Analysis of α-branched fatty acids by gas chromatography and mass spectrometry

Gas liquid chromatography and mass spectrometry were utilized in combination to identify isomeric α-branched chain fatty acid methyl esters. In a given isomeric series equivalent chain length, values decreased with increase in the number and size of α-alkyl substituents. Mass spectra of the α-monoalkyl derivatives are characterized by prominent McLafferty rearrangement ion peaks, whereas those of the α,α-dialkyl isomers contain the former ions plus an α-cleavage ion. Presented at the AOCS Meeting, Chicago, September 1973.     

Confirmation by gas chromatography/mass spectrometry of two unusualrans-3-monoethylenic fatty acids from the nova scotian seaweedsPalmaria palmata andChondrus crispus

The structures of two unusual fatty acids, the knownrans-3-hexadecenoic acid and a noveltrans-3-tetradecenoic acid, both isolated from the Nova Scotian sea-weedsPalmaria palmata andChondrus crispus, were positively identified. After the extraction of the total fatty acids by saponification, followed by methylation, the monoenoictrans fractions were isolated by thin-layer chromatography on silica gel impregnated with silver nitrate. The monoenoictrans fractions were derivatized with 2-amino-2-methyl-propanol prior to analysis by gas chromatography/mass spectrometry. The mass spectra showed prominent [M+1]⁺ ions but lacked the 12 amu interval useful for identifying the double bond position. Hence, alternative diagnostic peaks were used to confirm the position of the double bond in these two fatty acids. As thetrans-3-hexadecenoic acid is found in the photosynthetic tissue of all plants, it may also be present in ruminant fats and, presumably, in human adipose tissue.     

Gas chromatography/tandem mass spectrometry as an analytical tool for the identification of fatty acids

Structures of fatty acids present at very low quantities in mycobacteria are difficult to determine. A commonly used strategy is to introduce heteroatoms into functional groups by chemical means before subjecting them to gas chromatography/tandem mass spectrometry (GC/MS/MS) analysis. Routinely used methods give very low abundance diagnostic ions leading to ambiguities in structural conclusions. GC/MS/MS associated with electron capture ionization of pentafluorobenzyl esters was used to study very complex mixtures of fatty acids fromMycobacterium fallax andM. aurum. The charge-remote fragmentation of fatty acid carboxylate anions was used for structure determination at the nanogram level of a large number of unsaturated, branched, and cyclopropane-containing fatty acids. Some of them have not been observed previously in these Mycobacteria. On the basis of these studies, biosynthetic pathways of unsaturated, branched, and cyclopropane-containing fatty acid are proposed.     

Jojoba oil analysis by high pressure liquid chromatography and gas chromatography/mass spectrometry

Two analytical procedures for determining com-positions of jojoba liquid wax esters are described and compared. One, the more tedious, involves separation of wax ester homologs by high pressure liquid chro-matography followed by determination of the acid and alcohol moieties from each homolog. The second allows rapid determination of wax ester composition by gas Chromatographic separation of hydrogenated jojoba wax esters according to chain length, followed immediately by ancillary mass spectrometric identifi-cation of the acid and alcohol moieties. Double bonds in the alkyl chains in jojoba liquid waxes were almost exclusively (98%) ω-9, when examined by gas chro-matography/mass spectrometry (GC/MS) and ozonolysis/GC/MS. Presented in part at the 2nd International Conference on Jojoba and Its Uses, Ensenada, Mexico, February, 1976.     

Quantitative analysis of food fatty acids by capillary gas chromatography

The superior efficiency of capillary columns is desirable for the gas chromatographic analysis of complex mixtures of fatty acids, but there have been some reservations regarding quantitation and reproducibility. This paper discusses the use of wall-coated glass capillary columns in a semiautomated system for the determination of food fatty acids. Glass columns coated with SP2340 were used for extended periods at temperatures up to 200 C without appreciable deterioration. Up to 1900 samples were analyzed on a single column over an 11-month period, with only minor changes in retention ratios, response factors and column efficiency. Quantitative precision of results, calculated either as normalized weight percentage or as absolute amounts, based on the use of an internal standard, were typically within 2% relative deviation. Difficulties encountered in obtaining acceptable chromatograms and reproducible data are discussed, and typical analyses of the fatty acids from foods presented.     

Sperm whale oil analysis by gas chromatography and mass spectrometry

Gas liquid chromatography of winterized sperm oil showed that its wax esters with even carbon numbers range from C₂₄ to C₄₂ and are present in quantities resembling a normal distribution curve with C₃₄ as the mean. Between these even-numbered wax esters, ones with odd chain lengths were eluted. Triglycerides, similarly present in a normal distribution pattern, ranged from C₄₂ to C₅₈ and also included traces of odd chain species. The component acids and alcohols were analyzed by gas chromatography-mass spectrometry, and double bond positions in the monoenoid components were established. Branched chain and odd chain constituents, both saturated and unsaturated, were detected among both alcohols and acids. These moieties, when combined with those having even chains, are responsible for the wax esters and triglycerides with odd carbon numbers. ARS, USDA.     

Analysis of fatty acids by mass spectrometry in the selected ion monitoring mode

GC/EI‐MS‐SIM (gas chromatography / electron ionisation – mass spectrometry with selected ion monitoring) can be easily applied to the determination of fatty acids as methyl esters (FAME). SIM masses should have a high relative abundance and for this reason we suggest m/z 74 and m/z 87 for saturated and monoenoic fatty acids as well as m/z 79 and m/z 81 for FAME with more than one double bond. The relevance and ratio of these fragment ions along with GC retention times enables a good identification of a FAME represented by a peak detected in the chromatograms. Compared to GC with a flame ionisation detector, this simple method provides a higher selectivity and better sensitivity. It also offers the possibilities of using a wide range of internal standards which enables the individual quantification of FAME instead of the determination of their contributions to the fatty acid pattern.     

Detection of hydroxy fatty acids in biological samples using capillary gas chromatography in combination with positive and negative chemical ionization mass spectrometry

The most common method for use in the structural analysis of hydroxy fatty acids in biological samples is the gas chromatography-mass spectrometry (GC-MS) analysis of trimethylsilyl ethers of the methyl esters using electron impact ionization. A comparison of electron impact (EI) with chemical ionization mass spectrometry (CI-MS) shows that CI-MS is the superior technique. All ions necessary for structural analysis are observed at sufficiently high levels of intensity when methane or isobutane are used as reactant gases. The molecular weight can be determined from the ion group M+H, M-15 and M+H−90. The ionic series M+H−n×90 enables one to determine the number of hydroxyl groups. The position of the hydroxyl groups can be derived from the fragments of the α-cleavage of the fatty acid chain. The application of heptafluorobutyrates as derivatives for hydroxy fatty acid methyl esters shows advantages in the trace analysis of these compounds. Heptafluorobutyrates exhibit useful mass fragmentation patterns in the positive as well as in the negative CI mode. With methane as the reactant gas, M+H usually is base peak in positive mass spectra. The ionic series M+H−n×214 leads to the number of hydroxy groups in the molecule. In the negative mass spectra, M and M-20 are indicative for the molecular weight. The ion group m/z 213, 194 and 178 at high levels of intensity is typical for heptafluorobutyrates. The advantage of the application of heptafluorobutyrates is the high sensitivity which can be obtained in trace analysis using negative MS. Heptafluorobutyrates of hydroxy fatty acids gave a 20-fold higher response in the negative scan mode compared to that of the positive. The detection limit for heptafluorobutyrates in negative CI-MS was on the order of 1 fg (10⁻¹⁵ g).     

Pyrolysis/gas chromatography/mass spectrometry of glycidyl methacrylate—alkyl acrylate copolymers

Thermal degradation pattern of copolymers of glycidyl methacrylate with alkyl acrylates have been studied by pyrolysis/gas chromatography/mass spectrometry method. Various degradation products have been identified and based on the products obtained, the mechanisms of polymer degradation have been elucidated.     

Isolation and identification of trans -3,5-dimethoxystilbene from high quality tall oil fatty acids by liquid chromatography and mass spectrometry

trans-3,5-Dimethoxystilbene has been isolated from high quality tall oil fatty acids (unsaponifiables 1.5% maximum, rosin acids 1.5% maximum), using liquid column chromatography and low temperature solvent fractional crystallization. The structure of this compound was determined with the aid of IR, mass and NMR spectrometry. Thetrans-3,5-dimethyoxystilbene was found to be responsible for the development of a red color during the epoxidation of the tall oil fatty acids. Paper of the Journal Series, New Jersey Agricultural Experiment Station, Rutgers, The State University.     

Isolation and identification oftrans-3,5-dimethoxystilbene from high quality tall oil fatty acids by liquid chromatography and mass spectrometry

trans-3,5-Dimethoxystilbene has been isolated from high quality tall oil fatty acids (unsaponifiables 1.5% maximum, rosin acids 1.5% maximum), using liquid column chromatography and low temperature solvent fractional crystallization. The structure of this compound was determined with the aid of IR, mass and NMR spectrometry. Thetrans-3,5-dimethyoxystilbene was found to be responsible for the development of a red color during the epoxidation of the tall oil fatty acids.     

Analysis of the monoethylenic fatty acids of rapeseed oil by open tubular gas chromatography

A sample of rapeseed oil of known composition was analyzed by gas chromatography using open tubular columns and two types of polyester coating. The results confirm the previous analysis in most respects and elearly illustrate the possibility of using this technique to detect and determine isomeric forms of monoethylenic fatty acids. Some hitherto unreported trace components have been provisionally identified in rapeseed oil by linear log plots.     

The mass spectrometry of iso and anteiso monoenoic fatty acids

The normal, iso, and anteiso Δ⁸- and Δ⁹-17:1 fatty acid methyl esters were synthesized and their electron impact-induced fragmentation was studied by mass spectrometry. The mass spectra of the preterminal branched monoenoic fatty acid methyl esters present characteristic fragment ions, now understood to be indicative of the position of the methyl group. These fragment ions are in the iso compound m/e 227 [M-55]⁺, m/e 195 [M-87]⁺, and m/e 177 [M-105]⁺, while in the anteiso compound these fragments are shifted by 14 mass units to m/e 213, m/e 181, and m/e 163. The 15-D-iso Δ⁸- and Δ⁹-17:1 methyl esters were synthesized because the characteristic fragment ions in the methyl branched compounds indicated a key role of the tertiary hydrogen atom in the rearrangement process. A fragmentation mechanism consisting of a double bond migration triggered by the tertiary hydrogen and an allylic cleavage assuming a displacement mechanism is proposed.     

Analysis of conjugated bile acids by high performance liquid chromatography and mass spectrometry

Because of the known advantages of coupling high performance liquid chromatography with mass spectrometry (HPLC-MS) in biological fluids, studies on the reversed-phase HPLC-MS system for direct analysis of conjugated bile acids in human bile samples are described. Ten samples of gallbladder bile of apparently healthy subjects were examined. The amounts of each tauro- and glycoconjugated bile acid as trifluoracetate were determined by mass fragmentography. Quantitation of at least 1 ng of each bile acid was possible.     

The determination of specific free fatty acids in peanut oil by gas chromatography

Peanut oil is dissolved in 2,2,4-trimethylpentane standard. The free acids are extracted into N,N-dimethylformamide as their sodium salts and treated with methyl iodide. The resulting methyl esters are analyzed by gas chromatography. A typical result for oleic acid is about 200 ppm with a relative standard deviation of 6%. An average recovery of 95% is obtained from a sample spiked with an additional 200 ppm of oleic acid. Evidence is presented that the procedure does not give high values as a result of sample hydrolysis.     

Rapid determination of free fatty acids in vegetable oils by gas liquid chromatography

Determination of the free fatty acids in small quantities of vegetable oil is accomplished by gas liquid chromatography. The free fatty acids are isolated from a hexane solution of the vegetable oil into an aqueous solution of trimethylphenylammonium hydroxide (TMPH). Due to the alkalinity of TMPH, the free fatty acids readily partition into this aqueous phase. Injection of the free fatty acid-TMPH salts into a gas chromatograph results in pyrolytic methylation of the free fatty acid salts—yielding the methyl esters. Excellent results were obtained when this new procedure was used on neutral lipid oils containing known amounts of free fatty acids and compared with the results obtained by a modified BF₃/MeOH esterification procedure. When compared to the AOCS titration procedure, this new procedure gave comparable results. This new procedure has advantages over the AOCS procedure: it is more sensitive and gives quantitative results for individual free fatty acids. This new procedure also has several advantages over the modified BF₃/MeOH esterification procedure: it is easily and more rapidly performed, there is no deposition of glyceride on the column when the sample is injected, and because there is quantitative recovery, the new procedure is more sensitive and can be used on oils with a low weight percentage of free fatty acids.