N-Alkane Profile of Argemone Mexicana Leaves

Home , Hentriacontane, Nonacosane

n-Alkane Profi le of Argemone mexicana Leaves Indranil Bhattacharjeea, Anupam Ghosha, Nandita Chowdhurya, Soroj Kumar Chatterjeea, Goutam Chandraa,*, and Subrata Laskarb a Microbiology Research Unit, Parasitology Laboratory, Department of Zoology, The University of Burdwan, Burdwan-713104, West Bengal, India. E-mail: [email protected] b Natural Product Laboratory, Department of Chemistry, The University of Burdwan, Burdwan-713104, West Bengal, India * Author for correspondence and reprint requests Z. Naturforsch. 65 c, 533 – 536 (2010); received March 1/April 16, 2010 An n-hexane extract of fresh, mature leaves of Argemone mexicana (Papaveraceae), con- taining thin-layer epicuticular waxes, has been analysed for the fi rst time by TLC, IR and GLC using standard hydrocarbons. Seventeen long-chain alkanes (n-C18 to n-C34) were identifi ed and quantifi ed. Nonacosane (n-C29) was established as the n-alkane with the highest amount, whilst octadecane (n-C18) was the least abundant component of the extracted wax fraction. The carbon preference index (CPI) calculated for the hydrocarbon sample with the chain lengths between C18 and C34 was 1.2469, showing an odd to even carbon number predominance. Key words: Alkane, Epicuticular Wax, Carbon Preference Index

Introduction the alkane fraction by application of a nonpolar solvent (hexane) and to characterize the n-alkane Protecting coats, so-called waxes, present on the profi le of the epicuticular waxes of leaves of this surface of plant parts have an almost universal plant by gas liquid chromatography (GLC). occurrence. Their main functions are to protect plants from physical damage and, especially from Material and Methods the loss of water (Schoonhoven et al., 1998), with long-chain hydrocarbons/alkanes and their deriv- Wax extraction atives present as principle chemical constituents Fresh, mature leaves of A. mexicana were har- (Baker, 1982). Alkanes from epicuticular waxes vested randomly during July 2008 from plants are used as indicators of taxonomic relations be- growing in the surroundings of Department of tween plant species, and now attention has been Zoology, University of Burdwan, India. 50 g directed towards the possibility of using their dis- freshly collected mature leaves of A. mexicana tribution as a means of establishing a chemotaxo- were rinsed with distilled water and dried on pa- nomic system. per towelling. Leaves were then dipped in 2 L of Argemone mexicana L. (Papaveraceae), com- cold n-hexane for 45 min, and surface wax was monly known as prickly poppy, is a common me- extracted at room temperature. The crude extract dicinal plant. The antibacterial potential of an al- was fi ltered through Whatman fi lter paper no. 41 coholic and aqueous extract of the plant leaves has (Whatman, Maidstone, UK), and the solvent was been documented by Bhattacharjee et al. (2006). removed under reduced pressure. The extract was The petroleum ether extract of the plant exhibit- then fractionated by preparative thin-layer chro- ed larvicidal activity against laboratory-colonized matography (TLC) using carbon tetrachloride and fi eld-collected Culex quinquefasciatus larvae as mobile phase. The thin-layer chromatograph- (Karmegam et al., 1997). But neither the chemi- ic plates (thickness of 0.5 mm) were prepared cal composition nor the concentration of aliphatic with silica gel G (Merck, Mumbai, India) using compounds present in the thin-layer epicuticular a Unoplan coating apparatus (Shandon, London, wax of this plant have been determined so far. UK). The single hydrocarbon band was identifi ed The aim of the present paper was to isolate only through co-TLC studies with standard samples

(Sigma, St. Louis, MO, USA). The band was elut- ponents were characterized through co-TLC with ed from the layer with chloroform; it showed no authentic samples of n-alkanes (Sigma). absorption of any detectable functional group in All solvents employed were of analytical grade the infrared region. The presence of alkanes was and obtained from Merck (Mumbai, India). further conﬁ rmed by argentometric TLC.

Results and Discussion GLC analysis of surface wax The purifi ed hydrocarbon fraction was ana- Fig. 1 shows the GLC separation of the hydro- lysed directly by GLC on a Hewlett Packard carbons of the n-hexane extract of the epicuticular (Palo Alto, CA, USA) model 5890 series II instru- wax of mature leaves of A. mexicana. Seventeen ment fi tted with an HP-1 capillary column (25 m long-chain alkanes (n-C18 to n-C34) were identi- × 0.01 mm internal diameter) and a fl ame ioni- fi ed and quantifi ed. The carbon preference index zation detector; nitrogen was used as carrier gas (CPI) calculated for the hydrocarbon sample with at a fl ow rate of 16.5 mL/min. Oven temperature the chain lengths between C18 and C34 was 1.2469, was 170–300 °C at a 5 °C/min rise, initial period showing an odd to even carbon number predomi- was 1 min, and the fi nal period was 15 min. Com- nance.

Fig. 1. Gas liquid chromatogram of the n-hexane extract of fresh, mature leaves of A. mexicana. I. Bhattacharjee et al. · n-Alkane Proﬁ le of Argemone mexicana 535

The surfaces of leaves, ﬂowers, fruits, and non- Table I. Distribution of n-alkane constituents in the n- woody stems are covered by a cuticle made of hexane extract of mature leaves of A. mexicana. cutin and waxes (Jetter et al., 2006). Among the Peak Compound Retention Amount cuticular wax layer, an intracuticular and an epi- time [min] (mol%) cuticular layer can be distinguished according 1 Octadecane (n-C18) 7.678 1.2080 to the wax location inside the cutin matrix and 2 Nonadecane (n-C19) 9.676 1.4770 exterior to it, respectively (Jeffree, 1986). Epicu- 3 Eicosane (n-C20) 11.702 1.6337 4 Heneicosane ( -C ) 13.692 2.2129 ticular waxes serve many physiological functions, n 21 5 Docosane (n-C ) 15.510 1.0270 including protection against UV light (Reicosky 22 6 Tricosane (n-C23) 17.472 7.1622 and Hanover, 1978) and moderation of gas ex- 7 Tetracosane (n-C24) 19.236 3.8396 change through stomata (Jeffree et al., 1971). Be- 8 Pentacosane (n-C25) 20.934 6.9558 sides, the epicuticular waxes form the true sur- 9 Hexacosane (n-C26) 22.563 6.4799 face of the plant organs and, therefore, play an 10 Heptacosane (n-C27) 24.129 7.4189 11 25.630 7.5132 important ecological role in the interaction with Octacosane (n-C28) 12 Nonacosane (n-C ) 27.088 10.6814 insects (Müller, 2006) and pathogens (Carver and 29 13 Triacontane (n-C30) 28.635 9.4555

Gurr, 2006). There are signiﬁ cant differences in 14 Hentriacontane (n-C31) 30.443 10.4134 the individual and total n-alkane concentrations 15 Dotriacontane (n-C32) 32.592 5.8923 in plant leaves (Jetter et al., 2000; Piasentier et 16 Tritriacontane (n-C33) 35.197 5.2423 al., 2000; Barik et al., 2004; Jetter and Schäffer, 17 Tetratriacontane (n-C34) 38.380 4.8462 2001). Higher alkanes are known to be one of the main components of cuticular waxes of plant n-alkane with highest amount, whilst octadecane leaves and stems. Moreover, in the homologous (n-C ) was the least abundant (Table I). series the hydrocarbon fraction with an odd num- 18 In conclusion, A. mexicana is a medicinal plant ber of carbon atoms predominates considerably with established antimicrobial and mosquito- (Isidorov and Vinogorova, 2003; Buschhaus et al., larvicidal property found in different parts of 2007). Further, Hellmann and Stoesser (1992), the world. The extraction of epicuticular wax by Dutton et al. (2000), and Piasentier et al. (2000) a nonpolar solvent made it possible to broaden reported that heptacosane (n-C27), hentriacontane greatly the list of compounds detected previously. (n-C31), and tritriacontane (n-C33) were the most This analysis also provides a basis for further re- prominent alkanes present in the epicuticular search on interactions between leave surfaces of wax. However, our study produced different re- this plant and insect herbivores or pathogenic mi- sults as nonacosane (n-C29) was established as the croorganisms.

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