P-bromophenacyl Esters Research Articles

Enhanced levels of atmospheric low-molecular weight monocarboxylic acids in gas and particulates over Mt. Tai, North China, during field burning of agricultural wastes

To understand the source and atmospheric behaviour of low molecular weight monocarboxylic acids (monoacids), gaseous (G) and particulate (P) organic acids were collected at the summit of Mt. Tai in the North China Plain (NCP) during field burning of agricultural waste (wheat straw). Particulate organic acids were collected with neutral quartz filter whereas gaseous organic acids were collected with KOH-impregnated quartz filter. Normal (C1-C10), branched (iC4-iC6), hydroxy (lactic and glycolic), and aromatic (benzoic) monoacids were determined with a capillary gas chromatography employing p-bromophenacyl esters. We found acetic acid as the most abundant gas-phase species whereas formic acid is the dominant particle-phase species. Concentrations of formic (G/P 1 570/1 410 ng m−3) and acetic (3 960/1 120 ng m−3) acids significantly increased during the enhanced field burning of agricultural wastes. Concentrations of formic and acetic acids in daytime were found to increase in both G and P phases with those of K+, a field-burning tracer (r = 0.32–0.64). Primary emission and secondary formation of acetic acid is linked with field burning of agricultural wastes. In addition, we found that particle-phase fractions (Fp = P/(G + P)) of formic (0.50) and acetic (0.31) acids are significantly high, indicating that semi-volatile organic acids largely exist as particles. Field burning of agricultural wastes may play an important role in the formation of particulate monoacids in the NCP. High levels (917 ng m−3) of particle-phase lactic acid, which is characteristic of microorganisms, suggest that microbial activity associated with terrestrial ecosystem significantly contributes to the formation of organic aerosols.

Aliphatic Glucoside, Zanthoionic Acid and Megastigmane Glucosides: Zanthoionosides A-E from the Leaves of Zanthoxylum ailanthoides.

From the leaves of Zanthoxylum ailanthoides, 4'-O-p-E-coumaric acid esters of 2-propanol β-D-glucopyranoside, megastigmane and megastigmane glucosides were isolated. Their structures were elucidated by spectroscopic evidence. The absolute configurations of the megastigmane and aglycone of megastigmane glucosides were determined by the octant rule and modified Mosher's method after protection of carboxylic acids by p-bromophenacyl esters and primary alcohols by pivaloyl esters.

1,3‐Alternate calix[4]arene‐bonded silica stationary phases. Effect of calixarene skeleton substituents on the retention mechanism and column selectivity

Four novel 1,3-alternate calix[4]arene-bonded silica gel stationary phases possessing different aromatic and aliphatic substituents at the upper rim (CalixNph, CalixBph, CalixHex, and CalixDdc) were prepared and structurally characterized. The comparison and selectivity of these phases were done by using alkylbenzenes, fatty acid p-bromophenacyl esters, aromatic positional isomers, and polynuclear aromatic hydrocarbons as analytes. Quantum chemistry calculations have also been performed (using an ab initio method) to support the experimental findings. The effect of the type and content of organic modifier on the retention and selectivity of the alkylbenzenes was studied. The retention mechanism is also discussed. The results indicate that the stationary phases behave like RP packings. However, inclusion complex formation and hydrophobic and pi-pi interactions seem to be involved in the separation process.

Gas chromatography–mass spectrometry of cis-9,10-epoxyoctadecanoic acid ( cis-EODA) : I. Direct evidence for cis-EODA formation from oleic acid oxidation by liver microsomes and isolated hepatocytes

Oleic acid, cis-9-octadecenoic acid, is the major fatty acid in mammals. Its oxide, cis-9,10-epoxyoctadecanoic acid ( cis-EODA), has been identified in blood and urine of humans, its origin is, however, still unknown. Lipid peroxidation and enzyme-catalyzed epoxidation of oleic acid are two possible sources. In the present article, we investigated by HPLC and GC–MS whether cis-EODA is formed enzymatically from oleic acid by the cytochrome P450 (CYP) system. Oleic acid, cis-EODA and its hydratation product threo-9,10-dihydroxyoctadecanoic acid ( threo-DiHODA) were quantitated by HPLC as their p-bromophenacyl esters. For structure elucidation by GC–MS, the pentafluorobenzyl (PFB) esters of these compounds were isolated by HPLC and converted to their trimethylsilyl ether derivatives. Liver microsomes of rats, rabbits and humans oxidized oleic acid into cis-EODA. This is the first direct evidence for the enzymatic formation of cis-EODA from oleic acid. The epoxidation of oleic acid was found to depend on CYP, NADPH+H +, and O 2. cis-EODA was measurable in incubates of liver microsomes for up to 30 min of incubation. Maximum cis-EODA concentrations were reached after 5–7 min of incubation and found to depend upon oleic acid concentration. Isolated rat hepatocytes hydratated cis-EODA into threo-DiHODA which was further converted to unknown metabolites. However, from incubation of oleic acid with these cells we could not detect threo-DiHODA or cis-EODA. Our study suggests that circulating and excretory cis-EODA may originate, at least in part, from CYP-catalyzed epoxidation of oleic acid. GC–MS of intact cis-EODA as its PFB ester in the negative-ion chemical ionization mode should be useful in investigating the physiological role of cis-EODA in man.

High-performance liquid chromatographic determination of the rhamnolipids produced by Pseudomonas aeruginosa

The bacterial biosurfactants 3-[3′-( l-rhamnopyranosyloxy)decanoyloxy]decanoic acid (RL-1) and 3-[3′-(2″-O-α- l-rhamnopyranosyl-α- l-rhamnopyranosyloxy)decanoyloxy]decanoic acid (RL-2) were isolated from Pseudomonas aeruginosa DSM 2659 cultures. An HPLC method was developed for the p-bromophenacyl esters of the rhamnolipids. Separation was obtained within 25 min on a RP C 18 column using a linear gradient of water-acetonitrile (30:70 to 0:100) and UV detection (265 and 320 nm). Linearity of response existed for 1.2–25 μg of the RL-1- and 1.8–25 μg of the RL-2- p-bromophenacyl ester (0.9–19.2 μg RL-1 and 1.3–18.0 μg RL-2). The reproducibility of the entire analytical method (extraction, derivatisation) was tested.

Synthesis of gibberellins A8 and A56 from gibberellin A3

A mixture of gibberellin A3 derivatives with 1(10)-ene-2β,3β-diol and 1(10)-ene-2α,3β-diol (2:5) groups, readily obtained from gibberellin A3, has been used for a new and simple synthesis of gibberellin A8 and its esters. The hydrolysis of GA3 and the iodolactonization of a mixture of the 2-epimers was carried out in aqueous solution in a single flask, as also was a synthesis of GA56 from GA3 by a method that we have modified. The mixture of 1β-iodides of GA8 and GA56 was separated by chromatography on SiO2 in the form of methyl or p-bromophenacyl esters which were then deiodinated and the methyl or p-bromphenacyl ester of GA8 was isolated. Free GA8 was obtained by the dephenylation of the latter ester. By two-dimensional NMR spectroscopy we succeeded in assigning all the signals in the13C and1H NMR spectra of the methyl esters of GA8 and GA56. In an attempt to obtain GA5 methyl ester by the action of trimethylchlorosilane/sodium iodide on the 2α,3β-diol system in GA56 methyl ester, the 8,13-epimer of the latter was formed, the structure of its molecule being established from the results of X-ray structural analysis.

Characterization of Mycobacteria Species by HPLC and Pattern Recognition

Multivariate methods of analysis are used to characterize 36 mycobacteria species following high-performance liquid chromatography (HPLC) of extracts of mycolic acids. After growth of the bacterial cultures, cell wall mycolic acids are saponified, extracted, then derivatized to the p-bromophenacyl esters. Reversed-phase HPLC with gradient elution separates 38 peaks in 10 min, plus an internal standard. A training set of 276 samples is composed and classification models are developed to predict the mycobacteria species. Classifications during the training phase are 91 and 95% accurate for single-pass and multilevel approaches, respectively. Validation is performed with a 549-sample set; successful classifications of known species are 87 and 92%, respectively. Bunching the predictions of MAIS complex species into one category improves the latter result to 97% correctly classified species. Fourteen of the 27 species (including M. tuberculosis) are classified without error in the validation data set.

Identification of (24E)-3 alpha,7 alpha-dihydroxy-5 beta-cholest-24-enoic acid and (24R,25S)-3 alpha,7 alpha,24-trihydroxy-5 beta-cholestanoic acid as intermediates in the conversion of 3 alpha,7 alpha-dihydroxy-5 beta-cholestanoic acid to chenodeoxycholic acid in rat liver homogenates.

Studies of chemical structure of the intermediates in the biosynthetic sequence between 3 alpha,7 alpha-dihydroxy-5 beta-cholestanoic acid (DHCA) and chenodeoxycholic acid have been undertaken. Radiolabeled DHCA was incubated with a rat liver preparation. The reaction products were converted to the p-bromophenacyl esters, and analyzed by reversed-phase high performance liquid chromatography. Under the conditions used, the radioactivity was found in (24E)-3 alpha,7 alpha-dihydroxy-5 beta-cholest-24-enoic acid (31%) and (24R,25S)-3 alpha,7 alpha,24-trihydroxy-5 beta-cholestanoic acid (7%) along with the starting material (62%). Neither the 24Z isomer of the alpha,beta-unsaturated bile acid nor the other three isomers of the beta-hydroxy bile acid were detected. The findings support the proposed pathway for the side chain cleavage in chenodeoxycholic acid biosynthesis, which is thought to be identical to that of cholic acid biosynthesis.

Identification of mycobacteria by high-performance liquid chromatography

Mycolic acids extracted from saponified mycobacterial cells were examined as p-bromophenacyl esters by high-performance liquid chromatography (HPLC). Standard HPLC patterns were developed for species of Mycobacterium by examination of strains from culture collections and other well-characterized isolates. Relative retention times of peaks and peak height comparisons were used to develop a differentiation scheme that was 98% accurate for the species examined. A rapid, cost-effective HPLC method which offers an alternative approach to the identification of mycobacteria is described.

High pressure liquid chromatography methods for separation of ω- and (ω-1)-hydroxy fatty acids: Their applications to microsomal fatty acid ω-oxidation

Fatty acids (C 12C 18) and their ω- and (ω-1)-hydroxy derivatives, when converted to p-bromophenacyl (PBP) esters, can be completely separated from one another by high pressure liquid chromatography (HPLC) on a silicic acid column using 0.5% ( v v ) isopropanol in n-hexane. In this system, fatty acid PBP esters are eluted at the solvent front, whereas the retention times of the ω- and (ω-1)-hydroxy derivatives are 14–20 and 24–29 min, respectively. The PBP esters can also be separated by reverse phase HPLC on a μBondapak C 18 column, a method which has been developed by Fan et al. (Fan, L. L., Masters, B. S. S., and Prough, R. A. (1976) Anal. Biochem. 71, 265–272) for separation of methyl esters of fatty acids and their ω- and (ω-1)-hydroxy derivatives. In the latter method, however, the retention times of ω- and (ω-1)-hydroxy derivatives are only about 2 min apart and an increase in the solvent polarity is needed for elution of the esters of unmodified fatty acids. Fatty acid PBP esters, however, can be obtained as independent peaks which are not disturbed by the solvent front. An application of the former method to measure fatty acid ω oxidation by liver microsomes and by a reconstituted monooxygenase system containing purified cytochrome P-450 is described.

Rapid method for the analysis of red blood cell fatty acids by reversed-phase high-performance liquid chromatography

A high-performance liquid chromatographic method was developed for a rapid qualitative and quantitative analysis of p-bromophenacyl esters of red blood cell fatty acids in humans. Both free and bound fatty acids, extracted with hexane-2-propanol (3:2) from packed red blood cells were derivatized with p-bromophenacyl bromide and analysed. Ten identical samples taken from a mixed pool of packed red blood cells from healthy subjects were analysed on two different columns. The fatty acid p-bromophenacyl esters were analysed on a 10 RP-18 column with methanol-acetonitrile—0.01 M ammonium formate as mobile phase and also on a 10 RP-8 column with acetonitrile—0.01 M ammonium formate as mobile phase. The two methods gave analogous results except in total analysis time: that on a 10 RP-8 column is ca. 40% shorter. Furthermore, a quantitative analysis of a standard solution to evaluate the extraction procedure in the absence or in the presence of the red blood cell core indicated a significant difference when the core is present.

High-performance liquid chromatography analysis of mycolic acids as an aid in laboratory identification of Rhodococcus and Nocardia species.

High-performance liquid chromatography analysis of the p-bromophenacyl esters of mycolic acids from whole organisms gave chromatographic patterns that were useful in differentiation of Rhodococcus and Nocardia species. Rhodococcus equi, R. erythropolis, and R. rhodochrous contained more-polar mycolic acids and were easily separated from the less-polar mycolic acid-containing species of R. sputi, R. bronchialis, R. corallinus, R. rubropertinctus, and R. terrae. The less-polar mycolic acid-containing Rhodococcus species showed chromatographic patterns that partially overlapped (in elution times) the patterns of Nocardia asteroides, N. otitidiscaviarum, and N. brasiliensis, but the larger number of peaks in the last species made separation between the genera possible. Distinct chromatographic patterns were found for most species, except for R. equi strains that showed two different patterns. Strains of R. rubropertinctus and R. terrae appeared identical. N. asteroides and N. otitidiscaviarum showed similar mycolic acid patterns.

Determination of .OMEGA.-betaines in aquatic animals by high performance liquid chromatography.

A method is described for the HPLC analysis of glycinebetaine, β-alaninebetaine. and γ-butyrobetaine in aquatic animals. The betaines in marine animals were extracted with 80% methanol and converted to p-bromophenacyl esters by treatment with p-bromophenacyl bromide in the presence of 18-crown-6 and KH2PO4. The esters of betaines having UV absorption maximum at 262nm were separated on Shimpack FLC-CN using 10mM KH2PO4 containing 2% acetonitrile, pH 2.10 as a mobile phase, The recoveries of betaines added to the muscle of marine animals were in the range from 93 to 104%.

Determination of organic acids (C1-C10) in the atmosphere, motor exhausts, and engine oils

A method is described for the determination of volatile organic acids in the atmosphere, motor exhausts, and engine oils. Atmospheric organic acids were collected on a KOH impregnated quartz filter and derivatized to p-bromophenacyl esters. The derivatives were analyzed by high-resolution capillary gas chromatography and gas chromatography-mass spectrometry. C/sub 1/-C/sub 10/ aliphatic organic acids and benzoic acid were detected in Los Angeles air. Acetic and formic acids are dominant followed by propionic acids. Total concentrations measured were 0.37-7.45 ppb. Organic acids (C/sub 1/-C/sub 10/) were also detected in the motor exhaust from a single automobile at idle conditions and showed that the distribution of individual acids was similar to that in the air, but the concentration was 17 times higher than for the average atmospheric content. Formic, acetic, and benzoic acids were detected as major species of used engine oil, but their content is negligible in new oil.

Synthesis of heterocyclic 1,4-dihydropyridylacetic acid derivatives of 6-aminopenicillanic acid and D-α-aminobenzylpenicillin with antibacterial activity

Condensation of 4-substituted 3-(4,4-dimethyloxazolin-2-yl)-1,4-dihydropyridylacetic acids 3 with the C-2 p-bromophenacyl esters of 6-aminopenicillanic acid salt 4 and D-α-aminobenzylpenicillin 8 afforded the respective 1,4-dihydropyridylacetic acid analogs 6 and 10 which exhibited moderate antibacterial activity toward Gram-positive strains. The 6-APA derivatives 6 exhibited dual 1H nmr resonances for H2′, H2, H5, and one of the C3-Me groups, suggesting the presence of two rotamers which differ in configuration at the carbonyl-to-nitrogen bond. The D-aminobenzylpenicillin derivatives 9 may exist in more than one conformation, one of which may be an intramolecular hydrogen bonded species such as 11.

Determination of C20-C30 fatty acids by reversed-phase chromatographic techniques: an efficient method to quantitate minor fatty acids in serum of patients with adrenoleukodystrophy.

An analytical method for the determination of saturated very long chain (VLC) fatty acids in the serum has been devised. Free fatty acids obtained after hydrolysis of total lipid extracts were converted into p-bromophenacyl esters. The derivatives were purified in two sequential steps by clean-up on C18 reversed-phase cartridge and fractionation by reversed-phase thin-layer chromatography (TLC), and then quantitated by high performance liquid chromatography (HPLC) analysis. This technique provides a reliable and alternative method for the biochemical identification of patients and carriers of an inherited metabolic disease characterized by the accumulation of saturated VLC fatty acids (C24-C26) such as Adrenoleukodystrophy (ALD). In four cases of diagnosed ALD the fatty acid composition of serum total lipids was dramatically enriched in saturated VLC fatty acids compared to controls. The ratio of hexacosanoic acid (C26:0) to docosanoic acid (C22:0) in ALD patients was approximately six-fold higher than that of healthy controls or patients affected by metabolic or neurological disorders other than ALD.

Liquid Chromatographic Method for Quantitative Determination of Free Fatty Acids in Butter

A liquid chromatographic method has been developed for the determination of free fatty acids in butter. The fatty acids are converted to the p-bromophenacyl esters, via a crown ether-catalyzed reaction, without separation from the other butter components. The esters are separated on a C18-bonded silica column by using an acetonitrile-water solvent gradient and quantitated using the ester of heptadecanoic acid as internal standard. C16 and C18:1 co-elute in the acetonitrile-water system but are separated using an isocratic methanol-acetonitrile-water system. Limits of detection range from 7 ng for butyric acid to 45 ng for linoleic acid. The average coefficient of variation (n = 10) for 10 free fatty acids from a butter was 5.83%.

Determination of C1−C4 fatty acids as p-bromophenacyl esters using glass-capillary gas chromatography and electron-capture detection

A method for the determination of low relative molecular mass carboxylic acids (C1−C4) in water is reported. The acids are converted to p-bromophenacyl esters prior to a glass-capillary gas chromatographic separation. By utilizing electron-caputre detection the detectability is substantially improved compared to flame-ionization detection. A comparison of three different ways to treat the water samples and to produce the derivatives is made. It is shown that the α, p-dibromoacetophenone reagent decomposes to a small extent which limits the utility of the reagent. Nevertheless a detection limit for formic acid of approximately 2.5 mgl−1 is obtained. The method is applied to the determination of formic and acetic acids in a paper kraft water sample.

High-performance liquid chromatographic analysis of short chain carboxylic acids as p-bromophenacyl esters: identification and separation of a decay product

High-performance liquid chromatographic analysis of short chain carboxylic acids as p-bromophenacyl esters: identification and separation of a decay product

Simplified determination of free fatty acids using derivatization technique by high-performance liquid chromatography through the modified extraction column.

A more practical and rapid determination of free fatty acids (FFA) by use of high-performance liquid chromatography (HPLC) is described. Column extraction of FFA by Extrelut® was performed instead of the conventional liquid-liquid extraction. The most suitable extraction conditions of FFA by the Extrelut® column were investigated. FFA were quantitatively converted to their p-bromophenacyl esters to enhance their UV detectability. p-Bromophenacyl esters of FFA were determined by HPLC using a reverse-phase column. The recovery of four kinds of FFA added to human normal control serum was between 97-103% in the range of 50-200 nmol of FFA. The FFA contents of healthy human sera were measured by the proposed method. The correlation between the HPLC method and either enzymatic or gas chromatographic methods was investigated.