Pentoses determination

Schabort and Potgieter (1971) showed that each of the J -cyclopiazonate oxidocyclases contain one covalently linked molecule of flavin per molecule of enzyme. The purified enzymes had a yellow color, and their ultraviolet and visible spectra showed maxima at 276, 366, and 450 nm. The addition of sodium dithionite or of jSCA caused the disappearance of the 450 nm peak because of the reduction of the flavin moiety. The flavin residues released from the protein molecules by proteolytic digestion with pronase showed absorption peaks at 262, 370, and 450 nm. The released flavin residues, separated by paper chromatography, were found to contain a covalently bound amino acid or small peptide. It was concluded that in the native enzyme, the flavin was bound by covalent linkage to the protein and that, during digestion with pronase, proteolysis stops at points determined by the specificity of the pronase or by structural or steric hindrance by the flavin. Pentose determination on the flavin residues showed that the flavins were dinucleosides. 

In as far as other analytical methods are concerned, many specific reactions have been elaborated for the quantitative determination of 2-deoxy aldoses. 2-Deoxy-D-ribose (2-deoxy-D-erythro-pentose), a compound which was recognized early as playing an important role in biological systems, has been of particular interest. Overend and Stacey (43) have given a critical review of the methods available until 1952 for the estimation of 2-deoxy pentoses. A recent summary of specific methods for the identification and quantitative estimation of the different classes of deoxy sugars has been prepared by Dische (13). 

D-Xylulose 5-phosphate (ii-threo-2-pentulose 5-phosphate, XP) stands as an important metabolite of the pentose phosphate pathway, which plays a key fimction in the cell and provides intermediates for biosynthetic pathways. The starting compound of the pathway is glucose 6-phosphate, but XP can also be formed by direct phosphorylation of D-xylulose with li-xylulokinase. Tritsch et al. [114] developed a radiometric test system for the measurement of D-xylulose kinase (XK) activity in crude cell extracts. Aliquots were spotted onto silica plates and developed in n-propyl alcohol-ethyl acetate-water (6 1 3 (v/v) to separate o-xylose/o-xylulose from XP. Silica was scraped off and determined by liquid scintillation. The conversion rate of [ " C]o-xylose into [ " C]o-xylulose 5-phosphate was calculated. Some of the works devoted to the separation of components necessary while analyzing enzyme activity are presented in Table 9.8. 

In the determination of carbohydrates, sensitivity can often be increased by using fluorescence rather than absorbance for the final determination. With compounds that are not normally fluorescent, it becomes necessary to find fluorescent derivatives. Hirayama [160] concentrated the carbohydrates in coastal water samples, using electrodialysis and evaporation, and made fluorescent derivatives using anthrone and 5-hydroxyl-1-tetralone, determining pentoses separately from hexoses in the process. While this method does seem to have the extra sensitivity expected from fluorescent methods, the extra manipulations render it unsatisfactory for routine use. 

Hydrolysis products of neomycin may be an amino-sugar, a pentose or furfural depending on the reaction conditions chosen. Each of these entities has been utilised for indirect spectrophotometric determination of neomycin. 

The in vitro bioassays allowed to determine the inhibition constant of D-fructose transport by the CHO cells. This measure is carried out by competition with radioactive D-fructose. The study put in evidence that pentose-OZT derivatives are not recognized by the protein transporter. Only the ketohexose-OZT derivatives expressed some inhibition of GLUT5. These inhibition constants showed to be much effective with L-Sor derivatives than with D-Fru derivatives and even better than D-fructose itself (Kt = 15.5 mM) (Table 2). 

Figure 4.17 The trioses D-glyceraldehyde (aldose) and dihydroxyacetone (ketose), the pentose D-ribose, the hexoses D-galactose and D-glucose (aldoses) and the ketohexose D-fructose in their open chain forms. The configuration of the asymmetrical hydroxyl group on the carbon, the furthest away from the aldehyde or ketone group, determines the assignment of D- or L-configuration.

B. Luo, K. Groenke, R. Takors, C. Wandrey, and M. Oldiges, Simultaneous determination of multiple intracellular metabolites in glycolysis, pentose phosphate pathway and tricarboxylic acid cycle by liquid chromatography mass spectrometry. J. Chromatogr. A 1147, 153 164 (2007). 

Glyceraldehyde (2,3-dihydroxypropanal), acetol, and dihydroxyace-tone form 1-5% of biacetyl and a number of other products, including pyrocatechol and 33, after exposure to aqueous alkali at 300°. Such trioses as glyceraldehyde and dihydroxyacetone have been shown to form various hexoses by aldol reaction. Aldolization, followed by retro-aldoliza-tion, is undoubtedly a major consideration when three-, four-, and five-carbon sugars are subjected to elevated temperatures. Differences in thermolysis products, partially quantitative, are noticeable at 100°, but, at temperatures near 300°, it is quite difficult, if not impossible, to determine if the starting material was a triose, a tetrose, or a pentose. 

Sugars can be metabolized by either glycolysis or by the pentose phosphate pathway. The activities in these pathways can be determined by measuring the enzyme activities of selected dehydrogenases in each path. In soybean, ozone can alter the... 

As pentoses are readily accessible from wheat straw and bran [26, 27], the telomerization of 1 with a bran syrup having the composition given in Table 15 led to a crude mixture containing 1% bran symp, 67% monooctadienylethers (18, 23), 31% dioctadienylethers (24—26), and 1% trioctadienylethers (Fig. 21). The physical evaluation of this mixture is given in Fig. 22 and revealed satisfactory surface-active behavior of this crude mixture although no sharp value of CMC could be determined, as can happen with complex mixtures. Continuous decrease of... 

Fig. 2. Experimental design of a C tracer experiment for the determination of the flux partitioning ratio between pentose phosphate pathway and glycolysis ( ppp) C label distribution from l- C glucose through the network with C atoms (black) and C atoms (white)...

Minimum amounts of 2-3 /ig of sialic acids can be determined in, for example, the orcinol-Fe3+ assay, if the volumes of the Bial reagents23 are one-fifth of those originally described.107 By using this method, sialic acids can be accurately determined only if they have been prepurified, as free or glycosidically bound pentoses, hexoses, or alduronic acids interfere with the reaction by giving green chromo-phores.107 In our experience, an approximate determination of the sialic acid in a complex carbohydrate is only possible if its sialic acid content is >5%. 

Entry of glucose 6-phosphate either into glycolysis or into the pentose phosphate pathway is largely determined by the relative concentrations of NADP+ and NADPH. 

The position of the pyranose-furanose equilibria in solution has been determined for the four 5-acetamido-5-deoxypentoses the proportion of the pyranose form (which contains the nitrogen atom in the ring) is — 65% for the xylo, 50% for the lyxo, 25% for the arabino, and 10% for the ribo isomer,124 and this is the order found for the parent pentoses. The corresponding 5-(benzyloxycarbonyl)amino-5-deoxypentoses, however, exist in solution almost exclusively in the pyranose form,130 reflecting the diminished extent of deactivation of the amide nitrogen atom but a solution of 5-(benzyloxycarbonyl)amino-5,6-dideoxy-3-0-mesyl-L-idose was found to contain 20% of the furanose forms, because the steric effect of the N-acyl group forces it into the particularly unfavorable 4Cx(l) conformation (26) of the y3-pyranose form.132... 

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