The identification of 24-methylene-24,25-dihydrolanosterol and other possible ergosterol precursors in Phycomyces blakesleeanus and Agaricus campestris

The following sterols have been isolated from the fungi, Phycomyces blakesleeanus and Agaricus campestris: ergosterol, lanosterol, 24-methylene-24,25-dihydrolanosterol and episterol. 4,4-Dimethyl-5α-ergosta-8.24(28)-dien-3β-ol and 4α-methyl-5α-ergosta-8,24(28)-dien-3β-ol have been tentatively identified. Evidence for the incorporation of label from l-methionine-[methyl-¹⁴C] into some of these sterols in P. blakesleeanus has been obtained. The significance of these sterols in ergosterol biosynthesis is discussed.     

The occurrence of C29 sterols with different configurations at C-24 in Cucurbita Pepo as shown by 270 MHz NMR

The 270MHz NMR spectra of the major sterols of pumpkin seeds show that the configuration at C-24 of 24-ethyl-5α-cholesta-7,22,25-trien-3β-ol and 24-ethyl-5α-cholesta-7,25-dien-3β-ol is 24β_F = (24S) whereas the α-spinasterol has the 24α_F = (24S) configuration.     

Observations on the biosynthesis of 24-methylcholesterol and 24-ethylcholesterol by Zea mays

Examination of the sterols of Zea mays shoots has established that the 24-ethylcholesterol is predominately the 24α-epimer, sitosterol, but the 24-methylcholesterol is a mixture of the 24α- and 24β-epimers. After incubation of Z. mays shoots with [2-¹⁴C, (4R)4-³H₁]mevalonic acid the sitosterol had a ³H: ¹⁴C atomic ratio of 2.09:5 which is consistent with previous results indicating that a Δ²⁴⁽²⁵⁾ -sterol is implicated in its biosynthesis. By contrast, the 24α- and 24β-methylcholesterol mixture had a higher ³H: ¹⁴C atomic ratio of 2.82:5. This can be explained by the operation of two routes for the elaboration of the 24-methylcholesterol side chain. One may proceed via Δ²⁴⁽²⁵⁾- and Δ²⁴⁽²⁵⁾-sterols to produce the 24α-methylcholesterol with a ³H: ¹⁴C atomic ratio of 2:5. The other route may involve reduction of either a Δ²⁴⁽²⁸⁾-, a Δ²³- or a Δ²⁵-sterol intermediate to give the 24β1-methylcholesterol with a ³H: ¹⁴C atomic ratio of 3:5. The proportion of these two labelled compounds in the mixture then determines the observed ³H: ¹⁴C atomic ratio (2.82:5). Some evidence for the formation of a Δ²⁵-compound, cyclolaudenol, by Z. mays shoots was provided by incorporation studies employing either [2-¹⁴C]mevalonic acid or [Me-¹⁴C]methionine as the sterol precursor.     

Biosynthesis of Δ5.23 sterols in etiolated coleoptiles from Zea mays

In addition to the previously found ergosta-5, E-23-dien-3β-ol and 5α-ergosta-7, E-23-dien-3β-ol, the following Δ23 sterols have been identified in etiolated maize coleoptiles: cyclosadol, 4α, 14α-dimethyl-5α-ergosta-8, E-23-dien-3β-ol, 4α, 14α-dimethyl-9β, 19-cyclo-5α-ergosta-8, E-23-dien-3β-ol and 4α-methyl-5α-ergosta-7, E-23-dien-3β-ol. The incubation of maize coleoptile microsomes in the presence of cycloartenol and of [¹⁴C-methyl]S-adenosyl methionine gave a mixture of labelled 24-methylene cycloartanol and cyclosadol. No trace of cyclolaudenol could be detected in these conditions. It is suggested that Δ²³ sterols are products of the C-24 methyltransferase reaction and they probably do not arise from a Δ²⁴ → Δ²³ isomerization occurring at a later stage of the biosynthesis. The Δ¹³-sterols may play an intermediary role in the biosynthesis of 24-methyl sterols in this plant material.     

Sterols,sterol esters and fatty acids of Botrydium granulatum, Tribonema aequale and Monodus subterraneus

The composition of the sterols, sterol esters and fatty acids has been determined in 8-, 11- and 14-day cultures of three members of the Xanthophyceae, Botrydium granulatum, Tribonema aequale and Monodus subterraneus. The main sterols, whether esterified or unesterified, were cholesterol and clionasterol, whose proportions do not vary with age of culture. Much smaller quantities of cycloartenol and 24-methylenecycloartanol were also found in all three algae. The C₁₆ fatty acids are the most common fatty acids in all three algae with C_16:1 being particularly abundant. B. granulatum and T. aequale, however, differ from M. subterraneus in having polyunsaturated C₁₆ fatty acids and a smaller proportion of C_20:5.     

Fern constituents: Cycloartane triterpenoids and allied compounds from Polypodium formosanum and P. niponicum

From the rhizomes of Polypodium formosanum, new triterpenoids of the cycloartane group, (24R)-cyclolaudenol and (24R)-cyclomargenol, were isolated as the corresponding acetates, alcohols and ketones, and their structures were established. Also, from the rhizomes of P. niponicum eight acetates of cycloartane derivatives and two acetates of new methyl sterols were isolated and characterized.     

Co-occurrence of c-24 epimeric 24-ethyl-Δ7 -sterols in the roots of Trichosanthes japonica

¹³C NMR spectroscopy has demonstrated that the major components (～80%) of 24 - ethyl - 5α -cholest - 7 - en - 3β - ol and 24 - ethyl - 5α - cholesta - 7,trans - 22 - dien - 3β - ol isolated from the roots of Trichosanthes japonica are the 24α-epimers, 22-dihydrospinasterol and spinasterol, accompanied by minor amounts (～20%) of their 24β-epimers, 22-dihydrochondrillasterol and chondrillasterol, respectively. The possible biosynthetic pathway leading to these sterols is discussed. This seems to be the first instance of the detection of 22-dihydrochondrillasterol in a higher plant.     

Incorporation of methionine-[methyl-2H3] and mevalonic acid-[2-14C,(4R)-4-3H1] into Phycomyces blakesleeanus sterols

Ergosterol, episterol, 4α-methyl-5α-ergosta-8,24(28)-dien-3β-ol and 24-methylene-24,25-dihydrolanosterol, isolated from Phycomyces blakesleeanus grown in the presence of methionine-[methyl-²H₃], each contained two deuterium atoms; lanosterol, however, was unlabelled. The ¹⁴C:³H atomic ratio of the following sterols isolated from P. blakesleeanus grown in the presence of mevalonic acid-[2-¹⁴C,(4R)-4-³H₁], was: ergosterol, 5:3; episterol, 5:4; ergosta-5,7,24(28)-trien-3β-ol, 5:3; 4α-methyl-5α-ergosta-8,24(28)-dien-3β-ol, 5:4; 24-methylene-24,25-dihydrolanosterol, 6:5; lanosterol, 6:5. The significance of these results in terms of ergosterol biosynthesis is discussed.     

Sterol biosynthesis via cycloartenol in Saprolegnia

The Oomycete Saprolegnia ferax incorporates ³H from both cycloartenol-[2-³H] and lanosterol-[2-³H] into its normal sterols cholesterol, fucosterol, desmosterol, and 24-methylenecholesterol. It is concluded that sterol biosynthesis in this organism is via cycloartenol and the taxonomic implications are discussed.     

Dehydrodinosterol,dinosterone and related sterols of a non-photosynthetic dinoflagellate, Crypthecodinium cohnii

The heterotrophic dinoflagellate Crypthecodinium cohnii contained the 4α-methyl sterols, dinosterol, dehydrodinosterol (4α,23,24-trimethylcholesta-5,22-dien-3β-ol) and the tentatively identified 4α,24-dimethyl-cholestan-3β-ol and 4α,24-dimethylcholest-5-en-3β-ol. The major 4-demethyl sterol was cholesta-5,7-dien-3β-ol which was accompanied by a smaller amount of cholesterol and traces of several other C₂₇,C₂₈ and C₂₉ sterols. In addition, a 3-oxo-steroid fraction was isolated and the major component identified as dinosterone (4α,23,24-trimethylcholest-22-en-3-one). The possible biosynthetic relationships of these compounds are discussed.     

The structure of elasterol from Ecballium elaterium

The structure of elasterol, the major sterol of Ecballium elaterium, is revised to (24S)-24-ethyl-5α-cholesta- 7,22,25-trien-3β-ol from 24-ethyl-5α-cholesta-7,16,25-trien-3β-ol.     

Major sterols of Achlya bisexualis

Fucosterol, 24-methylene cholesterol, and cholesterol have been identified as major sterols in Achlya bisexualis. 7-Dehydrofucosterol has also been tentatively identified.     

Evidence for separate intermediates in the biosynthesis of 24α- and 24β-alkylsterols in tracheophytes

When mevalonate-[2-¹⁴C] was incubated with seeds of Pinus pinea, 23% of the label in sterols was found in trans-24-ethylidenecholesterol, 12% in a mixture of 24α- and 24β-methylcholesterol, and 65% in 24α-ethylcholesterol. However, when the radioactive substrate was lanosterol-[24-³H], label appeared only in the 24-ethylidene- (85%) and the epimeric 24-methylsterols (15%). From the ratios of labels in the ethylidene- and methyl-sterols it was possible to show that the tritium in the 24-C₁ -mixture was incorporated only into the 24β-methyl epimer. The labelling patterns are consistent with a pathway to 24β-alkylsterols via Δ²⁵⁽²⁷⁾-sterols bypassing 24-ethylidenesterols and to 24α-alkylsterols via Δ²⁴⁽²⁸⁾-sterols which are isomerized to Δ²⁴⁽²⁵⁾-sterols prior to reduction.     

Biosynthesis of steroidal withanolides in Withania somnifera

Administration of 24-methylene-cholesterol-[28-³H] to Withania somnifera, yielded [³H] radioactivity in the isolated withaferin A and withanolide D, whereas administered 24-(R,S)-methyl-cholesterol-[28-³H] was not incorporated into these compounds. 24-Methylene-cholesterol is, therefore, proposed as a sterol precursor of the withanolides. A novel procedure is described for the isolation of withanolides from W. somnifera. This method in conjunction with an improved procedure for administration of labelled sterols and mevalonolactone produces a greatly increased yield of labelled withanolides.     

Terpenoids of Rhododendron japonicum

From Rhododendron japonicum were isolated two diterpene glucosides, pieroside B and grayanoside B, and two new triterpenes, 2α,3α,24-trihydroxy     

Triterpenes of Lithocarpus species

The triterpenes of the five Lithocarpus species examined comprised members of the friedo- and unrearranged oleanane groups, viz. friedelin, friedelan-3β-ol, taraxerol and β-amyrin. Glutinol was also present, except in L. harlandi where friedelan-2α,3β-diol was found. In addition, three new cycloartane triterpenes, lithocarpolone (21,24-epoxy-24-hydroxymethyl-cycloartan-3-one), lithocarpdiol (21,24-epoxy-24-hydroxymethyl-cycloartan-3β-ol). and 24-methylenecycloartan-3β,21-diol were found in L. polystachya, and their structures determined.     

Sterols of roots and nitrogen fixing root nodules of vicia faba

Sterols of both roots and nodules of Vicia faba were found to be mainly 24-methylcholesterol, 24-ethyl-5,22-cholestadien-3β-ol and 24-ethylcholesterol. Nodules contained nearly twice the quantity of sterols present in roots, mainly due to an increase in 24-ethylcholesterol from about 40–60% of the total sterol content of root extract to over 80% of nodule extracts. Insignificant amounts of sterol were detected in nodule bacteroid fractions and in free-living Rhizobium leguminosarum.     

Sterols of the lichen Pseudevernia furfuracea

A mixture of C₂₇, C₂₈ and C₂₉ sterols was isolated from the lichen Pseudevernia furfuracea and characterized by means of GLC and MS. Mono-, di- and tri-unsaturated sterols were identified as well as a small amount of fully saturated sterols (stanols). Only a part of the total sterols present in the lichen tissue was easily extractable with organic solvents. Another portion was only obtained after saponification of the lichen residue remaining after extraction with organic solvents. The composition of these two fractions difrered considerably, the former contained predominantly 5a,8a-epidioxy-5a-ergosta-6,22-dien-3β-ol (ergosterol peroxide) and 24-ethylcholesta-5,22-dien-3β-ol while in the latter 24-ethylcholesta-5,22-dien- 3β-ol and C₂₈ triene sterols were the main components.     

Dammarane saponins of leaves of Panax pseudo-ginseng subsp. himalaicus

From dried leaves of Panax pseudo-ginseng subsp. himalaicus collected in Eastern Himalaya, new dammarane saponins, named pseudo-ginsenosides-F₁₁ and -F₈ were isolated along with the known Ginseng-root saponins, ginsenosides-Rb₃, Rd and -Re. Pseudo-ginsenoside-F₈ was proved to be a mono-acetyl-ginsenoside-Rb₃ and the location of its acetyl group was established mainly by ¹³C NMR spectroscopy. Pseudo-ginsenoside-F₁₁, was identified as the 6-O-α-rhamnopyransyl(1 → 2)-β-glucopyranoside of 3β,6α,12β,25-tetrahydoxy-(20S,24R)-epoxy-dammarane. The C-24 configuration of ocotillone and its related triterpenes was confirmed to be 24R excluding the recent comment by Lavie et al.     

The occurrence of C28 sterols in red algae

Lenormandia prolifera and Amansia glomerata contain large amounts of 24-methylenecholesterol. These two algae belong to the Amansia group of Rhodomelaceae as do Rytiphloea tinctoria and Vidalia volubilis, which are already known to be able to alkylate sterols at C-24. Halopythis pinastroides is an exception in the Amansia group, no C-28 sterols being present in this alga.