采用高效液相色谱技术分析生物体内维生素B6

维生素B6(VB6)是一类2-甲基-3-羟基吡啶类化合物的总称，基本类型有吡哆醇(PN)、吡哆醛(PL)和吡哆胺(PM)，磷酸酯型有磷酸吡哆醇(PNP)、磷酸吡哆醛(PLP)和磷酸吡哆胺(PMP)，其中磷酸吡哆醛和磷酸吡哆胺为活性形式，是多种酶的辅酶，哺乳动物尿中VB6的代谢产物主要是不具有生理活性的吡哆酸(PIC)，在植物体内还发现有数种吡哆醇的糖衍生物和氨基酸衍生物。     

Zur Analytik der Pyridoxine

Zusammenfassung Es werden 4 spezifische Nachweis- und Bestimmungsreaktionen für Pyridoxal und Pyridoxamin mitgeteilt. In Kombination mit dem Nachweis mit Gibbss'chem Reagens können nunmehr Pyridoxol, Pyridoxal und Pyridoxamin in Gemischen nebeneinander bestimmt werden. Die Überführbarkeit der 3 Komponenten ineinander wird aufgezeigt.

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On the analysis of pyridoxins

4 selective tests for pyridoxal and pyridoxamine are described. In combination with the test by means of 2,6-dichloroquinone-4-chloroimine it is possible to determine pyridoxol, pyridoxamine and pyridoxal in presence of each other in mixtures. It is shown that the three components are able to be transformed into one another.

     

Kinetic and mechanism of reactions of L-α-Glutamic acid and L-Glutamine with pyridoxal

The kinetics and mechanisms of condensation of pyridoxal with L-α-glutamic acid and L-glutamine were studied by UV spectroscopy and polarimetry. L-α-Glutamic acid reacts with pyridoxal to form a Schiff base whose subsequent hydrolysis gives rise to pyridoxamine and α-ketoglutaric acid. The reaction of Lglutamine with pyridoxal involves the Γ-NH₂ group and affords a Schiff base whose subsequent hydrolysis gives rise to pyridoxamine and L-α-glutamic acid.     

Quantitative profiling of biomarkers related to B‐vitamin status,tryptophan metabolism and inflammation in human plasma by liquid chromatography/tandem mass spectrometry

Vitamins B₂ and B₆ serve as cofactors in enzymatic reactions involved in tryptophan and homocysteine metabolism. Plasma concentrations of these vitamins and amino acids are related to smoking and inflammation, and correlate with other markers of immune activation. Large‐scale studies of these relations have been hampered by lack of suitable analytical methods. The assay described includes riboflavin, five vitamin B₆ forms (pyridoxal 5′‐phosphate, pyridoxal, 4‐pyridoxic acid, pyridoxine and pyridoxamine), tryptophan and six tryptophan metabolites (kynurenine, kynurenic acid, anthranilic acid, 3‐hydroxykynurenine, xanthurenic acid and 3‐hydroxyanthranilic acid), cystathionine, neopterin and cotinine. Trichloroacetic acid containing 13 isotope‐labelled internal standards was added to 60 µL of plasma, the mixture was centrifuged, and the resulting supernatant used for analysis. The analytes were separated within 5 min on a stable‐bond C8 column by a gradient‐type mobile phase containing acetonitrile, heptafluorobutyric acid and high concentration (650 mmol/L) of acetic acid, and detected using electrospray ionization tandem mass spectrometry (ESI‐MS/MS). The mobile phase ensured sufficient separation and high ionization efficiency of all analytes. Recoveries were 75–123% and within‐day and between‐day coefficients of variance (CVs) were 2.5–9.5% and 5.4–16.9%, respectively. Limits of detection ranged from 0.05 to 7 nmol/L. The method enables quantification of endogenous plasma concentrations of 16 analytes related to B‐vitamin status and inflammation, and may prove useful in large‐scale epidemiological studies. Copyright © 2009 John Wiley & Sons, Ltd.     

The composition of the complex formed in concentrated sulfuric acid between boric acid and 1,1'-dianthrimide

The complex formed in concentrated sulfuric acid between boric acid and 1,1'-dianthrimide was isolated in the solid state. It was found to consist of one six-membered boron-containing ring of the type reported by Gillespie and rohinson and two 1,1'-dianthrimide molecules, the molecular formula being C₅₆H₂₈N₂O₁₃B₂S. It is suggested that other reagents reacting with boric acid in concentrated sulfuric acid form complexes of the same type.     

Simultaneous determination of pyridoxal and pyridoxamine by different spectrofluorimetric techniques

Binary mixtures of pyridoxal and pyridoxamine can be resolved by using zero-crossing first derivative spectrofluorimetry, first devivative constant wavelength synchronous luminescence spectrometry and first derivative constant energy synchronous luminescence spectrometry. These methods do not require any previous separation steps. The lowest quantization limit is obtained with first derivative constant energy synchronous fluorescence (13.0 and 9.0 mug/1. for pyridoxal and pyridoxamine, respectively). The measurements were performed in aqueous medium at pH 7.0 provided by adding 0.05M phosphate buffer solution. In order to demonstrate the validity of these methods a complete and exhaustive statistical analysis of the experimental data was performed. Pyridoxal and pyridoxamine were determined by these methods in synthetic and real mixtures with good results.     

An N⋯H⋯N low-barrier hydrogen bond preorganizes the catalytic site of aspartate aminotransferase to facilitate the second half-reaction

Pyridoxal 5′-phosphate (PLP)-dependent enzymes have been extensively studied for their ability to fine-tune PLP cofactor electronics to promote a wide array of chemistries. Neutron crystallography offers a straightforward approach to studying the electronic states of PLP and the electrostatics of enzyme active sites, responsible for the reaction specificities, by enabling direct visualization of hydrogen atom positions. Here we report a room-temperature joint X-ray/neutron structure of aspartate aminotransferase (AAT) with pyridoxamine 5′-phosphate (PMP), the cofactor product of the first half reaction catalyzed by the enzyme. Between PMP N_SB and catalytic Lys258 Nζ amino groups an equally shared deuterium is observed in an apparent low-barrier hydrogen bond (LBHB). Density functional theory calculations were performed to provide further evidence of this LBHB interaction. The structural arrangement and the juxtaposition of PMP and Lys258, facilitated by the LBHB, suggests active site preorganization for the incoming ketoacid substrate that initiates the second half-reaction.

The neutron structure of pyridoxal 5′-phosphate-dependent enzyme aspartate aminotransferase with pyridoxamine 5′-phosphate (PMP) reveals a low-barrier hydrogen bond between the amino groups of PMP and catalytic Lys258, preorganizing the active site for catalysis     

Pyridoxamine,a scavenger agent of carbohydrates

Pyridoxamine has been found to inhibit protein glycation and to avoid the formation of advanced glycation end‐products (AGEs). One of the mechanisms by which pyridoxamine can inhibit glycation involves the scavenger of carbonyl groups with glycation capacity. In this work, we conducted a kinetic study of the reactions of pyridoxamine with various carbohydrates under physiological pH and temperature. The reactions involving hexoses were found to give a tricyclic compound ( 5 ) in addition to pyridoxal and pyridoxine. Such a tricyclic compound inhibits the Amadori rearrangement and the formation of other carbonyl compounds with glycating properties. The reactions involving pentoses gave compound 7 and pyridoxal—by transamination of the Schiff base. The transamination reaction enhances the inhibitory action of pyridoxamine. The formation rate constants for the Schiff base, k₃, were found to be similar to those for the reactions of D ‐glucose with amino acids, which suggests competition between pyridoxamine and terminal amino residues in proteins for glycating sites in sugars. These constants are dependent on the electrophilic character of the carbonyl carbon in the carbohydrate. © 2007 Wiley Periodicals, Inc. 39: 154–167, 2007     

Kinetics and mechanism of the condensation of pyridoxal hydrochloride with L-tryptophan and D-tryptophan,and the chemical transformation of their products

The kinetics and mechanism of interaction between pyridoxal and L-tryptophan, D-tryptophan, and their derivatives are studied. It is found that condensation reactions proceed via three kinetically distinguishable stages: (1) the rapid intraplanar addition of the NH₂ groups of the amino acids to pyridoxal with the formation of amino alcohols; (2) the rotational isomerism of amino alcohol fragments with their subsequent dehydration and the formation of a Schiff base with a specific configuration; (3) the abstraction of α-hydrogen in the product of condensation of pyridoxal with L-tryptophan, or the abstraction of СО₂ in the product of condensation of pyridoxal with D-tryptophan with the formation of quinoid structures, hydrolysis of which results in the preparation of pyridoxamine and keto acid or pyridoxal and tryptamine, respectively. Schiff bases resistant to further chemical transformations are formed in the reaction with tryptophan methyl ester.     

A spectroscopic investigation of the behavior of anthraquinone in sulfuric acid and oleum

Dissolution of anthraquinone in concentrated sulfuric acid and in oleum is accompanied by a coloration of the solution. It has been observed [1] that in 92–96% sulfuric acid anthraquinone gives a yellow solution, while in 20% oleum the color is red. The assumption has been made [2] that in the first case one of the carbonyl groups of the anthraquinone reacts with the acid, while in the second case both undergo reaction.     

Phosphorylation of pyridoxal azomethines. Synthesis of phosphorus containing azomethines and furopyridines

New O-phosphorylated pyridoxal derivatives have been synthesized through the reaction of azomethines with Р^V acid chlorides. 2-Chloro-2-thioxo-5,5-dimethyl-1,3,2-dioxaphosphinanes and diethylchlorothiophosphate have been employed as phosphorylating agents. Regardless of the nature of the phosphorylating agent, the reaction is regioselective at phenolic hydroxyl group. The structure of final products is determined by the nature of the substituent at the nitrogen atom. If R is alkyl or cycloalkyl group, the products of the reaction represent phosphorylated pyridoxal imines, whereas phosphorylated furopyridines are formed in the case R is aryl substituent.     

Quantities of B6 vitamers in human milk by high-performance liquid chromatography. Influence of maternal vitamin B6 status

A rapid, sensitive procedure is described for the analysis of the B6 vitamers pyridoxal, pyridoxamine, and pyridoxine in human milk from women taking and not taking supplements containing the vitamin using high-performance liquid chromatography with fluorometric detection. Vitamer values represent the sum of their phosphorylated and unphosphorylated forms. Minimum detectable quantities were 1-3 ng. Excellent recoveries of these vitamers in milk were obtained. Similar B6 vitamer concentrations of milk were obtained using the developed high-performance liquid chromatographic and the accepted microbiological techniques. Pyridoxal, actually consisting of pyridoxal plus pyridoxal phosphate, was the predominant B6 vitamer in human milk. The concentration of B6 vitamers in milk was reflective of the maternal vitamin B6 status.     

Complex formation in concentrated sulfuric acid between selenium(iv) and 1,1'-dianthrimide

The complex formation in concentrated sulfuric acid between selenium(IV) and 1,1'-dianthrimide (Di) was studied by spectrophotometry, infrared spectroscopy and chemical analysis. The system was found to contain two species, a Se₂Di complex and aselenium-1,2,7,8-diphthaloylcarbazole complex. The primary complex reaction was assumed to be the formation of the Se₂Di compound; unter certain experimental conditions one selenium atom is split off, resulting in the irreversible formation of a carbazole bond. The selenium-1,2,7,8-diphthaloylcarbazole was prepared in the solid state.     

N-dealkylation of 2-(tert-butyl)-3-hydroxy-3-(4-chlorophenyl)isoindolinone and n-(tert-alkyl)-2-aroylbenzamides in concentrated sulfuric acid

It has been established that 2-(tert-butyl)-3-hydroxy-3-(4-chlorophenyl)isoindolinone and N-(tert-Alkyl)-2-aroylbenzamides are dealkylated in concentrated sulfuric acid to give 3-hydroxy-3-arylisoindolinones. The reaction is realizable. only when there is a tert-alkyl group attached to the nitrogen atom. The reaction mechanism is discussed on the basis of data on the change with time in the electronic spectra of the investigated compounds in concentrated sulfuric acid and a comparison with the spectra of model structures.Translated from Khimiya Geterotsiklicheskikh Soedinenii, No. 6, pp. 763–765, June, 1977.     

The novel usage of thiourea nitrate in aryl nitration简

Thiourea nitrate （TN） was easily prepared from thiourea and nitric acid to explore its use as a new nitration reagent, Nitrations of various aromatic compounds utilizing TN in concentrated sulfuric acid were studied, TN could convert aromatic compounds to the corresponding nitrated derivatives with various abnormal yields under mild conditions. The results suggested that the reaction mechanism might be different from those of traditional nitration reagents.     

Complex formation in concentrated sulfuric acid between boric acid and quinalizarin or alizarin

The complex formation in concentrated sulfuric acid between boric acid and quinalizarin (1,2,5,8-tetrahydroxyanthraquinone) or alizarin (1,2-dihydroxyanthraquinone) was studied by spectrophotometry. Both systems contained only one species, viz. a complex between one boric acid and one hydroxyanthraquinone molecules. The reactions of boric acid with hydroxyanthraquinones are discussed and compared with the reaction of boric acid with 1,1'-dianthrimide.     

Production of formic and acetic acids from phenol by hydrothermal oxidation

Hydrothermal technology is a core environmental-protection technique which can be used for waste water treatment and biomass conversion. In this paper a novel idea, alkaline hydrothermal oxidation, is proposed for producing formic and acetic acids from wastewater containing phenolic compounds. The effects of the most important conditions??reaction temperature, reaction time, oxygen supply, and type of alkaline catalyst??on yields of formic and acetic acids were investigated. The results indicated that the optimum conditions for production of formic and acetic acids were: reaction temperature 300???C, reaction time 90?s, H₂O₂ equivalent to 60% oxygen, and NaOH concentration 1.5?mmol. Under the optimum conditions the yields of formic and acetic acids reached 4.8 and 23.5%, respectively. In addition, the effect of different alkalis on yields of formic and acetic acids was also investigated. The results showed that compared with use of NaOH addition of KOH had a more pronounced effect on improving the yield of acetic acid. This research indicated that high-value-added formic and acetic acids can be recovered as resources by hydrothermal oxidation of phenolic wastewater, and thus hydrothermal oxidation has high potential for converting phenolic compounds in wastewater into value-added products.     

Electrophilic substitution in the 1-phenyl-2-acylpyrazolidine series

Bromination, nitration, and sulfonation reactions in the acylphenylpyrazolidine series were investigated. 1-(p-Bromophenyl)-2-acylpyrazolidines are formed in good yields in the bromination of these compounds over a wide range of temperatures in various solvents. Removal of the acyl group takes place simultaneously with sulfonation in the para position of the phenyl ring in the sulfonation of phenylacylpyrazolidines with concentrated sulfuric acid at room temperature; the p-sulfophenylpyrazolidines formed in this case exist in the form of betain structures. The nitration of acylphenylpyrazolidines with concentrated nitric acid (sp. gr. 1.52) leads to 1-(2,4-dinitrophenyl)-2-acylpyrazolidines. However, the nitration of these compounds with dilute nitric acid (sp. gr. 1.35) is accompanied by pronounced resinification; both 2,4-dinitrophenyl and p-nitrophenyl-2-acylpyrazolidines, as well as dimers of the latter, were detected among the reaction products.Translated from Khimiya Geterotsiklicheskikh Soedinenii, No. 10, pp. 1377–1380, October, 1978.     

Reactions of methylcyclohexanones with concentrated sulfuric acid

Conclusions In reaction with concentrated sulfuric acid 2-, 3-, and 4-methylcyclohexanones undergo autocondensation and simultaneous intramolecular cyclization with formation of furan compounds.Translated from Izvestiya Akademii Nauk SSSR, Seriya Khimicheskaya, No. 11, pp. 1958–1963, November, 1966.