 1987), development of agrotechnology (Nair et al., 1979, Rao et al., 1985), genetic improvement (Ganguly et al., 1979, Maheshwari and Sethi, 1987) and breeding approaches (Kulkarni and Rajgopal, 1986, Rao and Sobti, 1987a, Shyalaraj and Thomas, 1993) of different Cymbopogon species and varieties appear scattered, and do not address the question of relationships among different taxa.

Molecular markers provide extensive polymorphism at DNA level used for differentiating closely related genotypes (Pecchioni et al., 1996) and also to find out the extent of genetic diversity (Jain et al., 2003). DNA based markers such as random amplified polymorphic DNA (RAPD) have been widely used in a number of plant groups for variety of purposes such as cultivar identification, diversity studies, parentage determination, developing breeding programmes and conservation strategies. Earlier, an attempt was made by our group using RAPD analysis to trace the ancestors of cultivar Java II within C. winterianus (Shasany et al., 2000), but at the inter-specific level, information on molecular characterization for establishing the phylogenetic relationships among Cymbopogon is scarce. This investigation deals with estimating the extent of genetic diversity using RAPD analysis in relation to chemo-variation observed in oil constituents among different commercially and agronomically used 19 taxa of Cymbopogon in India.

Section snippets

Plant material

Nineteen Cymbopogon accessions belonging to 11 species, two varieties, one hybrid taxon and four unidentified species collected from the wild were included in this study. The representative taxa had been previously collected either from the wild or received through the National Genebank in the genotype exchange programmes from other laboratories (as slips). These with individual identification are maintained in the National Genebank (field genebank) for medicinal and aromatic plants at Central

Analysis of essential oil profiles

The accessions representing different species of Cymbopogon showed remarkable variation in essential oil composition and yield, which ranged from 0.3% in Cymbopogon travancorensis to 1.2% in Cymbopogon martinii (Table 2). The major essential oil components citral ‘a’ and ‘b’, were detected in Cymbopogon pendulus, C. flexuosus and C. citratus with highest in C. citratus. Essential oil yield of about 0.6% was observed in all citronella taxa viz. C. nardus var. confertiflorus, C. nardus var. nardus

Discussion

The Cymbopogon species complex has been divided into three taxonomic series (Stapf, 1906) according to dominant essential oil components as markers for each. In the present analysis, most of the taxa are from series ‘Citrati’ except C. martinii and C. caesius which belong to series ‘Rusae’ and C. jwarancusa to series ‘Schoenanthi’. The taxa from ‘Citrati’ series form a separate larger cluster interrupted with series ‘Schoenanthi’ (C. jwarancusa) and series ‘Rusae’ (C. caesius) in the RAPD

Acknowledgements

We thankfully acknowledge the financial support of the Department of Biotechnology (DBT) and Council of Scientific and Industrial Research (CSIR), Government of India, New Delhi.

References (36)

  • S.C. Nath et al.

    Comparison of volatile inflorescence oils and taxonomy of certain Cymbopogon taxa described as Cymbopogon flexuosus (Nees ex Steud.) Wats

    Biochem. Syst. Ecol.

    (2002)
  • Anonymous
  • N.L. Bor

    The Grasses of Burma, Ceylon, India and Pakistan

    (1960)
  • J. Christopher et al.

    Karyomorphological and phylogenetic studies of Cymbopogon species from India

    J. Cytol. Genet.

    (1986)
  • A.K. Dhar et al.

    Changes in oil concentration and its constituents, herbage and oil yield in five genotypes of Cymbopogon jwarancusa following foliar application of nitrogen

    Indian Perfumer

    (1993)
  • E. Guenther
    (1950)
  • D. Ganguly et al.

    “RRLJOR-3-1970” an improved strain of Java Citronella (Cymbopogon winterianus Jowitt.)

    Indian Perfumer

    (1979)
  • B.K. Gupta

    Studies in the genus Cymbopogon Spreng III

    Proc. Ind. Acad. Sci.

    (1970)
  • B.K. Gupta et al.

    Cultivation and utilization of genus Cymbopogon in India

    Indian Perfumer

    (1978)
  • Hackel, E., 1887. Graminae in Engler and Prantl Pflanzen Familien Teil II. Anterling 2....
  • J.D. Hooker

    Graminae

    (1897)
  • A. Husain

    Essentials Oil Plants and Their Cultivation

    (1994)
  • K.S. Jagadish Chandra

    Recent studies on Cymbopogon Spreng. With special reference to Indian Taxa

    J. Plant Crops

    (1975)
  • K.S. Jagadish Chandra

    Cytogenetical Evolution in Some Species of Cymbopogon Cited in Advancing Frontiers in Cytogenetics

  • N. Jain et al.

    Molecular diversity in Phyllanthus amarus assessed through RAPD analysis

    Curr. Sci.

    (2003)
  • W. Jennings et al.

    Qualitative Analysis of Flavour and Fragrance Volatile by Capillary GC

    (1980)
  • S. Karthikeyan et al.

    Florae Indicae Enumeratio: Monocotyledonae

    (1989)
  • S.P.S. Khanuja et al.

    Rapid isolation of PCR amplifiable DNA from the dry and fresh samples of plants producing large amounts of secondary metabolites and essential oils by modified CTAB procedure

    Plant Mol. Biol. Report.

    (1999)
  • Cited by (110)

    View all citing articles on Scopus
    View full text