Synthesis of mono- and diglucosiduronates of metabolites of deoxycorticosterone and corticosterone and analysis by a new mass spectrometric technique

By condensing 3 alpha,21-dihydroxy-5 beta-pregnan-20-one, or its appropriate monoacetate, with methyl 2,3,4-tri-O-acetyl-1-bromo-1-deoxy-alpha-D-glucuronate in the Koenigs-Knorr reaction beta-D-glucosiduronates 10, 4, and 7 were obtained as polyacetate methyl esters. Alkaline hydrolysis of these substances cleaved the ester groups and gave the corresponding steroidal glucosiduronic acids 12, 6 and 8. Upon treatment with diazomethane, these acids produced the equivalent methyl esters. The C-3, the C-21 and the C-3,21 glucosiduronates of 3 alpha,21-dihydroxy-5 beta-pregnan-11,20-dione were prepared by previously reported methods and converted into the corresponding C-20 semicarbazones (14, 20 and 26). With C-20 stabilized by the semicarbazone group against reduction, it was possible to reduce the 11-oxo function in these substances to an 11 beta-hydroxyl group; after removal of the semi-carbazone moiety from these products at pH 2.0, glucosiduronic acids 18, 22 and 28 were obtained. The mass spectra of a representative group of the mono- and diglucosiduronic acids and esters were determined by utilizing fast atom bombardment and monitoring ions in both positive and negative modes of operation.     

Extraction of steroidal glucosiduronic acids from aqueous solutions by anionic liquid ion-exchangers

A pilot study on the extraction of three steroidal glucosiduronic acids from water into organic solutions of liquid ion-exchangers is reported. A single extraction of a 0.5mm aqueous solution of either 11-deoxycorticosterone 21-glucosiduronic acid or cortisone 21-glucosiduronic acid with 0.1m-tetraheptylammonium chloride in chloroform took more than 99% of the conjugate into the organic phase; under the same conditions, the very polar conjugate, beta-cortol 3-glucosiduronic acid, was extracted to the extent of 43%. The presence of a small amount of chloride, acetate, or sulphate ion in the aqueous phase inhibited extraction, but making the aqueous phase 4.0m with ammonium sulphate promoted extraction strongly. An increase in the concentration of ion-exchanger in the organic phase also promoted extraction. The amount of cortisone 21-glucosiduronic acid extracted by tetraheptylammonium chloride over the pH range of 3.9 to 10.7 was essentially constant. Chloroform solutions of a tertiary, a secondary, or a primary amine hydrochloride also will extract cortisone 21-glucosiduronic acid from water. The various liquid ion exchangers will extract steroidal glucosiduronic acid methyl esters from water into chloroform, although less completely than the corresponding free acids. The extraction of the glucosiduronic acids from water by tetraheptylammonium chloride occurs by an ion-exchange process; extraction of the esters does not involve ion exchange.     

Recovery of steroidal glucosiduronic acids from organic solvents containing anionic liquid ion-exchangers

Solutions of anionic liquid ion-exchangers in organic solvents are potentially useful for extracting steroidal glucosiduronic acids from biological fluids and for purifying mixtures of these acids by chromatography. If a glucosiduronic acid is to be isolated in pure form after either of these procedures, it is necessary to separate it from the ion-exchanger. Separation from an organic solution of tetraheptylammonium chloride may be accomplished by extraction with water under the following conditions, which promote transfer of a glucosiduronate to the aqueous phase: (1) an appropriate solvent (diluent) as the organic phase, (2) the presence in the two-phase mixture of an anion such as myristate or dodecyl sulphate to combine with the tetraheptylammonium ion, and (3) an increase of the pH of the aqueous phase in association with the presence of myristate or dodecyl sulphate. The foregoing factors apply also to removal of glucosiduronates from organic solutions of ion exchangers that are hydrochlorides of tertiary, secondary, or primary amines. Since these amines exert progressively less solubilizing effect for glucosiduronates as the pH of the aqueous phase is increased, the conjugates can be released from the organic phase by adjusting the pH to 10 and omitting the myristate or dodecyl sulphate.     

Phenolic O-methyltransferase from Lentinus lepideus (basidiomycete)

The fungus, Lentinus lepideus, produces crystalline methyl p-methoxycinnamate in stationary cultures. O-methylation and methyl ester formation of hydroxycinnamic acids were examined with enzyme preparations of the fungus. Using S-adenosylmethionine-¹⁴CH3, it was found that only the methyl esters of the hydroxycinnamic acids are substrates for O-methylation and not the free acids. Benzoic acids and their methyl esters are not substrates. The activity of the enzyme is p-specific and its specific activity decreases with increasing number of hydroxyl groups in the substrate. The same enzyme preparations catalyze the formation of the methyl ester of cinnamic acid from the free acid.     

Synthesis of cinnamoyl and hydroxycinnamoyl amino acid conjugates and evaluation of their antioxidant activity.

Fifteen amides of cinnamic, ferulic and sinapic acids with natural and unnatural C-protected amino acids have been synthesized. The amides (E)-N-(feruloyl)-L-tyrosine methyl ester (10), (E)-N-(feruloyl)-L-phenylalanine t-butyl ester (11), (E)-N-(sinapoyl)-L-tyrosine methyl ester (13) and (E)-N-(sinapoyl)-L-phenylalanine t-butyl ester (15) with a free carboxyl group of amino acids have been found in nature. The rest of the compounds are unknown. The hydroxycinnamoyl amino acid conjugates have been studied for their antioxidant activity (AOA) in bulk phase lipid autoxidation. The highest AOA has been found for the compounds 11 and 15, which contain the same phenylalanine moiety.     

In situ lipase-catalyzed reactive extraction of oilseeds with short-chained dialkyl carbonates for biodiesel production

Dimethyl/diethyl carbonate was adopted as extraction solvent and transesterification reagent at the same time for in situ lipase-catalyzed reactive extraction of oilseeds for biodiesel production in this work. Fatty acid methyl esters and ethyl esters were respectively obtained with higher yields than those achieved by conventional two-step extraction/transesterification. The augment ranged from 15.7% to 31.7%. The key parameters such as solvent/seed ratio and water content were further investigated to find their effects on the in situ reactive extraction. The highest yields of Pistacia chinensis Bunge methyl ester, P. chinensis Bunge ethyl ester, Jatropha curcas L methyl ester and J. curcas L ethyl ester could attain 89.6%, 90.7%, 95.9% and 94.5%, respectively under the optimized conditions.     

Identification of ornithine and arginine conjugates of cholic acid by mass spectrometry.

Nalpha-Cholylornithine, -arginine, and -histidine were prepared according to a method previously employed for the chemical synthesis of the monoamino acid conjugates of bile acids. The products were shown to involve the alpha amino group of the dibasic amino acids by examination of the mass spectra of the original compounds, their lactams, their methyl esters and the methyl ester acetates. Only the methyl ester acetates gave detectable amounts of molecular ion. The free acids and the methyl esters of Nalpha-cholylornithine and -arginine gave identical lactams upon sublimation from the direct insertion probe. The synthetic Nalpha-cholylarginine was shown to yield a mass spectrum identical to that of an arginocholic acid recovered from the bile of an isolated perfused rat liver.     

Determination of monoenoic fatty acid double bond position by permanganate-periodate oxidation followed by high-performance liquid chromatography of carboxylic acid phenacyl esters

This investigation was carried out to develop methods for a reverse-phase, high-performance liquid chromatography analysis of the monocarboxylic and dicarboxylic acids produced by permanganate-periodate oxidation of monoenoic fatty acids. Oxidation reactions were performed using [U-14C]oleic acid and [U-14C]oleic acid methyl ester in order to measure reaction yields and product distributions. The 14C-labeled oxidation products consisted of nearly equal amounts of monocarboxylic and dicarboxylic acid (or dicarboxylic acid monomethyl ester), with few side products (yield greater than 98%). Conversion of the carboxylic acids to phenacyl esters proceeded to completion. HPLC of carboxylic acid phenacyl esters was performed using a C18 column with a linear solvent gradient beginning with acetonitrile/water (1/1) and ending with 100% acetonitrile. Excellent resolution was achieved for all components of a mixture of C5 through C12 monocarboxylic acid phenacyl esters and C6 through C11 dicarboxylic acid phenacyl esters. Resolution was also achieved for all components of a mixture of C5 through C12 monocarboxylic acid phenacyl esters and C6 through C11 dicarboxylic acid monomethyl, monophenacyl esters. The resolution obtained by HPLC demonstrates that, for a wide range of monoenoic fatty acids, both products of a permanganate-periodate oxidation can be identified on a single chromatogram. Free fatty acids and fatty acid methyl esters were analyzed with equal success. Neither the oxidation nor the esterification reaction caused detectable hydrolysis of methyl ester. The method is illustrated for free acids and methyl esters of 14:1 (cis-9), 16:1 (cis-9), 18:1 (cis-6), 18:1 (cis-9), and 18:1 (cis-11).     

云南使君子仁油化学成分的GC-MS分析

以常规溶剂萃取得使君子仁油,取两份油,一份经甲酯化处理,别一份不甲酯化,然后采用重量法和气相色谱-质谱联用技术分别测定使君子仁油含量和脂肪酸成分。结果表明：使君子仁油含量为15％;从甲酯化脂肪油中共检测出5种成分,其中E-9-十八烯酸占脂肪酸总量的46．99％,十六烷酸甲酯占脂肪酸总量的28．25％;另外,从未甲酯化脂肪油中共检测出7种成分,其中含防十八烯酸63．19％,十六烷酸甲酯15．26％,同时还检测出11．79％的γ-生育酚。使君子仁油是具有抗氧化性的植物源脂肪油,是开发和利用E-9-十八烯酸,十六烷酸甲酯和γ-生育酚的理想原料,在食用、医疗保健方面具有巨大潜力和广阔前景。     

Synthesis of thromboxane B2 metabolites

This paper reports the synthesis of 11-dehydrothromboxane B₂ methyl ester (II), 15-dehydrothromboxane B₂ methyl ester (III), 15-dehydro-13,14-dihydrothromboxane B₂ (XII) and 2,3-dinorthromboxane B₂ methyl ester (XV). These compounds, as their free acids, have been reported to be thromboxane metabolites.     

Separation of methylated free bile acids from their taurine and methyl glycine conjugates by thin-layer chromatography

Class separation of methylated free bile acids from bile acids conjugated with taurine and methylglycine was accomplished using a solvent system of 2,2,4-trimethylpentane-absolute ethanol 10:1 (v/v). By developing a silica thin-layer plate two times with solvent in a Brinkmann sandwich tank, the difficult resolution between methyl cholate and methyl glycolithocholate was achieved. Evidence is presented that this separation system may be useful as a preparative step in the analysis of bile acids by gas-liquid chromatography or high pressure liquid chromatography.--Bolt, M. J. G. Separation of methylated free bile acids from their taurine and methyl glycine conjugates by thin-layer chromatography.     

Highly efficient transformation of waste oil to biodiesel by immobilized lipase from Penicillium expansum

An inexpensive self-made immobilized lipase from Penicillium expansum was shown to be an efficient biocatalyst for biodiesel production from waste oil with high acid value in organic solvent. It was revealed that water from the esterification of free fatty acids and methanol prohibited a high methyl ester yield. Adsorbents could effectively control the concentration of water in the reaction system, resulting in an improved methyl ester yield. Silica gel was proved to be the optimal adsorbent, affording a ME yield of 92.8% after 7 h. Moreover, the enzyme preparation displayed a higher stability in waste oil than in corn oil, with 68.4% of the original enzymatic activity retained after being reused for 10 batches.     

Synthesis and characterization of vegetable oil derived esters: evaluation for their diesel additive properties

Trans-esterification of four vegetable oils; canola oil, greenseed canola oil from heat-damaged seeds, processed waste fryer grease and unprocessed waste fryer grease, was carried out using methanol, and KOH as catalyst. The methyl esters of the corresponding oils were separated from the crude glycerol, purified, and characterized by various methods to evaluate their densities, viscosities, iodine values, acid numbers, cloud points, pour points and gross heat of combustion, fatty acid and lipid compositions, lubricity properties, and thermal properties. The fatty acid composition suggests that 80-85% of the ester was from unsaturated acids. Substantial decrease in density and viscosity of the methyl esters compared to their corresponding oils suggested that the oils were in their mono or di glyceride form. The lubricity of the methyl esters, when blended at 1 vol% treat rate with ISOPAR M reference fuel, showed that the canola methyl ester enhanced the fuel's lubricity number. From the analyses performed, it was determined that the ester with the most potential for being an additive or a substitute for diesel fuel is the canola methyl ester, whose physical and chemical characteristics are similar to diesel fuel.     

Solvent effects on lipase-catalyzed esterification of glycerol and fatty acids

The lipase-catalyzed acylglycerol synthesis with fatty acids of different chain length is studied. Measured ester mole fractions at equilibrium are compared with calculated mole fractions. For these calculations the computer program TREP (Two-phase Reaction Equilibrium Prediction) is used. This program is based on the UNIFAC group contribution method and is developed for nondilute two-phase reaction systems.With one set of equilibrium constants, namely 1.3, 0.8, and 0.6 for monoester, diester, and triester synthesis, respectively, the equilibrium position of the reaction between glycerol and all saturated fatty acids with a chain length from 6 to 18 and oleic acid (cis-9-octadecenoic acid) can be calculated. Deviations, expressed as the ratio between calculated and measured ester mole fractions, usually were between 0.7 and 1.2. In the presence of solvents, the deviations of the monoester mole fractions were higher and rose up to 3. Without addition of a solvent, the ester mole fractions at equilibrium are dependent on the fatty acid chain length. With the short-chain hexanoic acid, the monoester mole fraction is the highest ester mole fraction, while for the long-chain oleic acid, the diester mole fraction is the highest one. The ester mole fractions become independent on the chain length of the fatty acid with a solvent added in a sufficient high concentration. Both reactions, with saturated and unsaturated C(18) fatty acids, lead to the same equilibrium position. The program TREP is found to make good predictions of the equilibrium amounts of ester and fatty acid. However, systematic deviations arise between measured and calculated amounts of water and glycerol in the organic phase. The calculated water and glycerol amounts are always lower than the measured ones. These deviations seem to be highest in nonpolar media and are probably due to deficiencies in the UNIFAC calculation method. Some preliminary experiments show the effect of the choice of solvent on the reaction rates. In polar solvents, the monoester production rate is enhances by a factor of 1.5 as compared to the reaction rate in a system without solvent. (c) 1993 John Wiley & Sons, Inc.     

Preparation of S-( )-ketoprofen by coupling the enantioselective hydrolysis of ketoprofen methyl ester with the photo-oxidation of methanol

A novel method for preparation of S-(+)-ketoprofen is presented involving coupling enantioselective hydrolysis of ketoprofen methyl ester catalyzed by a surfactant-coated-lipase with the photo-oxidation of methanol in a water-saturated organic solvent. The effect of photocatalytic conversion of methanol into water and carbon dioxide on the hydrolysis of ketoprofen methyl ester and the stability of the enzyme was investigated. The photo-oxidation of methanol shifted the equilibrium of the hydrolysis toward the formation of ketoprofen, increasing the equilibrium conversion ratio and improving the enantioselectivity. Because the surfactant-coated lipase and ketoprofen methyl ester dissolved in the organic solvent and ketoprofen was absorbed on the TiO₂ photocatalyst particles, the separation procedures could be simplified and the stability of the enzyme was increased.     

Preparation of S-(+)-ketoprofen by coupling the enantioselective hydrolysis of ketoprofen methyl ester with the photo-oxidation of methanol

A novel method for preparation of S-(+)-ketoprofen is presented involving coupling enantioselective hydrolysis of ketoprofen methyl ester catalyzed by a surfactant-coated-lipase with the photo-oxidation of methanol in a water-saturated organic solvent. The effect of photocatalytic conversion of methanol into water and carbon dioxide on the hydrolysis of ketoprofen methyl ester and the stability of the enzyme was investigated. The photo-oxidation of methanol shifted the equilibrium of the hydrolysis toward the formation of ketoprofen, increasing the equilibrium conversion ratio and improving the enantioselectivity. Because the surfactant-coated lipase and ketoprofen methyl ester dissolved in the organic solvent and ketoprofen was absorbed on the TiO₂ photocatalyst particles, the separation procedures could be simplified and the stability of the enzyme was increased.     

A series of lysyldipeptide derivatives for racemization studies in peptide synthesis.

The series of diastereomeric peptide derivatives N-benzoyl-D,L-X-N epsilon-benzyloxycarbonyl-L-lysine methyl ester where X = alanyl, valyl, leucyl, phenylalanyl and isoleucyl are submitted as model systems for studying racemization in peptide synthesis. The diastereomers can be analyzed by quantitation of the separated ester methyl proton peaks of their nuclear magnetic resonance spectra. Data on the tendency to racemize of the different residues are presented. In polar solvents, valyl and isoleucyl residues racemize more readily than the other residues.     

The biology of methyl ketones

Examples of the biological occurrence of methyl ketones are reviewed. The lack of significant accumulations of these compounds in the biosphere indicates that a recycling of these organic molecules is occurring. Evidence for biodegradation of acetone by mammals and longer methyl ketones by microorganisms via terminal methyl-group oxidation is discussed. A new mechanism for the subterminal oxidation of methyl ketones by microorganisms is proposed whereby the first intermediate produced is an acetate ester which subsequently is cleaved to acetate and a primary alcohol two carbons shorter than the original ketone substrate. Methyl ketones can be produced by mammals and fungi by decarboxylation of beta-keto acids. Some bacteria are able to form methyl ketones via the oxidation of aliphatic hydrocarbons at the methylene carbon alpha to the methyl group. Speculations on the biosynthesis of methyl ketones by insects and plants and a discussion of the possible biological roles of methyl ketones in diverse biological systems are presented.     

Feeding stimulants eliciting the probing behavior for Peregrinator blannulipes Montrouzier et Signore (Hemiptera: Ruduviidae) from Tribolium confusum (Jacquelin du Val)

Four fatty acid methyl esters identified in the solvent extract of Tribolium confusum (Jacquelin du Val) larvae as kairomones were individually and collectively tested for probing behavior of Peregrinator biannulipes Montrouzier et Signoret. All identified fatty acid methyl eaters, methyl palmitate, methyl linolate, methyl oleate and methyl stearate, exhibited characterisitic kairomonal probing behavior of P. biannulipe toward the lure. These fatty acid methyl ester were active at 0.2 microg/lure but a synergistic effect was not observed among them. Commercially available C8-C14 even-numbered fatty acid methyl esters that were not detected in the extract of T. confusum larvae also elicited a probing behavior but their activities were weaker than those of four fatty acid methyl ester (C16:0, C18:0, C18:1 and C18:2) identified in the extract. On the other hand, C17 and C19 odd-numbered fatty acid methyl esters did not show any activity at all.     

Fatty Acid Profiling of Commercially Important Raw and Boiled Seer Fish <Emphasis Type="Italic">Scomberomorus commerson</Emphasis> (Lacepede, 1800) (Scombridae,Teleostei, Pisces)

The present study aims to determine the fatty acid profiling of commercially important fresh and boiled Scomberomorus commerson. Fatty acids in fresh and boiled fish were separated and quantitatively determined by gas chromatography–mass spectrophotometer using standard methods. The findings revealed that the predominant fatty acids in fresh S. commerson were octadecanoic acid methyl ester, octanoic acid, 1,2-benzenedicarboxylic acid and 3-cyclopentylpropionic acid representing respectively 45.91, 5.69, 6.75 and 8.65% of total fatty acids. Boiled S. commerson showed predominant changes in their fatty acid profiles. In the omega-3 and omega-6 families the dominant fatty acids were doconexent, 3-cyclopentylpropionic acid, octadeconoic acid methyl ester and Hexadecane representing respectively 3.87, 12.08, 44.26 and 3.11% of total fatty acids. After boiling, some fatty acids present in fresh fish are damaged and formed new fatty acids which belonged to ω-3 polyunsaturated fatty acids (PUFA). Boiling increased the concentration of PUFAs from 73.25 to 80.37% of total fatty acids and also formed new fatty acids.