Preparation and purification of pteroic acid from folic acid

Crude pteroic acid, obtained by microbiological degradation of folic acid, has been purified by column chromatography on cellulose CF11, eluting with 0.1 M glycine buffer of pH 10.0, containing 0.15% w/v ascorbic acid and saturated with isoamyl alcohol.     

Hydrolysis of glycocholic acid by fungi

Summary A range of fungi have been investigated for their ability to hydrolyse glycocholic acid to yield cholic acid. Using thin-layer chromatography the majority of fungi tested have been shown to possess this ability. In the case of Cochliobolus intermedius IMI 52980 the product of hydrolysis has been isolated and characterised as cholic acid. The same organism has been demonstrated to grow using glycocholic acid as the sole source of carbon and nitrogen.     

Enzymatic oxidation of mercaptoethanol to isethinic acid and isethionic acid.

The enzymatic oxidation of mercaptoethanol by purified cysteamine oxygenase has been studied. Products were identified by chromatography as isethinic acid (2-hydroxyethan-sulfinic acid) and isethionic acid. Other features of the reaction, as cofactor requirement, the influence of the enzyme concentration on the stage of oxidation, the biological significance of this reaction are also discussed.     

Quantitative analysis of ascorbic acid and dehydroascorbic acid by high-performance liquid chromatography

High-performance liquid chromatography with spectrophotometric detection has been used to separate and quantitate ascorbic acid and dehydroascorbic acid. These components of vitamin C are resolved on a Lichrosorb-NH₂ column. The technique is capable of quantitatively following oxidation of ascorbic acid to dehydroascorbic acid and the reverse reduction. The technique is demonstrated to be suitable for assay of vitamin C in biological samples, foods, and pharmaceutical vitamin preparations.     

1,4-Thiomorpholine-3,5-dicarboxylic acid, a novel cyclic imino acid detected in bovine brain

Gas-liquid chromatography of enriched bovine brain extract revealed the occurrence of several sulfur-containing compounds. By co-chromatography with authentic product and by mass-spectrometric analysis, one of these compounds has been identified as 1,4-thiomorpholine-3,5-dicarboxylic acid (TMDA). The possible derivation of TMDA from lanthionine is discussed. This represents the second S-containing cyclic amino acid so far discovered in a mammalian brain whose physiological significance has not yet been explored.     

Chlorogenic acid in a Nicotiana plumbaginifolia cell suspension

A phenylpropanoid compound has been characterized in a Nicotiana plumbaginifolia cell suspension. This compound has been isolated and purified by semi-preparative reverse phase-high performance liquid chromatography. Its structure has been identified by NMR spectroscopy as 5-O-caffeoylquinic acid, which is chlorogenic acid (CA). The influence of culture conditions on the accumulation of this metabolite by N. plumbaginifolia cell suspensions has been studied. Darkness strongly inhibits the CA accumulation. Moreover, it has been shown that feeding experiments with caffeic acid had a deleterious effect upon the CA content. This one was not influenced by a supplementation with quinic acid.     

Analysis of 3-indolylacetic acid and abscisic acid by high-performance liquid chromatography and gas-liquid chromatography

A method of analysis of 3-indolylacetic acid (IAA) and abscisic acid (ABA), allowing the simultaneous extraction of both regulators from plant material, has been developed. The method involves extraction with methanol, isolation of the acid fraction, diazomethane methylation, separation of the hormones through reverse-phase preparative high-performance liquid chromatography, and quantification of both compounds by gas-liquid chromatography. The recovery percentage at each step was monitored with radioactive compounds added at the beginning of the process. The final recovery was 70% for IAA and 96% for ABA. The method was applied to the analysis of the IAA and ABA content of stems of hazel (Corylus avellana L.).     

Structural parameters and natural occurrence of 2-deoxy-2,3-didehydro-N-glycoloylneuraminic acid

2-Deoxy-2,3-didehydro-N-glycoloylneuraminic acid has been found to occur in porcine, bovine and equine submandibular glands as well as in the urine of pig, horse and rat. This novel, unsaturated sialic acid was isolated by gel filtration and ion-exchange chromatography. Final purification was achieved by column chromatography or by preparative thin-layer chromatography on cellulose. The structural analysis was performed by combined capillary gas-liquid chromatography/mass spectrometry. The various data were compared with those from synthetic 2-deoxy-2,3-didehydro-N-glycoloylneuraminic acid. Besides of the unsaturated N-glycoloylated sialic acid, also the corresponding N-acetylated derivative was present in the materials analyzed. The inhibitory effect of 2-deoxy-2,3-didehydro-N-glycoloylneuraminic acid on Vibrio cholerae sialidase using N-acetylneuraminyl-(alpha 2----3)-lactose as substrate is slightly higher (50% inhibition at 10 microM) when compared with 2-deoxy-2,3-didehydro-N-acetylneuraminic acid (50% inhibition at 15 microM).     

Biosynthesis of folic acid

Summary It has been attempted to isolate and characterize the folate precursors in the culture filtrates of two folate-requiring organisms, Streptococcus faecalis R and Lactobacillus casei. On the basis of paper chromatography, bioautography, ultra violet absorption spectra, chemical reactions, and differential microbiological responses it has been concluded that L. casei cultures contain a compound similar to pteroic acid which can be utilized by S. faecalis R. The S. faecalis R cultures on the other hand appear to accumulate a pteridine derivative active for Crithidia fasciculata It has been confirmed that this pteridine is not derived from the folic acid usually added to the growth medium.Abbreviations PGA Pterolyglutamic acid - PABA p-aminobenzoic acid     

Liquid-gel partition chromatography of vitamin A compounds; formation of retinoic acid from retinyl acetate in vivo

A clear separation of retinol, retinal, and retinoic acid has been achieved by liquid-gel partition chromatography on Sephadex LH-20 with solvent mixtures of chloroform, Skellysolve B, and methanol. A mixture of retinyl esters, retinol, retinal, and retinoic acid has been resolved on hydroxyalkoxypropyl Sephadex using Skellysolve B and acetone. There is no decomposition of any of the vitamin A compounds during chromatography, and recovery is complete. The combination of mildness and potential for resolution makes liquid-gel partition chromatography a superior tool for the separation of vitamin A compounds. This method has been applied to the study of vitamin A metabolism at physiological levels in the vitamin A-deficient rat. Retinyl palmitate, an ester of retinoic acid, retinal, retinol, retinoic acid, and a polar metabolite have been demonstrated in various tissues of the rat 12 hr after a dose of 2 micro g of [11-(14)C]retinyl acetate.     

Postmortem changes in uric acid and ascorbic acid in human cerebral cortex tissues excised after cardiac death

There has been no report on the determination of uric acid (UA) in human brain and heart tissues. UA and ascorbic acid (AA) in human cerebral cortex and heart tissues excised after cardiac death have been studied by reversed-phase high-performance liquid chromatography (HPLC) with electrochemical detection (ECD). It has been found that the levels of AA and UA in the human cerebral cortex tissues tend to decrease and increase, respectively, after cardiac death as a function of time between death and forensic operation. In addition, it has been found that there is no special relationship between UA levels in human heart tissues and time after cardiac death, also that the UA levels in the heart are high as compared with those in human cerebral cortex tissues. We have emphasized that the HPLC-ECD method is useful in determining UA and AA in mammalian tissues by one-time chromatography to gain a better understanding of the relationship between disease and serum urate level.     

创新霉素产生菌吲哚丙酮酸甲基转移酶的研究

在创新霉素产生菌济南游动披线菌的无细胞提取物中检测到吲哚丙酮酸甲基转移酶活性,并进行了分离提取。该酶能利用S-腺苷-L-甲硫氨酸对吲哚丙酮酸进行甲基化,它可能作用于创新霉素中间体的甲基化。经过硫酸铵分部盐析和DEA F一纤维素柱层析,得到了纯化60倍的甲基转移酶,比活0．66mu／ms。酶的最适底物是吲哚丙酮酸,最适pH7．5,对于底物s一腺苷一L一甲硫氨酸和吲哚丙酮酸的米氏常数(KⅢ)分别是4×10-,mol／L和1．8×10-7moI／L。用Sephadcx G-150凝胶过滤测得分子量是55000士5000道尔顿。     

Estimation of dehydroascorbic acid in blood of diabetic patients.

A modified method has been described for the estimation of ascorbic acid (AA) and dehydroascorbic acid (DHA) in blood and plasma. DHA is practically absent in the blood of normal human beings. On the other hand, diabetic patients have a persistently high blood DHA level. The DHA from diabetic blood has been isolated as the 2,4-dinitrophenylhydrazone derivative and identified by thin-layer chromatography and spectrophotometry.     

Enzymatic reduction of dehydrocholic acid to 12-ketochenodeoxycholic acid with NADH regeneration

12-Ketochenodeoxycholic acid, an essential intermediate in the synthesis of chenodeoxycholic acid, has been enzymatically prepared from dehydrocholic acid. The specific reduction of dehydrocholic with NADH was catalysed by 3α-hydroxysteroid dehydrogenase (3α-hydroxysteroid: NAD(P)⁺ oxidoreductase, EC 1.1.1.50) and 7α-hydroxysteroid dehydrogenase (7α-hydroxysteroid:NAD⁺ 7-oxidoreductase, EC 1.1.1.159). Cofactor regeneration was obtained through the formate dehydrogenase (formate:NAD⁺ oxidoreductase, EC 1.2.1.2) catalysed oxidation of formate. Complete transformation of dehydrocholic acid to the 12-keto derivative was achieved with a coenzyme turnover number up to 1200. No steroid by-products were detected by high performance liquid chromatography and thin layer chromatography. The process yielded 9 g product l^?1 in 66–84 h. The high purity of the enzymatically prepared 12-ketochenodeoxycholic acid should drastically reduce the formation of the toxic by-product lithocholic acid, which occurs in the synthesis of chenodeoxycholic acid when using chemical methods alone.     

Abscisic acid in pea shoots

Summary The (+)-abscisic acid content of pea shoots has been determined using gas-liquid chromatography. In both tall and dwarf cultivars no significant difference was observed between plants grown in the dark or under red light. Nor was the difference between the tall and dwarf cultivars themselves significant.     

High-pressure liquid chromatographic analysis of amino acid phenylthiohydantoins: Comparison with other techniques

High-pressure liquid chromatography, utilizing reverse phase μ Bondapak C₁₈ columns and elution with increasing acetonitrile concentrations, has been used to resolve amino acid phenylthiohydantoins obtained from the automated Edman degradation of proteins. Assignment of identity to residues which are difficult to distinguish or identify conclusively by other conventional techniques is easily achieved by high-pressure liquid chromatographic techniques. The use of high-pressure liquid chromatography, in parallel with gas-liquid and polyamide thin-layer chromatography, allows unequivocal assignments of identity to amino acid phenylthiohydantoins obtained in protein sequencing. Single protein sequence determinations can be extended by 20 to 100% by the use of high-pressure liquid chromatography with rapid, accurate, and quantitative identifications of amino acid phenylthiohydantoins.     

Identification of abscisic acid from shoots of Salix pentandra

Abscisic acid isolated from vegetative shoots of Salix pentandra L. has been identified by high performance liquid chromatography and combined gas chromatography-mass spectrometry.     

Dry-column chromatographic isolation of fatty acid esters of phorbol from croton oil

The family of mixed fatty acid esters of 4,9,12,13,20-pentahydroxy-tiglia-1,6-dien-3-one (phorbol) has been isolated from croton oil by column chromatography on silica gel and preparative dry-column chromatography on silica gel HF/254 packed in a cellophane membrane. The tumorpromoting agents are obtained rapidly and with minimum difficulty.     

gamma-Carboxyglutamic acid in invertebrates: its identification in hermatypic corals

The presence of a γ-carboxyglutamate-containing protein in hermatypic corals has been established. γ-Carboxyglutamate has been isolated from the alkaline hydrolysate of protein extracted from the coral Lobophyllia corymbosa, by chromatography of the hydrolysate on Dowex AG 1-X8 (formate form), followed by chromatography on an amino acid analyzer column. This procedure achieves complete separation of γ-carboxyglutamate from an acid-stable compound which is also present in the alkaline hydrolysate of coral protein, and which has proved difficult to separate from γ-carboxyglutamate by other methods. The identity of the γ-carboxyglutamate thus isolated has been established unequivocally by determining the yield of glutamic acid after acid treatment (2 M HCl, 6 h, 110°C). The color factor for the γ-carboxyglutamate isolated from Lobophyllia is 0.45₄ times the value for glutamic acid, in good agreement with the value determined for authentic γ-carboxyglutamic acid (0.45₈) under exactly the same conditions. Virtually identical results have been obtained for the coral, Acropora cuneata. These experiments provide the first secure evidence for the presence of γ-carboxyglutamate in an invertebrate species, and they clearly have important implications for our understanding of invertebrate mineralization.