柑橘真棉蚧蜡泌物的超微形态、红外光谱与化学成分特征

采用扫描电镜、红外光谱、气相色谱/质谱联用技术研究了柑橘真棉蚧Eupulvinariacitricola(Kuwana)(半翅目:蚧科)蜡泌物的超微形态、红外光谱和卵囊蜡质的化学成分特征。结果发现:该蚧雌虫背面腺体分泌湿蜡,在体表面形成薄的半透明蜡壳,背面蜡壳分为6个小区,表面由湿蜡凝结成片状和蜡块状构造。雌成虫产卵期由腹面多格腺分泌丝状蜡结成卵囊。雄若虫也分泌湿蜡,雄蛹的蜡茧薄,表面有浅的凹凸花纹。用雌成虫卵囊、背面蜡壳和雄茧的蜡质作红外光谱分析,它们的谱图具有共同的基本特征,吸收峰群分为5个区域:2900cm-1附近的3个强吸收峰组成的峰群是>CH2和—CH3的特征吸收峰;1800cm-1~1500cm-1之间的吸收峰群是羰基(>C=O)和C=C键的伸缩振动;1500cm-1~1400cm-1区域是>CH2和—CH3基团的吸收峰;1400cm-1~1000cm-1之间为饱和碳链—C—C—C—骨架振动。730cm-1附近为环状化合物的基团吸收峰群。由此确定它们的组成都是长碳链结构的脂肪酸、脂肪醇、烃类、酯类或带有环状结构的化合物。其主要区别在第2、3、4区吸收峰的形式、强度和数量,反映出基团的种类和数量不同。雌成虫卵囊蜡质甲酯化后GC/MS检测出7个组分(表1),全部为长链脂肪酸。未经甲酯化的蜡质检测出4类16个组分(表2),可分为长链烃、酸、醇、酯。第一类为饱和长链烃,5个组分,占总组分的38.38%;第二类是长链脂肪酸,包括直链饱和酸和不饱和酸共6个组分,占31.59%;第三类是直链饱和醇,占2.87%;第四类是酯类化合物,4个组分,占总组分的27.16%。     

In vitro synthesis of mycolic acids by the fluffy layer fraction of Bacterionema matruchotii

Biosynthetic activity for mycolic acid occurred in the fluffy layer fraction but not in the 5000g supernatant of Bacterionema matruchotii. With [1-¹⁴C]palmitic acid as precursor for the in vitro system, the predominant product was identified as C_32:0 mycolic acid by radio-gas-liquid chromatographie (radio-GLC) and gas chromatographic/mass spectroscopic analyses; if [1-¹⁴C]stearic acid was used, two major radioactive peaks appeared on GLC: one corresponding to the peak of (C_34:0 + C_34:1) mycolic acids and the other to (C_36:0 + C_36:1) mycolic acids. By pyrolysis/radio-GLC analysis, C_32:0 mycolic acid synthesized by [1-¹⁴C]palmitic acid was pyrolyzed at 300 °C to form palmitaldehyde (the mero moiety) and methyl palmitate (the branch moiety). The pH optimum for the incorporation of [1-¹⁴C]palmitate into bacterionema mycolic acids was 6.4 and the reaction required a divalent cation. The in vitro system utilized myristic, palmitic, stearic and oleic acids (probably via their activated forms) well as precursors, among which myristic and palmitic acids were more effective than the rest. Avidin showed no effect on the biosynthesis of mycolic acid from ¹⁴C-palmitate whereas cerulenin, a specific inhibitor of β-ketoacyl synthetase in de novo fatty acid synthesis, inhibited the reaction at a relatively higher concentration. Thin-layer chromatographic analysis of lipids extracted from the reacting mixture without alkaline hydrolysis showed that both exogenous [1-¹⁴] fatty acid and synthesized mycolic acids were bound to an unknown compound by an alkali-labile linkage and this association seemed to occur prior to the condensation of two molecules of fatty acid.     

Ultralong C100 mycolic acids support the assignment of Segniliparus as a new bacterial genus

Mycolic acid-producing bacteria isolated from the respiratory tract of human and non-human mammals were recently assigned as a distinct genus, Segniliparus, because they diverge from rhodococci and mycobacteria in genetic and chemical features. Using high accuracy mass spectrometry, we determined the chemical composition of 65 homologous mycolic acids in two Segniliparus species and separately analyzed the three subclasses to measure relative chain length, number and stereochemistry of unsaturations and cyclopropyl groups within each class. Whereas mycobacterial mycolate subclasses are distinguished from one another by R groups on the meromycolate chain, Segniliparus species synthesize solely non-oxygenated α-mycolates with high levels of cis unsaturation. Unexpectedly Segniliparus α-mycolates diverge into three subclasses based on large differences in carbon chain length with one bacterial culture producing mycolates that range from C58 to C100. Both the overall chain length (C100) and the chain length diversity (C42) are larger than previously seen for mycolic acid-producing organisms and provide direct chemical evidence for assignment of Segniliparus as a distinct genus. Yet, electron microscopy shows that the long and diverse mycolates pack into a typical appearing membrane. Therefore, these new and unexpected extremes of mycolic acid chemical structure raise questions about the modes of mycolic acid packing and folding into a membrane.     

Purification and structure analysis of mycolic acids in <Emphasis Type="Italic">Corynebacterium glutamicum</Emphasis>

Corynebacterium glutamicum is widely used for producing amino acids. Mycolic acids, the major components in the cell wall of C. glutamicum might be closely related to the secretion of amino acids. In this study, mycolic acids were extracted from 5 strains of C. glutamicum, including ATCC 13032, ATCC 13869, ATCC 14067, L-isoleucine producing strain IWJ-1, and L-valine producing strain VWJ-1. Structures of these mycolic acids were analyzed using thin layer chromatography and electrospray ionization mass spectrometry. More than twenty molecular species of mycolic acid were observed in all 5 strains. They differ in the length (20–40 carbons) and saturation (0–3 double bonds) of their constituent fatty acids. The dominant species of mycolic acid in every strain was different, but their two hydrocarbon chains were similar in length (14–18 carbons), and the meromycolate chain usually contained double bonds. As the growth temperature of cells increased from 30°C to 34°C, the proportion of mycolic acid species containing unsaturated and shorter hydrocarbon chains increased. These results provide new information on mycolic acids in C. glutamicum, and could be useful for modifying the cell wall to increase the production of amino acids.     

Thermally adaptive changes of mycolic acids in Mycobacterium smegmatis

The effect of growth temperature on mycolic acid composition in eight strains of Mycobacterium smegmatis was investigated by gas chromatography/mass spectrometry. A change in growth temperature from 45 to 20 degrees C caused a shift in the subclass and molecular species composition of mycolic acids. The relative amount of alpha'-mycolic acids to alpha-mycolic acids decreased, and that of hydroxy mycolic acids increased at lower temperatures. Moreover, the proportion of shorter-chain species of alpha-mycolic acids increased, and those of longer-chain species of alpha-mycolic and hydroxy mycolic acids decreased. This observation seems to be due to the changes of the chain length of meromycolates because the alpha-alkyl chain unit of mycolic acids was not affected. The ratio of odd to even carbon-numbered alpha-mycolates decreased as the growth temperature was lowered. In contrast, the molecular species composition of alpha'-mycolic acid was not influenced by the growth temperature.     

Characterization of anguibactin, a novel siderophore from Vibrio anguillarum 775(pJM1).

Anguibactin, a siderophore produced by cells of Vibrio anguillarum 775 harboring the pJM1 plasmid, has now been isolated from the supernatants of iron-deficient cultures. This iron-reactive material was purified by adsorption onto an XAD-7 resin and subsequent gel filtration on a Sephadex LH-20 column. The resulting neutral compound produced an ion at m/z 348 in mass spectrometry and contained one sulfur, four oxygen, and four nitrogen atoms as determined by elemental analysis. Its strong UV absorbance and blue fluorescence were suggestive of a phenolic moiety. In colorimetric reactions anguibactin behaved like a catechol. The catechol assignment was supported by the appearance of a new absorption band at 510 nm in the ferric complex and by the appearance of peaks at 1,367, 1,447, 1,469, and 1,538 cm-1 in the resonance Raman spectrum. In addition, the infrared spectrum gave evidence of a secondary amide function, but no free carboxylic acid or hydroxamic acid groups were observed. A third iron-ligating group was suggested by the liberation of three protons during iron binding; mass spectrometry of the resulting material yielded a molecular ion characteristic of a 1:1 complex of ferric anguibactin. The purified anguibactin exhibited specific growth-promoting activity under iron-limiting conditions for a siderophore-deficient mutant of V. anguillarum 775(pJM1). A novel structure for anguibactin was indicated by the failure of a large number of known siderophores and synthetic chelators to yield a similar type of specific cross-feeding in the V. anguillarum bioassay.     

藏药蕨麻多糖的光谱性质及单糖组成分析

本研究对藏药蕨麻多糖进行了分离提纯,并测定其水溶性多糖含量为99.4%;通过紫外光谱与红外光谱分析表明,蕨麻多糖为分子量较小的α-吡喃糖,并含有氨基糖;蕨麻多糖的水解单糖经过NMP衍生后进行毛细管电泳分析,测得其单糖组成为木糖、阿拉伯糖、葡萄糖、鼠李糖、甘露糖、岩藻糖、半乳糖、葡萄糖醛酸和半乳糖醛酸,含量分别为3.945、77.445、17.568、17.646、3.942、2.165、65.268、13.037μg/mg和33.484μg/mg,与GC-MS的定性分析结果一致。     

Mass spectrometry as a tool for identifying group D2 corynebacteria by their fatty acid profiles

Corynebacterium group D2 (CGD2) are lipophilic antibiotic-multiresistant bacteria involved in some infections of immunocompromised patients. The fatty acid composition and structure of different strains was established by several mass spectrometric methods, particularly negative ion tandem mass spectrometry coupled with capillary gas chromatography. Non-hydroxylated fatty acid profiles of three strains of CGD2 (ATCC 43042, ATCC 43043, ATCC 43044) were almost identical and revealed the presence of several straight chain unsaturated fatty acids from the omega-9 series, with even carbon numbers ranging from 14 to 24. Branched saturated fatty acids were mainly anteiso-heptadecanoic acid and tuberculostearic acid. Surprisingly, a relatively large quantity of 10-methylene octadecanoic acid was found. The non-hydroxylated fatty acid profile of one rare beta-lactam susceptible strain (SC1) was different; 10-methylene octadecanoic acid was lacking whereas tuberculostearic acid was much more abundant. In contrast, the four CGD2 strains displayed highly similar mycolic acid patterns. The major mycolic acid species corresponded to C32, C30 and C28 bis-unsaturated with a double bond on each branch at the omega-9 position. The comparison of the mycolic acid composition and structure with those of other medically important corynebacteria strains, revealed a characteristic pattern for CGD2 strains, and CGD2 strains were easily distinguished from Corynebacterium jeikeium (CIP 82.51).     

Comprehensive analysis of mycolic acid subclass and molecular species composition of Mycobacterium bovis BCG Tokyo 172 cell wall skeleton (SMP-105)

The mycobacterial cell envelope consists of a characteristic cell wall skeleton (CWS), a mycoloyl arabinogalactan peptidoglycan complex, and related hydrophobic components that contribute to the cell surface properties. Since mycolic acids have recently been reported to play crucial roles in host immune response, detailed molecular characterization of mycolic acid subclasses and sub-subclasses of CWS from Mycobacterium bovis BCG Tokyo 172 (SMP-105) was performed. Mycolic acids were liberated by alkali hydrolysis from SMP-105, and their methyl esters were separated by silica gel TLC into three subclasses: alpha-, methoxy-, and keto-mycolates. Each mycolate subclass was further separated by silver nitrate (AgNO(3))-coated silica gel TLC into sub-subclasses. Molecular weights of individual mycolic acid were determined by MALDI-TOF mass spectrometry. alpha-Mycolates were sub-grouped into cis, cis-dicyclopropanoic (alpha1), and cis-monocyclopropanoic-cis-monoenoic (alpha2) series; methoxy-mycolates were sub-grouped into cis-monocyclopropanoic (m1), trans-monocyclopropanoic (m2), trans-monoenoic (m3), cis-monocyclopropanoic-trans-monoenoic (m4), cis-monoenoic (m5), and cis-monocyclopropanoic-cis-monoenoic (m6) series; and keto-mycolates were sub-grouped into cis-monocyclopropanoic (k1), trans-monocyclopropanoic (k2), trans-monoenoic (k3), cis-monoenoic (k4), and cis-monocyclopropanoic-cis-monoenoic (k5) series. The position of each functional group, including cyclopropane rings and methoxy and keto groups, was determined by analysis of the meromycolates with fast atom bombardment (FAB) mass spectrometry and FAB mass-mass spectrometry, and the cis/trans ratio of cyclopropane rings and double bonds were determined by NMR analysis of methyl mycolates. Mycolic acid subclass and molecular species composition of SMP-105 showed characteristic features including newly-identified cis-monocyclopropanoic-trans-monoenoic mycolic acid (m4).     

Three types of mycolic acid from Mycobacterium tuberculosis Brévanne: implications for structure-function relationships in pathogenesis.

Saponification of the chloroform-soluble wax from Mycobacterium tuberculosis Brévanne led to the isolation of three classes of mycolic acid containing characteristic functional groups along the methylene backbone: type alpha (two cyclopropane rings); type beta (methoxyl, methyl, and cyclopropane); and type gamma (ketone, methyl, and cyclopropane). The structures of these acids were elucidated principally by mass spectrometry. The high mass region of the keto mycolate is presented showing the meromycolal and molecular ion regions. This is first time a molecular peak for this mycolic acid has been reported. The structure of the keto mycolate was further substantiated by study of the mass spectral fragmentation of its dithioketal derivative. Within each type of acid, a series of homologs was encountered, varying according to the number of methylene units in the backbone chain. Chromatographic and infrared spectrophotometric evidence is presented for the alkali-induced isomerization of the three types of mycolates.     

Aliphatic medium chain tricarboxylic acids in rat urine.

Three aliphatic tricarboxylic acids have been found in rat urine. They have been identified as 6-carboxy-5-undecenedioic acid, 6-carboxy-5-dodecenedoic acid, and 6-carboxy-5-tridecenedioic acid. The carbon skeleton structure was determined by mass spectra of the hydrogenated methyl esters. The double bond position was determined after osmium tetroxide oxidation followed by trifluoroacetylation and mass spectrometry and by infrared spectrometry. The compounds were present in the urine when the rats were fed on pellets but disappeared when they received sucrose and water. The acids were not present in the pellets, and a metabolic relation to compounds of longer chain length, possibly mycolic acids, is likely.     

Changes in molecular species composition of nocardomycolic acids in nocardia rubra by the growth temperature

Nocardomycolic acids from Nocardia rubra were fully separated and characterized by a combination of argentation thin-layer chromatography and gas chromatography — mass spectrometry (GCMS). The occurrence of 20 or more different molecular species of mycolic acids was demonstrated. GCMS analysis of each subclass of mycolic acids after separation on AgNO₃ thin-layer chromatography revealed that in general the major species consisted of the even-carbon mycolic acids ranging from C₃₈ to C₅₂. However, the most abundant species differed by the subclasses; C₄₄ being in saturated, C₄₆ in monoenoic and C₄₆ in dienoic mycolic acids, respectively. All these acids were shown to possess C₁₂ or C₁₄ alkyl branch at 2 position, while double bonds were located in longer straight chain alkyl unit.By using this method, distinctive changes in mycolic acid composition by growth temperature were observed. The ratios of saturated, monoenoic to dienoic mycolic acids in a mixture of certain carbon numbered mycolic acids varied greatly, according to the shift of growth temperature. The mass fragmentographic analysis, monitoring M-15 ions derived from the loss of methyl group from the molecular ions showed the lower temperature (15°C) grown cells contained more unsaturated (especially dienoic) mycolic acids, while the higher temperature (40°C) grown cells contained more saturated mycolic acids in both extractable and cell-wall bound lipids. These changes in mycolic acid composition occurred shortly after shifting up the growth temperature from 20°C to 43°C at a logarithmic stage of the bacterial growth.     

Characterization of mycolic acids from the pathogen Rhodococcus equi by tandem mass spectrometry with electrospray ionization

We describe a simple tandem mass spectrometric approach toward structural characterization of mycolic acids, the long-chain α-alkyl-β-hydroxy fatty acids unique to mycobacteria and related taxa. On collisionally activated dissociation in a linear ion trap or tandem quadrupole mass spectrometer, the [M−H]⁻ ions of mycolic acid generated by electrospray ionization undergo dissociation to eliminate the meroaldehyde residue, leading to formation of carboxylate anions containing α-alkyl chains. The structural information from these fragment ions affords structural assignment of the mycolic acids, including the lengths of the meromycolate chain and the α-branch. This study revealed that the mycolic acids isolated from pathogenic Rhodococcus equi 103 contained a series of homologous ions having C₃₀ to C₅₀ chain with 0–2 double bonds. The α-branch ranged from C₁₀ to C₁₈ with 0 to 1 double bond, in which 16:0 and 14:0 are the most prominent, whereas the meromycolate chain ranged from C₁₄ to C₃₄ with 0 to 2 double bonds. The major molecular species consisted of more than 3 isomers that differ by the lengths of the α-branch or meromycolate chain, and up to 10 isobaric isomers were identified for some minor ions. We also employed tandem quadrupole mass spectrometry with precursor ion and neutral loss scans for profiling mycolic acid with specific structure in mixtures. The tandem spectra obtained from precursor ion scans of m/z 255 (16:0-carboxylate anion) and m/z 227 (14:0-carboxylate anion) may provide a simple specific means for classification of Rhodococci species, whereas tandem spectra from neutral loss of meroaldehyde residue scans provided a simple approach to reveal the mycolic acid molecules with specific meromycolate chain in mixtures.     

Mass spectra of methyl-branched hydrocarbons from eggs of the tobacco hornworm

Hydrocarbons from three homologous series of branched alkanes from the eggs of the tobacco hornworm, Manduca sexta (L.), were identified by mass spectrometry. Gas-liquid chromatography (GLC) peaks 37-A (equivalent chain length of 37.2) and 39-A (equivalent chain length of 39.2) were mixtures of 13-, 15-, 17-, and 19-methylheptatriacontane and 13-, 15-, 17-, and 19-methylnonatriacontane, respectively. GLC peaks 33-B, 37-B, and 39-B with equivalent chain lengths of 33.4, 37.4, and 39.4, respectively, were mixtures of 13,17- and 15,19-dimethyltritriacontane, 13,17-, 15,19-, and 17,21-dimethylheptatriacontane, and 13,17-, 15,19-, and 17,21-dimethylnonatriacontane, respectively. GLC peak 37-C (equivalent chain length of 37.6) was a mixture of 11,15,19-, 13,17,21-, and 15,19,23-trimethylheptatriacontane.     

Isonicotinic acid hydrazide induced changes and inhibition in mycolic acid synthesis in Nocardia and related taxa

The mycolic acid compositions of Nocardia rubra and related bacteria grown in media containing different concentrations of antituberculous isonicotinic acid hydrazide (INH) were determined in detail by gas chromatography-mass spectrometry. On the basis of molecular species composition, average carbon numbers of mycolic acids were calculated. In Nocardia rubra, N. lutea and Rhodococcus rhodochrous IFO-13161, the ratio of mycolic to non-mycolic fatty acids and the average carbon numbers of mycolic acids were decreased at the INH concentrations of higher than 1 g/ml, paralleling with the significant inhibition of growth. In above three species the synthesis of longer chain mycolic acids (longer than C₄₄ or C₄₆) was inhibited more significantly than shorter homologues such as C₃₈ or C₄₀. In contrast, neither growth inhibition nor change in corynomycolic acid composition was observed in Corynebacteria xerosis and Rhodococcus rhodochrous IFO-13165 at the concentration region of INH up to 100 g/ml. The direct mass fragmentographic analysis of the trimethylsilylated (TMS) derivatives of mycolic acid methyl esters, monitoring [M-15] ions of individual molecular species, revealed that the chain shortening of total mycolic acid molecule by INH occurred more greatly in more highly unsaturated subclasses than in less unsaturated subclasses. Furthermore, mass fragmentographic analysis, monitoring fragment ions (A) and (B), due to straight chain and branched chain alkyl units, respectively, demonstrated the inhibition of mycolic acids was not attributed to the shortening of -alkyl chain, but to the inhibition of chain elongation of C₂₈ to C₃₂ straight chain meromycolic acids. It was also indicated the amounts of trehalose mono- and di-mycolate (cord factor) decreased significantly with the addition of INH (1 to 20 g/ml) in the above strains. From the results obtained above, INH appeared to inhibit the synthesis of mycolic acids longer than C₄₄ or C₄₆ specifically by inhibiting chain elongation or desaturation of precursor long chain fatty acids longer than C₂₈ or C₃₀.     

Defective mycolic acid biosynthesis in a mutant of Mycobacterium smegmatis.

A mutant of Mycobacterium smegmatis defective in mycolic acid biosynthesis was isolated following chemical mutagenesis. Fatty acids were extracted from the mutant and subjected to structural analysis by thin-layer chromatography and high-performance liquid chromatography (HPLC) of both methyl and p-bromophenacyl ester derivatives. Thin-layer chromatography did not show the presence of any fatty acid of RF comparable to that of standard methyl mycolate. The HPLC profile revealed a broad peak in the standard mycolic acid ester region. No characteristic peaks of mycolic acid esters comparable to the wild-type could be resolved. Mass spectral analysis of the HPLC-purified peak demonstrated the presence of shorter-chain fatty acids in the mutant. These data support the idea that the mutant accumulates precursors of mycolic acids and is incapable of carrying out the final conversion to mycolic acids of 60-90 carbon atoms.     

Influence of Tween 80 on the mycolic acid composition of three cutaneous corynebacteria

Changes in the mycolic acid composition of three cutaneous strains of corynebacteria were caused by the addition of Tween 80 to the culture medium. Gas chromatography-mass spectrometry showed that the carbon chain length and the degree of unsaturation had been affected: the levels of corynomycolic acid with 36 carbon atoms and two double bonds increased significantly.     

Molecular weight determination of methyl esters of mycolic acids using thermospray mass spectrometry.

Methyl esters of normal fatty acids, corynomycolate and corynomycolenate were used as model compounds for thermospray mass spectrometric procedures for molecular weight determination of the related nocardial mycolic acids. By using ammonium acetate at the positive ion generator, in both cases, a family of ions was produced. The following members were found and corresponded to the adducts: (1) M + H; M + NH4 and M + H + NH4 for methyl esters of normal fatty acids, whereas M + H, M + 2H and M + H + NH4 were the adducts most frequently observed with methyl corynomycolates. The methyl esters of C40-C48 mycolic acids from Rhodococcus rhodochrous exhibited prominent peaks corresponding to adducts M + H + NH4 whereas those corresponding to M + 2H showed slightly lower intensities. The structure M + H had no significant representatives with this subclass of mycolic acids. A similar pattern was observed with methyl esters of C50-C54 mycolic acids from Nocardia asteroides GUH-2. Ion peaks C50-C54 representing adducts M + 2H and M + H + NH4 prevailed in the mass spectrum. In this case, the intensities of peaks corresponding to M + 2H were slightly higher than those of the M + H + NH4. Essentially three main species of nocardomycolic acids were detected: (1) monounsaturated C50:1, C52:1 and C54:1; (2) diunsaturated C50:2, C52:2 and C54:2 and (3) triunsaturated C52:3 and C54:3 mycolic acids. The most abundant mycolic acid was C52:2 followed in decreasing abundance by C52:1, C54:2, C50:2, C52:3 and C54:3 mycolic acids.     

Effect of creatine on contents of myosin heavy chain and myosin-heavy-chain mRNA in steady-state chicken muscle-cell cultures.

The i.r. spectra of bilirubin isomers that differ in number and position of vinyl groups were examined to verify the assignment of the 988 cm-1 peak of bilirubin (991 cm-1 peak in calcium bilirubinate) to its pendant vinyl groups. There were only small changes in this peak with changes in position of vinyl groups (exo-2- and -18-vinyl versus endo-3- and -17-vinyl), but progressive loss of peaks in this region was observed when vinyl groups were reduced to ethyl groups (dihydrobilirubin and mesobilirubin). Methylvinylmaleimide, a monopyrrole derived from the outer (A and D) rings of bilirubin, has a pendant vinyl group and exhibits a prominent peak at 986 cm-1, but haematinic acid methyl ester derived from the inner (B and C) rings has no vinyl group and shows no peak near 988 cm-1. These observations verify the assignment of the 988 cm-1 peak of bilirubin to its pendant vinyl groups. This supports our previous proposal that a decrease in the peak at 985-995 cm-1 in the i.r. spectra of pigment gallstones, as compared with unconjugated bilirubin or calcium bilirubinate, indicates a consumption of vinyl groups in the process of formation of the polymer in the pigment stones.     

Purification and characterization of C28-55 fatty acids from Mycobacterium smegmatis

The nonmycolic C16 to C55 fatty acids obtained from Mycobacterium smegmatis ATCC 356 by saponification were enriched with respect to the C28 to C55 acids by successive chromatography on silicic acid and Sephadex LH-20 columns. These partially purified fatty acids were then derivatized to the p-bromophenacyl ester and further fractionated by argentation thin-layer chromatography and reverse-phase high-performance liquid chromatography into their individual components. The esters were characterized by electron impact mass spectrometry. Two structural series of C28:1 to C42:1 and C45:2 to C55:2 fatty acids were identified as possible precursors of the monoenyl and dienyl mycolic acids, respectively. These acids were structurally related to the alpha-alkylhydroxyl group of the corresponding mycolic acid. The results suggest that these C28 to C55 fatty acids (meromycolic acids) of M. smegmatis might be precursors of mycolic acids.