Unusual lipid composition of a Bacillus sp. isolated from Lake Pomorie in Bulgaria

The lipid composition of a Bacillus sp., isolated from Lake Pomorie in Bulgaria, was unusual and consisted of 26 different fatty acids between C12 and C26, with anteiso C15-C17 saturated fatty acids predominating. The furan fatty acid, 10,13-epoxy-11-methyloctadeca-10,12-dienoic acid, was also identified, a new finding for this genus. The hydrocarbons consisted of 30 different monounsaturated hydrocarbons, between C25 and C30, with the iso-iso, iso-anteiso, anteiso-anteiso, iso-normal, and anteiso-normal methyl branching for odd-numbered chains, and the iso-iso, iso-anteiso, iso-normal, and anteiso-normal methyl branching for even-numbered chains. The double bond positions in these hydrocarbons were determined by dimethyl disulfide derivatization followed by GC-MS, and the double-bond cis configuration was confirmed by infrared spectroscopy. Some previously unknown hydrocarbons in bacteria, such as (Z)-3,21-dimethyl-9-tricosene, (Z)-3,21-dimethyl-10-tricosene, (Z)-2,24-dimethyl-11-pentacosene, and (Z)-2,25-dimethyl-13-hexacosene were identified. Sterols were detected and were based on the sitosterol nucleus.     

Cuticular Hydrocarbons of Buffalo Fly, <Emphasis Type="Italic">Haematobia exigua</Emphasis>, and Chemotaxonomic Differentiation from Horn Fly, <Emphasis Type="Italic">H. irritans</Emphasis>

We determined the quantity and chemical composition of cuticular hydrocarbons of different strains, sexes, and ages of buffalo flies, Haematobia exigua. The quantity of cuticular hydrocarbons increased from less than 1 μg/fly for newly emerged flies to over 11 μg/fly in 13-d-old flies. The hydrocarbon chain length varied from C₂₁ to C₂₉, with unbranched alkanes and monounsaturated alkenes the major components. Newly emerged flies contained almost exclusively C₂₇ hydrocarbons. Increasing age was accompanied by the appearance of hydrocarbons with shorter carbon chains and an increase in the proportion of alkenes. 11-Tricosene and 7-tricosene were the most abundant hydrocarbons in mature H. exigua. Cuticular hydrocarbons of H. exigua are distinctly different from those of horn flies, Haematobia irritans. The most noticeable differences were in the C₂₃ alkenes, with the major isomers 11- and 7-tricosene in H. exigua and (Z)-9- and (Z)-5-tricosene in H. irritans, respectively. Cuticular hydrocarbon analysis provides a reliable method to differentiate the two species, which are morphologically difficult to separate. The differences in cuticular hydrocarbons also support their recognition as separate species, H. exigua and H. irritans, rather than as subspecies.     

Characterization of branched monounsaturated hydrocarbons ofSarcina lutea andSarcina flava

The exact position of the double bond and the probable branch assignments of each hydrocarbon isomer ofSarcina lutea andSarcina flava were determined by derivatives and gas chromatography and combined gas chromatography-mass spectrometry. It was shown by mass spectral data that in all isomers the double bond position was in or near the center of the molecule and, that some of the resolved isomers contained an additional positional isomer. The hydrocarbons were identified as containing methyl branches in the iso or anteiso or both configurations, symmetrically and asymmetrically disposed on the ends of the isomers. The assignment of the methyl branches for each tetrad of four isomers as the isomers emerged from the gas chromatographic column was iso-iso’; anteiso-iso; anteiso-anteiso’; and isonormal for odd carbon-numbered chains, and iso-iso’; anteiso-iso; iso-normal; and anteiso-normal for even carbon-numbered chains. A fifth isomer was identified as a normal-olefin.     

The structures of the branched fatty acids in the wax esters of vernix caseosa

By combined gas liquid chromatography-mass spectrometry a series of monomethyl branched fatty acids was found in the fatty acid moiety of the wax esters of vernix caseosa. The methyl branch occurred on the even C-atoms of chains ranging from C₁₁ to C₁₇ (some 43 compounds in all). Except for the iso acids and possibly some of the anteiso acids, these could be formed by replacement of malonyl CoA with a molecule of methyl malonyl CoA at the point of the branch. Smaller amounts of fatty acids also were found with two methyl branches occurring on the even C-atoms of chains ranging from C₉ to C₁₅.     

Peroxidative formation of C3-hydrocarbons from an ω-4 polyunsaturated fatty acid(16∶3ω4) in the algaBumilleriopsis

Under peroxidative conditions (i.e., illumination in the presence of Cu²⁺ or ap-nitro diphenylether herbicide), the xanthophycean microalga,Bumilleriopsis filiformis, evolves C₂ and C₅ hydrocarbons besides substantial amounts of propane and propene. Fatty acids were separated as methyl esters by argentation and reversed-phase thin layer chromatography and the fractions subsequently peroxidized by illuminated and copper-supplementedAnacystis thylakoids. These membranes do not contain polyunsaturated fatty acids and are, therefore, unable to evolve volatile hydrocarbons by itself. The C₂ and C₅ hydrocarbons formed by the fractions added match with their content of ω-3 and ω-6 fattyacid species having 2–4 double bonds. The fractions yielding C₃ hydrocarbons contain a fatty acid hitherto unknown forBumilleriopsis, which was isolated and identified as 16∶3ω4.     

Characterization of skin-surface lipids from the monkey (Macaca fascicularis)

Skin-surface lipids from the monkeyMacaca fascicularis are composed of sterol esters (38%), cholesterol (4%) and two types of wax diesters, identified as Type II (IIa and IIb, 17% and 40%, respectively). Type IIa contained diesters of 1,2-alkanediols esterified with two molecules of long-chain (C₁₄−C₃₄) fatty acids having straight and branched chains. In the diesters IIa, fatty acids shorter than C₁₉ predominated in position 1, and fatty acids longer than C₂₀ predominated in position 2. Type IIb contained diesters of 1,2-alkanediols esterified with C₄ and C₅ branched-chain fatty acids (predominantly isovaleric acid) at position 1 and long-chain (C₁₄−C₂₇) acids, having straight and branched chains, at position 2. The shortchain acids were converted to 2-nitrophenylhydrazides and analyzed by high-performance liquid chromatography (HPLC). Ammonia chemical ionization (CI)-gas chromatography (GC)-mass spectrometry (MS) resolved the intact diesters IIb into 12 peaks corresponding to molecular weights ranging from 597 to 748, and showed that the molecular species, such as C₂₁−C₁₆−C₅ (diol, fatty acid in position 2, fatty acid in position 1), C₂₂−C₁₆−C₅ and C₂₃−C₁₆−C₅, were prevalent. The fatty acids from both diesters were mostly (>98%) saturated. The 1,2-alkanediols from both diesters consisted of C₁₆−C₂₆ saturated straight- and branched-chain components. The acyl groups of sterol esters contained 86% C₁₄−C₃₄ branched-chain acids. The unsaturated fatty acids (5.4%) belonged to a straight-chain monoenoic series having extremely long chains (C₁₈−C₃₄). The branched-chain structures in the fatty acids and diols were iso and anteiso. These results show the species-specific profile for the skin-surface lipid synthesis.     

Chromatograpahic Analysis of Eriobotrya japonica Fat

Thin layer chromatography and infrared spectophotometry of the fat of Eriobotrya japonica shows the presence of hydrocarbons (3.1%), was esters (5.3%), triglycerides (786%) and the last polar fraction containing fatty acids, colouring matter etc. (13.0%). After the saponification of the fat, the fatty acids C₁₂-C₂₄ and fatty alcohols C₁₂-C₂₆ determined first time as methyl esters and acetates respectively were identified by the application of gas liquid chromatography.     

Volatile hydrocarbons in the dufour's gland of the parasiteNemeritis canescens (Grav.) (Hymenoptera; ichneumonidae)

The Dufour's gland of the parasitic waspNemeritis (= Venturia) canescens (Grav.) was shown by gas chromatography and mass spectrometry to contain a mixture of C₂₁, C₂₃, and C₂₅ saturated and monounsaturated hydrocarbons. The main component (62%) was (Z)-10-tricosene. The biological activity of the components is discussed.     

Cuticular lipids of insects: II. Hydrocarbons of the cockroachesPeriplaneta australasiae,Periplaneta brunnea andPeriplaneta fuliginosa

The major cuticular hydrocarbons of the cockroachesPeriplaneta australasiae, Periplaneta brunnea andPeriplaneta fuliginosa aren-tricosane,cis-9-tricosene, 3-methyltricosane, 11-methyltricosane and 13-methylpentacosane. There are as yet unexplained quantitative differences between the hydrocarbon compositions of males and females ofP. australasiae andP. fuliginosa, cis-9-tricosene being a major hydrocarbon of the males only. A series of mono-methyl internally branched hydrocarbons ranging in chain length from 23 to 26 carbons with the methyl branch on the 13th carbon from one end was observed. Minor quantities of other hydrocarbons have been identified.     

Some monomethyl-branched fatty acids from ruminant fats: Open-tubular GLC separations and indications of substitution on even-numbered carbon

Isomeric methyl esters of fatty acids in three groups (C₁₅, C₁₇, C₁₉) have been isolated from ruminant fats. Basic structural analysis by physiochemical techniques indicated that these odd-numbered fatty acids were even chain with a single methyl branch on the chain. High resolution open-tubular gas liquid chromatographic studies indicate that, with the exception of iso acid impurities in these fractions, only even-numbered carbons of the fatty acid chains bear the methyl branch.     

Isolation and identification of constituents of Jasminum auriculatum (Vahl) leaves

Jasminum auriculatum (Vahl) (family: Malvaceae) widely grown in India was analysed for its fatty acids and waxy constituents. Straight-chain hydrocarbons (C₂₀-C₃₄), fatty acids (C₁₄-C₂₃) and fatty alcohols (C₂₁-C₃₂) were found in the 95% aqueous ethanol extract of the leaves of the plant. Malvalic acid was the only cyclic acid identified. Hydrocarbons (C₂₉ and C₃₁), fatty acids (C₁₆, C₁₈, C_18:1, C_18:2, C_18:3 and C₂₂) and fatty alcohols (iso-C₂₆, C₂₈ and C₃₀) were the major components. Four polyalcohols, namely D-mannitol, xylitol, inositol and sorbitol, have also been found in the alcoholic extract of the leaves.     

Sponge fatty acids. 3. Occurrence of series of n−7 monoenoic andiso-5,9 dienoic long-chain fatty acids in the phospholipids of the marine spongeCinachyrella aff.schulzei keller

The fatty acid composition of phospholipids from the New Caledonian spongeCinachyrella aff.schulzei Keller was studied. More than 60 fatty acids were identified as methyl esters andN-acyl pyrrolidides by gas chromatography and gas chromatography/mass spectrometry. Two isoprenoid fatty acids also were shown to be present, namely 4,8,12-trimethyltridecanoic and 5,9,13-trimethyltetradecanoic acids. The unusual 6-tetradecenoic, 6-pentadecenoic, 12-nonadecenoic and 26-methylheptacosanoic (iso-28∶0) acids were found for the first time in sponge phospholipids. A series of six n−7 monoenoic long-chain fatty acids (C₂₃ to C₂₈) were identified, including the rare 16-tricosenoic, 18-pentacosenoic and 21-octacosenoic acids. Fifteen fatty acids possessing the typical 5,9 dienoic moiety accounted for 30% of the total fatty acid mixture. Two new fatty acids were identified, namely 5(Z)-octacosenoic and 27-methyl-5(Z),9(Z)-octacosadienoic (iso-5,9-29∶2). Based on gas chromatography/Fourier transform infrared experiments, the double bonds were assigned the (Z) configuration. For part 2 of this series, see Reference 1.     

Mass spectrometric analysis of the fatty acids and nonsaponifiable lipids ofEimeria tenella oocysts

The fatty acids and nonsaponifiable lipids ofEimeria tenella oocysts were analyzed by gas liquid chromatography and combined gas liquid chromatographymass spectrometry. The fatty acids detected were identified as C_14∶0, C_16∶0, C_16∶1, C_18∶0, C_18∶1, and C_18∶2. Though the wt of the fatty acid fraction decreased during sporulation from 91 μg per 10⁶ oocysts to 47 μg per 10⁶ oocysts, the relative amounts of these fatty acids did not change appreciably. The nonsaponifiable lipids ofE. tenella consisted of cholesterol and unbranched primary alcohols of 22, 24, 26, 28, 30, and 32 carbons. Mass fragmentography demonstrated that each species of alcohol consisted of saturated and monounsaturated derivatives. Trimethylsilyl ethers of fatty alcohols were found to offer several important advantages over free alcohols for mass spectrometric characterization. Before sporulation, most fatty alcohols were in the oocyst wall. During sporulation, the wt of the nonsaponifiable lipids increased from 16 μg per 10⁶ oocysts of 44 μg per 10⁶ oocysts due largely to synthesis of C₂₄ and C₂₆ alcohols. The newly synthesized fatty alcohols were not deposited in the oocyst wall.     

Composition of the silk lipids of the spider Nephila clavipes

A detailed analysis of the lipids of spider silk is given for the first time. Extracts of the silk from the golden orb weaver, Nephila clavipes, were studied by gas chromatography, mass spectrometry, and chemical derivatizations. The major group of the lipids consisted of methyl-branched 1-methoxyalkanes (methyl ethers) with up to four methyl groups in the chain (chain length between C₂₈ and C₃₄), which are unique to spiders. The position of the methyl branches was determined by conversion into cyanides, which allowed easy location of methyl branches. The second-largest group included alkanes with a wide structural variety; 2-methyl-branched, even-numbered hydrocarbons predominated. A general numerical method for the estimation of retention indices of alkanes and their derivatives is presented. Further components of the web included alkanols and alkanediols, fatty acids, and glyceryl ethers. Some comments on the biosynthesis of these compounds are also given.     

Phospholpid studies of marine organisms: 2.1 Phospholipids,phospholipid-bound fatty acids and free sterols of the spongeAplysina fistularis (Pallas) formafulva (Pallas) (=Verongia thiona)2. Isolation and structure elucidation of unprecedented branched fatty acids

The free sterols and phospholipids of the demospongeAplysina fistularis were isolated and analyzed. The free sterols consisted mainly of the unusual 26-methylated sterols aplysterol (53%) and 24(28)-dehydroaplysterol (7%) together with 7 commonly occurring sterods. The major phospholipids were phosphatidylcholine, phosphatidylglycerol, phosphatidylinositol, phosphatidylethanolamine, phosphatidylserine and diphosphatidylglycerol. The major fatty acyl components of the phospholipids consisted of 85% C₁₄−C₂₀ acids, including the unprecedented 2,6,10-trimethyl-5-tetradecenoic acid and 11-methyloctadecanoic acid. The remaining 15% were C₂₇−C₃₀ demospongic acids, including 2 novel acids tentatively assigned the structures 5,9,23-octacosatrienoic acid and 5,9,23-nonacosatrienoic acid, and 3 novel acids proven to be 5,9,21-octacosatrienoic acid, Z,Z-20-methyl-5,9-hexacosadienoic acid and Z,Z-22-methyl-5,9-octacosadienoic acid. The biosyntheses of the novel demospongic acids are proposed to occur by chain elongation of monoenoic or branched precursors followed by desaturation. The large quantities of typically bacterial phospholipids and fatty acids found implied the presence of bacteria in the sponge, in agreement with microscopic studies. Analysis of the phospholipid-bound fatty acids in a sponge cell-enriched fraction indicated that the demospongic acids, including the 2 branched structures, were the major acids of the sponge cells. The presence inA. fistularis of demospongic acids containing membrane disordering groups—methyl branches or double bonds—on the ω7 carbon is proposed to be due to the need by the sponge for membranes possessing fluidity near the middle of the phospholipid bilayer. It is also proposed that the C₂₆ methyl group of aplysterol causes disordering of the phospholipid bilayer in the same region, and thus also evolved in response to this need. For part 1, see ref. 1. Phylum porifera, class demospongia, subclass ceractinomorpha, order verongida, family aplysinidae (2). Formerly known asVerongia thiona de Laubenfels. Major diagnostic fragments from peak 23:M⁺ 471 (2.9%); m/z 442 (C₂₆, 0.2%); m/z 428 (C₂₅, 0.2%); m/z 414 (C₂₄, 0.1%); m/z 400 (C₂₃, 0.3%); (absence of C₂₂ peak); m/z 374 (C₂₁, 0.2%); m/z 360 (C₂₀, 0.2%); m/z 234 (C₁₁, 1.0%); m/z 220 (C₁₀, 0.7%); m/z 206 (C₉, 0.5%); m/z 194 (C₈, 0.4%); m/z 180 (C₇, 24.9%); m/z 166 (C₆, 1.2%); m/z 152 (C₅, 0.8%); m/z 140 (C₄, 1.3%); m/z 126 (C₃, 18.7%). Major diagnostic fragments from peak 25:M⁺ 485 (0.9%); m/z 456 (C₂₇, 0.1%); (absence of C₂₆, C₂₅ peaks); m/z 414 (C₂₄, 0.3%); m/z 400 (C₂₃, 0.1%); m/z 402 (C₂₃, 0.1%); (absence of C₂₂ peak); m/z 374 (C₂₁, 0.1%); m/z 360 (C₂₀, 0.1%); m/z 234 (C₁₁, 0.5%); m/z 220 (C₁₀, 0.3%); m/z 206 (C₉, 0.1%); m/z 194 (C₈, 0.2%); m/z 180 (C₇, 18.0%); m/z 166 (C₆); m/z 152 (C₅), m/z 140 (C₄); m/z 126 (C₃, 15.0%).     

Studies of jasmine wax: I. Physical properties and chemical composition

Jasmine wax was fractionated using thin layer chromatography. Six fractions were isolated, purified and identified using combined gas liquid chromatography and mass spectrometry. Infrared spectra were adopted for the chemical structure confirmation of each fraction. The fractions were 49.44% hydrocarbons, 15.73% esters, 3.49% aldehydes and ketones, 7.91% primary fatty alcohols, 8.31% free fatty acids and 8.99% was considered origin. The hydrocarbon fraction was found to consist mainly of saturated chains with C₂₉, C₃₁ and C₂₇ the most predominant. The aldehyde fraction was characterized by straight chains with even numbers of carbon atoms ranging between C₁₀ and C₂₈, whereas the ketone fraction was found to contain a longer chain structure, i.e., C₃₆. However, the C = O was found to be at the C₈ and C₁₁ positions. Free primary alcohols were found to occur in short chains, indicating their existence in the liquid phase, whereas the free fatty acids were found in long chains and were more likely in the solid phase.     

Identification of novel octadecadienoic fatty acids in the seaweedCladophora rupestris through oxidative ozonolysis of the alcohols prepared from the acids

The BF₃−MeOH reagent for ozonolysis of ethylenic unsaturation does not oxidize alcohols. It is therefore feasible to determine the position of ethylenic unsaturation in long chain fatty alcohols of synthetic or natural origin by recovering the methyl ester products intact and silylating the alcohol function of half-ester, half-alcohol, products prior to gas liquid chromatographic analysis. The C₃ fragment from methylene-interrupted alkyl chains is not recovered, but, by first reducing carboxyl ester groups to alcohols, the terminal difunctional products can be identified in nonmethylene-interrupted dienoic fatty acids. The seaweedCladophora rupestris is shown to contain Δ5,Δ11-,Δ8,Δ11-, and Δ11, Δ14- as well as Δ9,Δ12-octadecadienoic acid.     

Cuticular lipids of insects: VIII. Alkanes of the mormon cricketAnabrus simplex

The cuticular hydrocarbons of the Mormon cricket,Anabrus simplex, are all saturated and consist of n-alkanes (29+%), 3-methylalkanes (12%), internally branched monomethylalkanes (26+%), and dimethylalkanes (28+%). The principal n-alkane is the C₂₉ component, with a range from C₂₃ to C₃₃. The 3-methylalkanes range from C₂₈ to C₃₂, and the internally branched monomethyl-and dimethylalkanes range from C₂₉ to C₃₉. When the branched alkanes ofA. simplex are compared to those from other insects in the order Orthoptera, interesting patterns of methyl branching are observed. Scientific Journal Series 631, Agricultural Experiment Station, University of Montana, Bozeman, Montana 59715.     

Metabolically blocked analogs of housefly sex pheromone: II. Metabolism studies

Analogs of (Z)-9-tricosene (Z9–23: Hy) bearing methyl substituents, cyclopropyl groups, fluorine substituents, and additional double bonds were used to probe the substrate requirements for the monooxygenase system that converts Z9–23: Hy to the corresponding epoxide and ketone. Three of the seven analogs tested, 10-fluoro-(Z)-14-tricosene, 10,10-difluoro-(Z)-14-tricosene, and 14-methyl-(Z)-9-tricosene, were metabolized to the corresponding epoxide. Compounds with two methyl groups, a cyclopropane group, a hydroxy group, or an additional double bond at the 14 position were not epoxidized at the 9,10 position. This suggests that only minimal structural change at the 14-position of Z9–23: Hy is allowed with retention of metabolic activity. None of the analogs tested were hydroxylated at the position equivalent to the 14 position of Z9–23:Hy. Of the 13 analogs tested as inhibitors of Z9–23:Hy metabolism, the two compounds that were the most effective inhibitors in both male and female houseflies were (Z)-14-tricosen-10-one and 1-nonyl-1-[(Z)-4-tetradecen-1-yl]-cyclopropane. These data show that the poly substrate monooxy genase that metabolizes Z9–23: Hy in the housefly has very strict structural requirements for the substrate.     

Phospholipid studies of marine organisms: New branched fatty acids fromStrongylophora durissima

The phospholipids of the spongeStrongylophora durissima were analyzed. The major phospholipids present were phosphatidylethanolamine (PE), phosphatidylcholine (PC), phosphatidylserine (PS), phosphatidylglycerol (PG) and phosphatidylinositol (PI). The major fatty acid components of the phospholipids consisted of short chain (C₁₄−C₁₉) and very long chain (C₂₅−C₃₀) “Demospongic” acids. Three novel branched Δ⁵ monounsaturated acids,Z-19-methyl-5-pentacosenoic,Z-19-methyl-5-hexacosenoic andZ-19-methyl-5-heptacosenoic acids were encountered in the sponge. The 3-saturated counterparts of these compounds, 19-methylpentacosanoic, 19-methylhexacosanoic and 19-methylheptacosanoic acids, as well as 19-methylpentacosanoic and 20-methyloctacosanoic acids also are hitherto undescribed acids present in the sponge. Trace amounts of 2 very long chain acids also were detected and their structures tentatively assigned as 19,21-dimethylheptacosanoic and 20,22-dimethyloctacosanoic acids. The distribution of these fatty acids according to phospholipid head groups also was described.