Effect of the carbon dioxide modifier on the lipid composition of wool wax extracted from raw wool

Wool wax or lanolin is a unique substance secreted by sheep and forms a natural protective coating on wool fibres. It is widely used in pharmaceutical and cosmetic formulations. However, different systems of wool wax recovery from scouring liquour provide a dark impurified greasy product. This product has a lipid composition that differs from the wool wax present on wool fibres. The wool wax extraction method from raw wool with pressurised CO₂ and different modifiers at constant pressure and temperature was studied. Thin-layer chromatography coupled to an automated flame ionisation detection system (TLC/FID) was used to analyse the different lipid classes present in the collected extracts. Moreover, a detailed structural comparison of the cholesteryl esters and hydroxycholesteryl esters was carried out by means of sub-ambient pressure chromatography mass spectrometry in the electron impact and in the ammonia positive chemical ionisation modes. For comparison, qualitative and quantitative analyses of the lanolin extracted in Soxhlet with dichloromethane and commercial cosmetic lanolin were carried out. Differences in the quantity of wool wax extraction and in the lipid composition of different wool wax extracts were detected by changing the modifier polarity.     

共反应剂法合成纤维素高级脂肪酸酯

采用乙酸酐为共反应剂合成了纤维素高级脂及酸酯。运用IR,^1H NMR对合成产物的结构进行了表征。结果表明,在纤维素分子骨架上同时接技了高级脂肪酸酯基以及乙酸酯基,产物为纤维素混合酸酯。产物的酯化度值通过^1H NMR确定,可达到1.66。研究了纤维素与系列高级脂肪酸的酯化反应,结果表明,随着高级脂肪酸中碳原子数目的增加,产物的酯化度逐渐减小,而产物质量有一个先增加后减小的趋势。     

Rapid analysis of non-esterified fatty acids as methyl esters from different biological specimens by gas chromatography after one-step esterification

A rapid gas chromatographic method for the determination of medium-chain and long-chain free fatty acids (C14:0 to C24:0 fatty acids) from different biological specimens is presented. After a rapid one-step transesterification method in methanol-acetyl chloride (50:1, v/v), fatty acid methyl esters were extracted into n-hexane and analysed on a 15-m Durabond-Wax column within a 12-min chromatographic run. The detection limit is 500 pg per injection.     

Characterization of a lipid-rich fraction synthesized by Streptomyces avermitilis

Isolation of the macrocyclic lactone parasiticide avermectin and other closely related natural products produced by Streptomyces avermitilis also yields a lipid-rich fraction. The latter has been characterized by techniques based on gas-liquid chromatography (GLC) and mass spectrometry (MS). Initial examination of the lipid-rich fraction by direct probe electron-impact (EI) MS and packed-column GLC showed that it consists primarily of a mixture of triglycerides possessing C14-C17 acyl groups. Further examination of this fraction by capillary column GLC-MS demonstrated that it contains low levels of C15-C17 free fatty acids, squalene and diglycerides and, as the major components, at least ten mixed acyl triglycerides (total number of acyl carbon atoms ranging from 43 to 50). Prominent among the triglycerides were a C15-C15-C16 species, a C15-C16-C16 species and a C15-C16-C17 species. Capillary-column GLC and GLC-MS of the fatty acid methyl esters resulting from transesterification demonstrated that the major triglyceride acyl groups are anteiso-C15 (12-methyltetradecanoyl), iso-C16 (14-methylpentadecanoyl), n-C16 (hexa-decanoyl) and anteiso-C17 (14-methylhexadecanoyl). Lower levels of the methyl esters of the following fatty acids were observed: iso-C14 (12-methyltridecanoic), n-C14 (tetradecanoic), iso-C15 (13-methyltetradecanoic), n-C15 (pentadecanoic), iso-C17 (15-methylhexadecanoic) and n-C17 (heptadecanoic). Little evidence was seen for either unsaturated acyl groups or acyl groups of less than 13 or more than 18 carbon atoms. Desorption chemical ionization MS (ammonia reagent gas) analysis confirmed the nature of the lipid-rich fraction, and is an attractive one-step approach for determining the molecular weights and distribution of triglycerides in a mixture.     

High-resolution analysis by nano-electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry for the identification of molecular species of phospholipids and their oxidized metabolites

Nano-electrospray ionization (ESI) Fourier transform ion cyclotron resonance mass spectrometry (FTICRMS) was applied to identify the molecular species of phosphatidylethanolamine of Caenorhabditis elegans, which has a high concentration of phospholipids with a fatty acyl chain having an odd number of carbon atoms. The molecular species of diacyl phosphatidylethanolamine with one fatty acyl chain having an odd number of carbon atoms and one fatty acyl chain having an even number of carbon atoms was identified separately from alkyl-acyl phosphatidylethanolamine with an alkyl chain having an even number of carbon atoms and a fatty acyl chain having an even number of carbon atoms. Furthermore, nano-ESI-FTICRMS was applied to the direct identification of oxidized phosphatidylcholine from soybean. The mass peaks of individual molecular species of oxidative phosphatidylcholine, such as 34:3 diacyl phosphatidylcholine with peroxide (+2O) (m/z 788.544) or 34:2 diacyl phosphatidylcholine with peroxide (+2O) (m/z 790.560), can be separated from the peaks of the molecular species of the non-oxidative phospholipids. This suggests that the mass peaks with a difference of less than 0.1 mass units in their molecular weight can be separated and that their individual exact molecular compositions can be obtained by the FTICRMS analysis. The high resolution and high accuracy of FTICRMS are very effective in the analysis of molecular species of phospholipids and their derivatives.     

Identification of intact long-chain p-hydroxycinnamate esters in leaf fibers of abaca (Musa textilis) using gas chromatography/mass spectrometry

The study of acetone-extractable components from the leaf fibers of the non-wood plant abaca (Musa textilis) resulted in the isolation and identification of series of intact hydroxycinnamate esters consisting of ferulic and p-coumaric acids esterified to long-chain fatty alcohols (C20 to C28) and omega-hydroxyfatty acids (C22 to C28). These series of compounds were characterized by high-temperature gas chromatography/mass spectrometry (GC/MS) using capillary columns (12 m length) with thin films that allowed the analysis of intact (i.e., without prior saponification) hydroxycinnamate esters. Characterization of intact individual compounds was achieved based on the mass spectra obtained by GC/MS of the underivatized compounds and their methyl and/or trimethylsilyl ether derivatives.     

Proton affinities of saturated aliphatic methyl esters

The kinetic method was used to determine the proton affinities of methyl esters of several saturated fatty acids. Decompositions of the proton-bound dimers of the methyl esters, AHB⁺, were observed under different conditions with two instruments. The proton affinities (PAs) of the methyl esters increase continually with increasing carbon number in the acid. Equilibrium and initial rate experiments were performed with a Fourier transform ion cyclotron resonance mass spectrometer on the methyl ester of the C₂₂ saturated acid (methyl behenate). These experiments give values for PA (methyl behenate) that are perhaps slightly lower than those obtained with the kinetic method. The PAs of the methyl esters of the fatty acids could be correlated with the equation: PA (ester) = (40.0 ± 2.5)*log(n) + (784.7 ± 3.9) kJ/mol or PA (ester) = (864 ± 2) − (479 ± 41)/n, wheren = number of atoms in the molecule. Proton affinities of smaller sets of 1-alkylamines and 1-alkanols can be fit to similar equations.     

A density functional study on beryllium-doped carbon dianion clusters CnBe2- (n = 4-14)

Making use of the software of molecular graphics, we designed numerous models of C(n)()Be(2-) (n = 4-14). We carried out geometry optimization and calculation on vibration frequency by means of the B3LYP density functional method. After comparison of structure stability, we found that the ground-state isomers of C(n)()Be(2-) (n = 4-14) are linear with the beryllium atom located inside the C(n)() chain. When a side carbon chain is with an even number of carbon atoms, it is polyacetylene-like, whereas when a side chain is with an odd number of carbon atoms, it is cumulene-like. The C(n)Be(2-) (n = 4-14) clusters with an even number of carbon atoms are more stable than that with an odd number of carbon atoms, matching the peak pattern observed in accelerator mass spectrometry (AMS) and Coulomb Explosion Imaging (CEI) investigations of C(n)()Be(2-) (n = 4-14). The trend of such odd/even alternation is explained based on concepts of bonding characteristics, electronic configuration, electron detachment, and incremental binding energy.     

Analysis of isomeric long-chain hydroxy fatty acids by tandem mass spectrometry: application to the diagnosis of long-chain 3-hydroxyacyl CoA dehydrogenase deficiency

Acetyl trimethylaminoethyl ester iodide derivatives have been used to selectively analyze isomeric long-chain hydroxy fatty acids by electrospray ionization tandem mass spectrometry (ESI-MS/MS). The binary derivatives of 2-, 3-, 12- and 16-hydroxypalmitic acids afford remarkably different product ion spectra. Further discrimination between isomers is possible by acylating with pivaloyl chloride. 2- and omega-Hydroxy long-chain fatty acids form pivaloyl esters in quantitative yield whereas other secondary alcohols only partially react. Cotton-based filter paper used for blood collection contains substantial amounts of esterified long-chain hydroxy fatty acids. From the product ion spectra of the acetyl trimethylaminoethyl esters the hydroxydocosanoic and -tetracosanoic acids are >90% omega-hydroxy. All remaining saturated and unsaturated hydroxy acids are >90% 2-hydroxy acids. A method for the quantification of free 3-hydroxypalmitic acid in plasma by ESI-MS/MS for the diagnosis of long-chain 3-hydroxyacyl CoA dehydrogenase deficiency (LCHAD) is described. Median plasma concentrations of 0.43 micromol/L (control, n = 22) and 12.2 micromol/L (LCHAD, n = 3) were obtained from 5 microL plasma samples.     

Characterization of internal lipids from wool

The internal lipids of wool were isolated after solubilizing the wool keratin with a mixture of papain and dithioerythritol 1.4. Thin layer chromatography of the internal lipids was applied to classify the extracted components. The internal wool lipids gave 11 spots of which some were identified as cholesterol and free fatty acids, C-16 and C-18 being predominant. A very small amount of triglycerides and cholesterol esters were also found. A characteristic difference between internal wool lipids and wool wax appears to be the limited number of well defined components of which free cholesterol and fatty acids constitute the main bulk. Furthermore another feature of keratin membrane lipids is the extremely reduced phospholipid: cholesterol ratio of 0.3. The internal lipids originate mainly from the cell membrane complex of the fiber. The existence of a chemically modified bilayer membrane structure without the essential phospholipids must be taken into consideration.     

Analysis of Polyunsaturated Fatty Acids Using High Performance Liquid Chromatography – Atmospheric Pressure Chemical Ionization Mass Spectrometry

Analysis of polyunsaturated fatty acids using high performance liquid chromatography–atmospheric pressure chemical ionization mass spectrometry is described. The standard fatty acid methyl esters from 16 to 22 carbons were analyzed by LC‐MS with APCI. The effect of orifice voltage and total carbon atoms versus number of double bonds in each homologue on the mass spectra is discussed. The correction coefficients for homologues from saturated fatty acids to hexaenoic acid are also mentioned.     

Identification by gas chromatography and mass spectrometry of lipids from the rat Harderian gland

Lipids from rat Harderian glands were extracted with ethyl acetate, hydrolysed with base and examined by gas chromatography (GC) and gas chromatography-mass spectrometry (GC-MS) as trimethylsilyl (TMS), [2H9]TMS, methyl ester-TMS, picolinyl, nicotinate and nicotinylidene derivatives. The latter three derivatives were used to reveal the structures of the alkyl chains of fatty acids, alcohols and glycerol ethers, respectively. Forty-eight compounds were identified, representing about 97% of the total extracted lipids as measured by GC peak areas. The major constituents were fatty acids with chain lengths from 12 to 22 carbon atoms (mainly C18 and C20) and fatty alcohols (C16 to C26) derived from wax esters. Most of these acids and alcohols were unsaturated in the omega-7 position and were accompanied by smaller amounts of the saturated and omega-5 monounsaturated analogues. glycerol ethers were also identified for the first time in this secretion; the ether chains contained from 14 to 19 carbon atoms (mainly 16) and were straight-chain saturated, unsaturated (omega-5 and omega-7) and branched (iso). The only sterol found was cholesterol amounting to 1.24% of the total extract.     

Equivalent chain lengths of all C4-C23 saturated monomethyl branched fatty acid methyl esters on methylsilicone OV-1 stationary phase

Isomer mixtures of monomethyl branched saturated C7-C23 fatty acid methyl esters (FAME) were prepared by performing a methylene insertion reaction to the straight chain FAME and this study model was completed by using commercially available standards of C4-C7 FAME. The equivalent chain lengths (ECL) of all 220 C4-C23 monomethyl branched FAME on OV-1 stationary phase were measured, achieving an average repeatability of ±0.0004 ECL units. The monomethyl branched FAME was identified by GC on the basis of regularity of the fractional chain lengths (FCL) dependence on the number of carbon atoms (C(z)) of individual homologous series of methyl 2-, 3-, …, 21-FAME. The prediction of retention of the first homologues, having the new position of methyl group beginning at higher carbon atoms number, and analogously for the second, third, fourth, and other members of the homologous series, allowed the dependence FCL=f(C(z)) for the first and subsequent members of beginning homologous of monomethyl derivatives of FAME. The identification was confirmed by mass spectrometry. All of the methyl isomers of FAME, which could not be completely separated by gas chromatography due to having a methyl group in surroundings of the middle of the carbon chain, were resolved by mass spectrometry using deconvolution in a SIM-mode. Measured gas chromatographic and mass spectrometric data were applied for identification of the monomethyl branched saturated FAME in tongue coating.     

液相色谱-四极杆-飞行时间串联质谱法高通量分析美国红参的脑苷脂分子种

利用液相色谱-四极杆-飞行时间串联质谱法(LC-Q-TOF MS/MS)分析美国红参(Parastichopus californicus)脑苷脂的分子种组成。以氯仿-甲醇(2：1,v/v)溶液提取,经皂化和固相萃取柱(SPE)净化后,在正离子模式下,通过自动二级质谱方式同时获取样品脑苷脂分子的一级和二级质谱图。筛取含180 Da中性丢失碎片的分子,再结合二级质谱中的长链碱和脂肪酸碎片,确定每种脑苷脂分子的结构。通过该法一次性共分析出123种美国红参脑苷脂分子种;含有的长链碱共18种,其中植物鞘氨醇型与鞘氨醇型长链碱的相对含量比为1：2;脂肪酸为18~25碳的饱和或单不饱和脂肪酸,以24碳脂肪酸为主,2-羟基脂肪酸占58.62%,饱和与单不饱和脂肪酸的相对含量比约为1：3。LC-Q-TOF MS/MS法分析海参脑苷脂分子种的灵敏度高,准确,简便,为海参脑苷脂的构效关系研究及功能食品开发提供理论依据。     

Temperature-programmed gas chromatography linear retention indices of all C4-C30 monomethylalkanes on methylsilicone OV-1 stationary phase. Contribution towards a better understanding of volatile organic compounds in exhaled breath

Monomethylated alkanes have been proposed to be contained in exhaled breath, their concentration pattern serving for identification of lung carcinoma, breast carcinoma and rejection of foreign tissue after heart transplant rejection. Improving the accuracy of identification for monomethylated alkanes will enhance work on their biochemical background which presently is unknown. The programmed temperature linear retention indices of all 196 C(4)-C(30) monomethylalkanes on OV-1 stationary phase were measured with an average repeatability of +/-0.07 index units (i.u.). The mixture of C(9)-C(30) monomethylalkanes was prepared by methylene insertion reaction to C(8)-C(29)n-alkanes mixture. The preliminary identification of monomethylalkanes was performed on the basis of the dependence of homomorphy factors on the number of carbon atoms of individual homologous series of monomethylalkanes (retention indices extrapolated with s=0.15 i.u.). The prediction of retention of isomers with new position of methyl group beginning at higher carbon atoms number, as well as for second, third, fourth, etc., member of homologous series allowed the dependence H(P)=f(C(n)) for first, second, third, etc., members of beginning homologous of monomethylalkane series (retention indices extrapolated with s=0.17 i.u.). The identification was confirmed by mass spectrometry. All gas chromatographic unseparated monomethylalkane isomers with methyl-group near the middle of molecule carbon chain were resoluted by mass spectrometric deconvolution. Obtained regular dependences H(P)=f(C(n)) allow precise retention prediction of monomethylalkanes C(30).     

Improved and simplified tissue extraction method for quantitating long-chain acyl-coenzyme A thioesters with picomolar detection using high-performance liquid chromatography.

A method has been developed that permits rapid and easy tissue extraction of long-chain acyl-coenzyme A (acyl-CoA) thioesters with sensitive quantitation by reversed-phase high-performance liquid chromatography (RP-HPLC). Tissue homogenants are extracted using a reserve Bligh-Dyer technique, and long-chain acyl-CoA esters are harvested in the methanolic aqueous phase. Complex lipids and phospholipids are removed in the chloroform-rich organic Bligh-Dyer second phase, and long-chain acyl-CoA compounds are further purified from the methanolic aqueous Bligh-Dyer first phase on C18 extraction columns after removal of the methanol. The eluted and purified acyl-CoA esters are then quantitated by RP-HPLC using heptadecanoyl-CoA as an internal standard resulting in a detector sensitivity of about 12 pmol. Ten long-chain acyl-CoA esters from C12:0 to C20:4 were identified and separated from canine renal cortex and murine liver samples. The predominant acyl-CoA peaks from both kidney and liver were 14:0, 16:1, 16:0, 18:1, 18:2 and 20:4. Murine liver also produced 18:0 and all peaks disappeared after alkaline hydrolysis of the samples. This extraction and quantitation technique can successfully be used for tissue samples as small as 20 mg, and many samples can be processed in a short period of time. The simplicity of the extraction procedure and the sensitivity of the assay make this an attractive alternative approach to quantitating long-chain acyl-CoA thioesters from complex biological samples such as tissues.     

Molecular characterization of a highly heterogeneous mixture of glucosylceramides from a deep-water Mediterranean scleractinian coral Dendrophyllia cornigera

We report here on the structural characterization of a highly heterogeneous mixture of glucosylceramides (GlcCers) isolated from a deep-water Mediterranian dendrophylliid coral, Dendrophyllia cornigera. The neutral glycosphingolipid (GSL) components of the coral were separated into three HPLC fractions which were structurally characterized by nuclear magnetic resonance (NMR) and mass spectrometry (MS). NMR analysis revealed a beta-glucosylpyranose, a methyl branched conjugated sphingadienine and alpha-hydroxy fatty acid moieties characteristic for the species. Molecular mass distributions of the HPLC fractions were monitored using single-stage MS. At least 17 different GlcCer constituents with variable long-chain base and fatty acid residues were observed based on the molecular ion peaks in the liquid secondary ion (LSI) survey spectra. Structures of the individual components were revealed by product ion spectra of the alkali-cationized molecules ([M + Cat](+)), which resulted in two characteristic fragment ions, F(F) and F(S). Tandem MS of the same fragment ions formed in the ion source showed that F(F) carries the hydoxy fatty acid, while F(S) carries the long-chain sphingoid base, thus providing complementary structural information for the characterization of ceramide composition. Based on the tandem mass spectra of the molecular ions [M + Na](+), 26 different GlcCers of the coral were identified. The ceramide moiety showed heterogeneity in both the sphingoid portion (d18:2, d19:2, d20:2 and d20:3) and the alpha-hydroxy fatty acid chain (h19-h24, either saturated or unsaturated), forming an extremely heterogeneous mixture. The method is generally applicable to the characterization of structurally heterogeneous GlcCer mixtures.     

Fatty acids and amino acids of entomopathogenic fungus <Emphasis Type="Italic">Conidiobolus coronatus</Emphasis> grow on minimal and rich media

Entomopathogenic fungi are referred to as potential candidates as insect pest control agents. The objective of the study was to identify fatty acids and amino acids from Conidiobolus coronatus cultured on two different media. Each medium was extracted with ethyl acetate and its mixtures with isopropanol, acetonitrile and methanol. Analyses of fatty acids and amino acids of entomopathogenic fungus C. coronatus were performed by means of gas chromatography coupled with mass spectrometry. The analysis showed that the fungus C. coronatus produces the following groups of compounds: fatty acids and amino acids; α- and β-glucopyranose were also identified. The identified fatty acids included 12–20, 22 and 24 carbon atoms per chain. The highest content of fatty acids was detected in a mycelium sample cultured in a liquid minimal medium extracted with ethyl acetate. The lowest content of these organic compounds was identified in mycelium cultured in a liquid nutrient-rich medium extracted with ethyl acetate–methanol mixture. Fatty acids were found to account for 62.0 mass % to 94.4 mass % of all organic compounds in the analyzed mycelia. C18:1 acids were detected in the highest amounts when ethyl acetate was used as the extracting agent. The identified amino acids accounted for 4 mass % to 21 mass % of all organic compounds. Upon extraction of C. coronatus mycelium samples with the ethyl acetate—methanol mixture, two anomeric forms of glucose were also identified. An analysis of the studied material confirmed, that the entomopathogenic fungus C. coronatus is a very rich source of organic compounds, which might encourage its further research so as to identify an even larger number of compounds being produced by this species.     

Preparation of 6-O-(4-alkoxytrityl)celluloses and their properties

Cellulose was reacted with a series of 4-alkoxytrityl chlorides (C(n)TCl, n: number of carbon atoms in a saturated alkyl chain) under homogeneous reaction conditions in LiCl-N,N-dimethyl acetoamide to give a series of 6-O-(4-alkoxytrityl)celluloses (C(n)TC) with a high degree of substitution (DS), from 0.94 to 0.99, and with high regioselectivity at the 6-O position. Solubility of the C(n)TC in nonpolar solvents depended on the alkyl chain length: as the alkyl chain lengthens, cellulose derivatives become more hydrophobic and are readily soluble in nonpolar solvents, but not in polar solvents. Acetates of the C(4)-C(18)TC (C(4)-C(18)TCAc) showed anisotropic structures over melting temperatures (T(m)) examined under a polarized optical microscope (POM). Over isotropization temperatures (T(i)), flow birefringence were detected for C(12)-C(18)TCAc. The T(m) and T(i) decreased linearly with an increasing number of carbon atoms in the alkyl substituent. Wide-angle X-ray scattering (WAXS) studies of C(n)TC indicated that the fully extended side chains were perpendicular to the polymer backbone and interdigitated. These C(n)TC with the improved solubility may be used as starting materials for further derivatization focused on the secondary hydroxyl groups at the C-2 and C-3 positions.     

Identification by combined gas chromatography-mass spectrometry of constituent long-chain fatty acids and alcohols from the meibomian glands of the rat and a comparison with human meibomian lipids

Constituent long-chain fatty acids and alcohols from the meibomian secretions of the rat were examined as trimethylsilyl (TMS) and methyl ester-TMS derivatives by capillary gas chromatography and by combined gas chromatography-mass spectrometry. The positions of double bonds and methyl branch points were determined by the mass spectra of picolinyl esters and nicotinates for long-chain fatty acids and alcohols, respectively. Fatty acids had chain lengths from C12 to C34 and were of the straight-chain iso, anteiso and monounsaturated types. The unsaturated acids had double bonds in the omega-7 and omega-9 positions. The alcohols had corresponding structures. In common with the constituent acids and alcohols of other meibomian secretions, the chain lengths of the constituents showed a biphasic distribution with maxima around C16-C18 and C25-C27. The profile was qualitatively similar to that obtained from human meibomian secretion but with some differences in the relative proportions of certain acids and alcohols.