N-caproic Acid Research Articles

Solubilities of pyruvic acid and the lower (C1-C6) carboxylic acids. Experimental determination of equilibrium vapour pressures above pure aqueous and salt solutions

Henry's law constantsK′ H (mol kg−1 atm−1) have been determined at 298.15 K for the following organic acids: formic acid (5.53±0.27×103); acetic acid (5.50±0.29×103); propionic acid (5.71±0.34×103);n-butyric acid (4.73±0.18×103); isobutyric acid (1.13±0.12×103); isovaleric acid (1.20±0.11×103) and neovaleric acid (0.353±0.04×103). They have also been determined fromT=278.15 K toT=308.15 K forn-valeric acid (ln(K′ H)=−14.3371+6582.96/T);n-caproic acid (ln(K′ H)=−13.9424+6303.73/T) and pyruvic acid (ln(K′ H)=−4.41706+5087.92/T). The influence of 9 salts on the solubility of pyruvic acid at 298.15 K has been measured. Pyruvic acid is soluble enough to partition strongly into aqueous atmospheric aerosols. Other acids require around 1 g of liquid water m−3 (typical of clouds) to partition significantly into the aqueous phase. The degree of partitioning is sensitive to temperature. Considering solubility and dissociation (to formate) alone, the ratio of formic acid to acetic acid in liquid water in the atmosphere (at equilibrium with the gas phase acids) is expected to increase with rising pH, but show little variation with temperature.

Determination of portal short-chain fatty acids in rats fed various dietary fibers by capillary gas chromatography.

A simple, rapid and sensitive capillary gas chromatographic method was investigated to measure portal short-chain fatty acids (SCFAs). A 20-microliters sample of portal plasma was denatured with sulfosalicylic acid and then extracted with diethyl ether before the removal of protein precipitate. The resultant extract was concentrated by a transfer to 50 microliters of 0.2 M NaOH, thus avoiding tedious further concentration steps. This reduced the sample volume to one-fourth. Since the ratio of acetic acid, a major SCFA, to other acids varies widely, ranging from 10-fold to 100-fold, acrylic and methacrylic acids were used as internal standards to simultaneously measure SCFAs having a carbon number of 2-6. As a result, good recovery (90.38-103.17%) and reproducibility (coefficient of variation 0.83-8.85%) were observed over a wide range. Furthermore, portal SCFAs in rats fed various dietary fibers were determined by the present method. We showed that the amounts not only of the major acids such as acetic acid and propionic acid, but also of the minor fermented products such as n-valeric acid and n-caproic acid, could be significantly changed by dietary manipulation. Thus, the present method is simple and reliable, and requires only a small amount of sample.

Experimental evidence for the existence of duoselective (Type III) enantiomer separations in the capillary electrophoretic analysis of chiral weak acids

Experimental evidence has been found for the existence of the theoretically predicted duoselective (Type III) enantiomer separations in the capillary electrophoretic (CE) analysis of chiral weak acids. The mobilities of the enantiomers of 3,5-dinitrobenzamido phenylalanine were determined in the pH 3–6 range using constant ionic strength ϵ-amino- n-caproic acid background electrolytes and β-cyclodextrin as resolving agent. The complex formation constants and ionic mobilities of the free and complexed enantiomers were determined using the multiple equilibria-based electrophoretic mobility model. The extended peak resolution equation of CE was then used to calculate the peak resolution surfaces as a function of the pH and the β-cyclodextrin concentration of the background electrolyte, the dimensionless electroosmotic flow coefficient, and the effective portion of the applied potential. Though the observed peak resolution values were lower than predicted by the theory due to the presence of electromigration dispersion, the predicted and observed loci of the extrema of the resolution surface agreed well. The desionoselective/ionoselective/duoselective (DID) separation selectivity model and the peak resolution model permit the rational optimization of the CE separation of weak acid enantiomers.

Effect of volatile fatty acids and trimethylamine on nitrification in activated sludge

The effect of volatile fatty acids and trimethylamine on the nitrification activity of activated sludge was studied in laboratory batch experiments. The critical concentration of inhibitor I K at which the activity ceases was determined by modelling. I K values for ammonia oxidation were found to be 6 mM for isobutyric, 37 mM for n-valeric, 6 mM for isovaleric 36 mM for n-caproic acid and 3 mM for trimethylamine. I K values for nitrite oxidation were found to be 2 mM for formic, 115 mM for acetic, 68 mM for propionic, 33 mM for n-butyric, 8 mM for isobutyric, 75 mM for n-valeric, 7 mM for isovaleric and 81 mM for n-caproic acid. For propionic acid and trimethylamine, the undissociated compound appears to act as an inhibitor. For formic, isobutyric and isovaleric acids, the degree of inhibition seems to be determined by the total concentration of acid. Experiments with anaerobically treated wastewater stripped of sulphide showed that volatile fatty acids and trimethylamine alone cannot account for the inhibition of the nitrification activity, indicating that other factors are also involved.

Comparative Cytotoxic Effects of Short-chain Fatty Acids Produced by Periodontal Pathogens on Two Cultured Fibroblast Lines

The cytotoxic effects of a range of short-chain fatty acids produced by anaerobic periodontopathic bacteria were compared using two types of fibroblast culture: Vero cells and human gingival fibroblasts. Morphological changes in cells exposed to 1.56—25 mM acetic, n-butyric, isobutyric, n-caproic, isocaproic, phenylacetic, propionic, n-valeric and isovaleric acids for three days were monitored using phase contrast microscopy and metabolic activity of cultures assessed by measuring reduction of the tetrazolium salt MTT. Vero cells were considerably more sensitive than human gingival fibroblasts to both morphological and metabolic changes. Reduction in metabolic activity of Vero cells was n-butyric > propionic, n-valeric and isovaleric > phenylacetic, n-caproic and isocaproic. N-butyric acid (≥ 12.5 mM) caused progressive cell rounding and death of >90 per cent of the cells by three days. Exposure to n-caproic, n-valeric and isovaleric acids (25 mM) produced spindle-shaped Vero cells with one or more long dendritic projections and reduced metabolic activity to ≤60 per cent of control values ( P <0.001). Comparable morphological changes were not observed in human gingival fibroblasts exposed to the maximum concentrations of any acid and only n-butyric, propionic and phenylacetic acids reduced metabolic activity significantly. Thus although Vero cells have traditionally been used to test for the cytotoxic effects of potential virulence factors from periodontal pathogens, their behaviour may not be representative of the cells normally exposed to these components in vivo . Keywords - Cytotoxicity, Short-chain fatty acids, Human gingival fibroblasts, MTT, Vero cells.

Determination of Organic Acids in Natural and Processed Foods by Isotacophoresis.

A simple and rapid analytical method was developed by using isotachophoresis for simultaneous determination of organic acids including citric, tartaric, malic, fumaric, lactic, succinic, ascorbic, acetic, aspartic, glucuronic acids in fresh fruits and processed juices, and in carbonated and fermented drinks. The detection limits of this method are about 0.04 to 0.1 mg within the coefficient of variation of ca. 6%, and the entire analytical procedure can be accomplished within 30 min without any pretreatment of samples. Using this method, a leading electrolyte solution was prepared with 7 parts of 0.1% polyvinyl alcohol in 0.01 N HCl, buffered at pH 3.0 with β alanine, and 3 parts of methanol. A solution of 0.01 N n-caproic acid was used as the terminal electrolyte. There was a two stage migration procedure : the first stage in a 0.1 mm i.d. ×10 cm tube with 250 μA current for 10 min and the second stage in a 0.05 mm i.d. × 15 cm tube with 100 μA current for 10 min. The oven temperature was kept at 25°C. The nitrogen carrier gas was introduced at 0.25 Kg/cm2 pressure.

3-Bromomethyl-7-methoxy-1,4-benzoxazin-2-one as a highly sensitive fluorescence derivatization reagent for carboxylic acids in high-performance liquid chromatography

A fluorescence labelling reagent, 3-bromomethyl-7-methoxy-1,4-benzoxazin-2-one (BrMB), was synthesized from 2-amino-5-methoxyphenol and ethyl pyruvate by the Wislecenus reaction followed by bromination. Picomole to nanomole amounts of saturated aliphatic fatty acids were converted into the corresponding fluorogenic esters in the presence of anhydrous potassium carbonate and a crown ether as catalysts. BrMB derivatives of fatty acids were separated on a reversed-phase column and were sensitively detected fluorimetrically at 440 nm with excitation at 345 nm. Quantitative studies revealed that n-caproic acid was esterified completely under mild conditions and with sufficient reproducibility. The detection limit of the BrMB derivative of n-caproic acid was just below 2 fmol.

Determining the extent of labeling for tetramethylrhodamine protein conjugates

A new, relatively simple, spectrophotometric technique has been developed which is useful for accurately determining the extent of chromophore labeling of proteins. Often the absorbance spectra and extinction coefficients of dye/protein conjugates are strongly affected by changes in the chromophore microenvironment that may occur at high dye/protein ratios. In the method being presented, the microenvironment effects have been significantly reduced by denaturing the dye/protein complex in 6 M guanidine hydrochloride prior to making the necessary spectrophotometric measurements. With this approach, extinction coefficients were obtained under native and denatured conditions for tetramethylrhodamine isothiocyanate (TRITC) when bound to a model protein receptor, the sugar binding protein concanavalin A (ConA). The extinction coefficients used for TRITC/ConA conjugates under native and denaturing conditions were 6.52 × 10 4 M −1 cm −1 and 6.96 × 10 4 M −1 cm −1, respectively. These values were obtained from a model dye complex formed between TRITC and ϵ-amino- n-caproic acid which closely resembles the sidechain of lysine residues. Additional dye/ConA conjugates were prepared with tetramethylrhodamine succinimidyl ester (RHS) and eosin isothiocyanate (EITC), and the effects of microenvironment changes on these conjugates were examined. Extinction coefficients for these dyes in native and denaturing conditions, as a function of the degree of labeling, were not appreciably different indicating that changes in the microenvironment did not have a significant affect on the spectral properties of these two dyes. In summary, with this new approach it is quite easy to accurately determine the dye/protein ratio for TRITC conjugates. Also, it is expected that RHS would be a better dye than TRITC for protein conjugation because more accurate values for dye/protein ratios can be obtained under native conditions.

A novel process of cyclodextrin production by the use of specific adsorbents: Part I. Screening of specific adsorbents

Novel adsorbents that are composed of ligand, spacer and support were chemically synthesized and the two consecutive screenings of them made it possible to determine the adsorbents that were most recommendable for α- and β-cyclodextrin (CD) production. First, ligands of high adsorption selectivity for each CD were screened from among the candidates (carboxylic acids) that are tied in ionic bonds to two types of strongly basic anion exchange resins as support. Secondly, ligand derivatives (as ligand and spacer) were bound in covalent bonds onto chilosan beads as support and then, the most suitable spacer length for the CDs' adsorption selectivity and capacity were investigated. An optimal mol ratio of ligand to amino group of the beads was also examined. Stearic acid was the most effective ligand for α-CD, whereas cyclohexanepropanamide- n-caproic acid was the best for β-CD. Adsorption selectivity of adsorbents derived from carboxylic acids (stearic and/or palmitic) and chitosan beads was almost 100%, while their adsorption capacities were large enough to meet the demand for economic production and purification of CDs on an industrial scale.

Sensitive fluorescence labelling for analysis of carboxylic acids with 4-bromomethyl-6,7-methylenedioxycoumarin

A fluorescence labelling reagent, 4-bromomethyl-6,7-methylenedioxycoumarin (BrMDC), was synthesized from sesamol and citric acid by Pechman condensation followed by bromination. Nanomole amounts of saturated aliphatic fatty acids were converted into the corresponding fluorogenic esters in the presence of anhydrous potassium carbonate and a crown ether as a catalyst and were separated by reversed-phase high-performance liquid chromatography (HPLC). Quantitative studies revealed that n-caproic acid was esterifted completely at low temperature and with sufficient reproducibility. The detection limit was just below 15 fmol per injection at a signal-to-noise ratio of 3. The fluorescence quenching of the BrMDC derivative was the lowest in conventional mixed solvent systems in comparison with those of two previously reported coumarin compounds. BrMDC was also applied to the simultaneous analysis of some acidic non-steroidal anti-inflammatry agents by reversed-phase HPLC.

Gas chromatographic analysis of the carboxylic acid composition of valeriana extracts

The gas chromatographic analysis for the qualitatived and quantitative analysis of carboxylic acids present in valeriana extracts in the form of their n-butyl and trifluoroacetylated n-butyl esters is presented. On the basis of the proposed method the amount of four valtrate species, as well as the free carboxylic acid content of valeriana tinctures can be calculated. It is shown that extracts of Valeriana officinalis obtained by using ethanol, acetone, or chloroform as the solvent contain the same carboxylic acids. The determined components, are: formic, acetic, propionic, isobutyric, n-butyric, lactic, isovaleric, n-valeric, 2-hydroxy-isovaleric, isocaproic, n-caproic, caprylic, malic, capric, lauric, myristic, valerenic, palmitic, stearic, arachic, behenic, erucic and lignoceric acids.

Quantitation of sodium mercaptoundecahydrododecaborate by capillary isotachophoresis

A simple, specific and accurate method for the quantitation of sodium mercaptoundecahydrododecaborate, which is a very hygroscopic agent for boron neutron-capture therapy, was developed by using capillary isotachophoresis with sodium p-toluenesulphonate as an internal standard. Compound 1a was separated and identified using 0.01 M hydrochloric acid and β-alanine as the leading electrolyte and 0.01 M n-caproic acid as the terminating electrolyte. It could be resolved well from the oxidation products, sodium sulphidobis (undecahydrododecarborate) and sodium sulphidobis (undecahydrododecarborate) S-oxie, and the detection limit was 0.7 μmol/ml.

Isotachophoretic analysis of mercaptoundecahydrododecaborate anion in human serum and urine.

An isotachophoretic assay method was developed for sodium mercaptoundecahydrododecaborate (1), Na210 B12H11SH, which is used for boron neutron-capture therapy. The method is simple, specific and suitable for the determination of 1 as an anion in human serum and urine. Compound 1 was separated from serum and urine constituents with 0.01 M hydrochloric acid and β-alanine (leading electrolyte, pH 3.70) and 0.01 M n-caproic acid (terminating electrolyte). The recoveries of 1 from serum and urine were about 85 and 101%, respectively. The detection limit was 7×10-7 mol/ml in serum and urine. The procedure could also detect the oxidation product, sodium di (thioundecahydrododecaborate), at 5×10-7 mol/ml or more in urine. The present method was used to determine the levels of 1 in urine samples of patients with malignant brain tumor being treated by boron neutron-capture therapy.

Hydroxy acid metabolites of branched-chain amino acids in amniotic fluid

A method is described in which ammonia chemical ionization gas chromatography-mass spectrometry was utilized in the selected ion monitoring mode to provide an accurate, selective approach to the quantification in amniotic fluid of a number of hydroxylated organic acids derived from the metabolism of the branched-chain amino acids. 2-Hydroxy- n-caproic acid was employed as an internal standard and the hydroxy acids were isolated from amniotic fluid by liquid partition chromatography and the trimethylsilyl derivatives were quantified. Normal values have been obtained for 2-hydroxyisovaleric acid, the sum of 2-hydroxyisocaproic acid and 2-hydroxy-3-methylvaleric acid, 2-methyl-3-hydroxybutyric acid, 3-hydroxyisovaleric acid and 2-ethyl-3-hydroxypropionic acid. The method also provides data on the concentration of methylmalonic acid. The concentration of 2-hydroxyisovaleric acid was not useful in the prenatal diagnosis of a fetus with maple syrup urine disease. Elevated concentrations of 2-methyl-3-hydroxybutyric acid as well as methylmalonic acid were found in the amniotic fluid of two fetuses with methylmalonic acidemia.

Short-Chain Fatty Acids in the Normal Human Feces

The short-chain fatty acids ( SCFAs ) have been studied in the feces of 20 healthy subjects--10 methane excretors and 10 non-methane excretors. The analytical procedure included homogenization of fecal samples followed by vacuum distillation and subsequent gas chromatography. This method for analysis of fecal SCFAs showed recoveries of the individual acids from 90% to 109% and coefficients of variation for the inter-assay reproducibility from 6.0% to 19.7%, highest for those acids present in the smallest concentrations. There was no difference in the concentrations or relative compositions of SCFAs between methane-excreting subjects and non-methane-excreting subjects. The concentrations of SCFAs , given as mmol/kg feces (wet weight), were (median and range): total, 76.8 (27.9-187.7); acetic acid, 37.4 (12.8-103.4); propionic acid, 12.5 (4.5-27.8); i-butyric acid, 2.2 (0.7-3.8); n-butyric acid, 12.4 (4.0-53.0); i-valeric acid, 3.2 (0.8-5.9); n-valeric acid, 2.4 (0.6-3.8) and n-caproic acid, 0.5 (0.0-3.6). The study shows that the SCFAs are quantitatively the most important anions in the feces of healthy subjects. The pronounced individual variations in the concentrations of SCFAs are real biological variations and cannot be explained by methodological variations.

Decreased Fc and C3 receptor function in macrophage populations which are refractory to migration inhibitory factor, C3 activators, and immune complex

Guinea pig macrophage populations previously established to be either responsive or refractory to activation by migration inhibitory factor (MIF) and Lotus fucolectin in the macrophage migration inhibition (MMI) assay were further characterized for their MMI response to diverse effectors as correlated with their Fc and C3b receptor function. MIF-refractory populations were found to be uniformly unresponsive to the complement activators: bacterial lipopolysaccharide, cobra venom factor, zymosan, and immune complex. MIF-responsive macrophages were responsive to the same activators. Fc-mediated binding and phagocytosis of IgG-coated sheep erythrocytes (EA) were markedly depressed in freshly harvested refractory macrophages as compared to responsive cells. Fc phagocytosis by refractory populations increased rapidly during 24–28 hr in vitro culture to levels equal to that of responsive cells which corresponded with an increase in their MMI response to MIF. Refractory macrophages also had decreased C3b receptor function as shown by reduced binding and phagocytosis of EAC or serum-coated zymosan and displayed a greater loss in C3b binding capacity than responsive cells during 48 hr in vitro culture. Trypsinization of responsive macrophages rendered them refractory in their MMI response to the various activators and selectively reversed C3b-dependent binding without effect on Fc binding. The plasmin esterase inhibitors, ϵ-amino- n-caproic acid, tranexamic acid, and l-lysine, previously established to reverse the MMI response to MIF, FBP, and C3 activators were found to inhibit both Fc- and C3-dependent phagocytosis. These results indicate that macrophage populations which are refractory to migration inhibition by MIF and C3 activators also have reduced Fc- and C3b-mediated phagocytic functions as compared to more mature responsive populations.

A new esteroproteinase (proteinase F) from the submandibular glands of female mice

One of the esteroproteinases present in the submandibular glands of female mice was purified and characterized. The enzyme, designated proteinase F in this report, had a p I value of 4.6 and a molecular weight of 27 600, being comprised of two subunits of 10 000 and 18 000 daltons. The amino acid composition of proteinase F resembled that of the epidermal growth factor-binding protein, but antiserum against proteinase F only reacted weakly against the binding protein. Proteinase F had an optimum pH at around 9.0 and was strongly inhibited by Cu 2+ and Hg 2+ (42 and 76% inhibition, respectively, at a concentration of 4·10 −6 M). It was also inhibited by aprotinin, phenylmethylsulfonylfluoride, iodoacetamide, leupeptin, antipain, and benzamidine but neither by trypsin inhibitors from pancreas, soybean or ovomucoid, nor by TLCK, TPCK, and ϵ-amino- n-caproic acid. Although its actual physiological function has yet to be determined, these properties indicate that proteinase F is a new enzyme, being distinguished from known proteinases, kallikrein, plasmin, trypsin, chymotrypsin, tonin, angiotensin-converting enzyme, proteinase A (β-nerve growth factor endopeptidase), proteinase D (epidermal growth factor-binding protein), P-esterase, renin A, and renin C. Proteinase F was present in the submandibular glands of female mice more abundantly than in those of males, but it increased in males following castration. Thus, proteinase F appears to be affected by male hormones in vivo.

Measurement of adenosine triphosphate and some other metabolites in blood cells by isotachophoresis. I. Preparative technique and enzymatic confirmation.

This study investigated the optimal conditions for detection of nucleotides in blood using an IP-1B capillary isotachophoretic apparatus. The system used 10 mM HCl-beta-alanine (pH 4.2) as the leading electrolyte and n-caproic acid as the terminal electrolyte. Direct application of lysed red blood cells was shown to be inaccurate, and a method of deproteinization based on heat in a microwave oven was developed. The zones for 2,3-diphosphoglycerate, ATP, inorganic phosphate, and lactate were identified enzymatically by withdrawal of pure samples of each zone via a special withdrawal cell. The quantitative values obtained by isotachophoresis were also confirmed enzymatically. The technique is now available for convenient and accurate identification of these metabolites simultaneously.

Macrophage migration inhibition by complement activators, migration inhibitory factor, and Lotus fucolectin: Evidence for common involvement of cell-associated esterase activity

Activators of the complement pathway were compared with migration inhibitory factor (MIF) and Lotus fucolectin, which mimics MIF, for their macrophage migration inhibitory (MMI) activity. Endotoxin (LPS), cobra venom factor (CVF), and zymosan, known complement (C3) activators, were found to produce dose-dependent MMI activity, similar to MIF, independent of requirements for nonadherent lymphocytes and serum complement. Comparable activity was observed by all migration inhibitors in the presence of freshly harvested unheated or heat-inactivated guinea pig serum as well as zymosan-adsorbed (C3-depleted) serum. Iscove's serum-free medium also promoted migration inhibition confirming a lack of requirement for serum complement in the reaction. Polymyxin B reversed MMI by LPS, but had no effect on the other inhibitors, indicating that the MMI activity of CVF, zymosan, MIF, and Lotus fucolectin was not primarily due to LPS contamination. Peritoneal macrophages (PM), depleted of nonadherent lymphocytes, responded as well as unpurified PM to the complement activators and MIF. ϵ-Amino- n-caproic acid (EACA), l-lysine, and tranexamic acid (TA), known inhibitors of the fibrinolytic activity of plasminogen activator (PA), were found to reverse migration inhibition of C3 activators, MIF, and Lotus fucolectin. In contrast bovine pancreatic trypsin inhibitor (BPTI) and soybean trypsin inhibitor (STI) had no effect on MMI activity. These results suggest a common mechanism for mediation of migration inhibition by complement activators and MIF which may involve activation of cell-associated complement to produce esterolytic end products capable of triggering the activation process.

Quantitative analysis of trifluoroacetate in the urine and blood by isotachophoresis.

Trifluoroacetate (TFA), the major metabolite of halothane, was assayed by a newly developed isotachophoretic technique. This technique has several advantages over the presently used methods of analysis. It requires no special preparation of urine or blood samples. The sample volume is small (5--100 microliters) and the analysis time is short (30--90 min per sample). In addition, the method provides an analysis that is both qualitative and quantitative over a wide range of concentrations (from 2 nanomoles in 200 microliters to 200 nanomoles in 5 microliters). In this study, the assay was performed using HCl (0.001 M) in 1 per cent Triton X-100, titrated with beta-alanine to a pH value of 3.6--3.9 as the leading electrolyte and n-caproic acid (0.01 M) as the terminal electrolyte (50--100 muA migration current). Using this technique, daily urinary TFA excretion of seven patients was measured during halothane anesthesia and for 14 days postoperatively. The TFA values were highest on the second postoperative day (317--1,259 mg). The mean values of the urinary TFA excreted during the entire study (2,501 +/- 493 mg, mean +/- SEM) were much higher than those reported previously. The isotachophoretic technique provides a sensitive assay for future research into the biotransformation of halothane.