Studies on Phytosterol Biosynthesis: the Sterols of Larix decidua Leaves

Abstract

The sterols of Larix decidua leaves consist of B-sitosterol, campesterol, cycloartenol, 24-methylene cycloartanol, cycloeucalenol, 24-methylene lophenol, 24-ethylidene lophenol, together with a number of unidentified 4a-methyl sterols. The pattern of incorporation of [2-14C]mevalonic acid into these compounds has been examined.Recent reports have described the isolation from plants of sterols which may be biosynthetic intermediates between squalene and the typical phytosterols [I -71 and the incorporation of [2J4C] mevalonic acid, [I-'%] acetate and [methyl-14C] methiorline into such compounds has been described [6,7]. During studies in our Department on the biosynthesis of the phytosterol side chain [8] a plant tissue was required which contained ,4-sitosterol as the major sterol component. Preliminary examination by gasliquid chromatography showed that a number of conifer species contained predominantly /3-sitosterol together with minor amounts of campesterol. This paper reports the results of an analysis of the sterols of the newly opened leaves of the deciduous conifer larch ( L a r i x decidua). Some observations on the incorporation of [2-l4CC] mevalonic acid into sterols by the leaves of this tree are also included. EXPERIMENTAL PROCEDUREExperimental methods were generally those described previously [7]. Two batches of larch leaves were collected locally in the springtime of successive years when the leaves had been developing for about two to three weeks. Batch 1The non-saponifiable lipid (6.25 g) was extracted from the leaves (650g) by homogenisation in 1.31 of ethanol followed by addition of 200ml of water and 200 g of potassium hydroxide and refluxing for 2l/, h. The leaf debris was removed by filtration through a pad of glass wool and the non-saponifiable lipids obtained by extraction with diethyl ether in the usual manner. The total sterol (213 mg) was obtained by digitonin precipitation [7,9], the digitonides washed with diethyl ether only and then cleaved with pyridine in the usual way. The sterols were first separated by chromatography on 20 g of alumina (Brockmann grade 111) and eluting with 24'200ml portions of light petroleum containing increasing amounts of diethyl ether ["]. Three sterol fractions were obtained, the 4,4'-dimethyl sterols (24 mg), the 4a-methyl sterols (7.3 mg) and the 4-desmethyl sterols (142.6 mg). These were further purged by preparative thin layer chromatography on silica gel plates incorporating Rhodamine 6 G and with chloroform as developer [7]. Separation of the 4,4'-dimethyl and 4a-methyl fractions into their constituent components (see below) was achieved by acetylation followed by chromatography on thin layers of silica gel impregnated with loo/, silver nitrate with 40°/0 benzene in n-hexane as developer. Sterols were located by spraying with an acetone solution of Rhodamine 6 G followed by examination in ultraviolet light [lo].Batch 2 2.5 kg of larch leaves were extracted exhaustively with acetone followed by diethyl ether-light petroleum (1 : l, v/v) to yield 51 *...

Intra-Species Variation in the Chemical Composition of Needles of Larix Leptolepis

“…Although in the total lipid fraction co-chromatography indicated the possible presence of traces of~-sitosterol and campesterol, these compounds were not found in the sterol fraction. Thus, contrary to the investigation by GoAD & GoODWIN (1967) of needle extracts from Larix decidua, the free sterols were not found in the wax of L. leptolepis needles. Saponification of the 10%fraction, on the other hand, yielded a new peak indicating a comparatively high concentration.…”

Plastochromanol in the Leaves of Hevea brasiliensis

“…However, intermediates of types (iii) and (iv) are allowed, as schemes involving these do not necessarily require loss of the 3H on C-24. This argument holds whether the compound that is alkylated is desmosterol (Lederer, 1964) or, as seems more likely, some compound closely related to lanosterol (Goad & Goodwin, 1965).…”

The conversion of lanosterol, cycloartenol and 24-methylenecycloartanol into poriferasterol by Ochromonas malhamensis

“…The Conversion of Lanosterol, Cycloartenol and 24-Methylenecycloartanol into Poriferasterol by Ochromonas malhamensis By JUDITH HALL, A. R. H. SMITH, L. J. GOAD and T. W. GOODWIN Department of Biochemistry, University of Liverpool, P.O. Box 147 (Received 30 December 1968) It is postulated that the first cyclized sterol precursor in higher plants and algae is cycloartenol (I) and not lanosterol (II), the known sterol intermediate in animals and fungi (Ardenne, Osske, Schreiber, Steinfelder & Tiimmler, 1965;Benveniste, Hirth & Ourisson, 1966;Goad & Goodwin, 1966, 1967, 1969Goad, 1967;Gershengorn et al 1968). This suggestion is based on the widespread occurrence of cycloartenol (I) and 24-methylenecycloartanol (III) in plants and their rapid labelling from [2-14C]mevalonate and [1-14C]acetate under conditions where phytosterols are being actively synthesized (see references above).…”