Formylpyruvic acid

Exposure to periodate yields 3-formylpyruvic acid, which reacts with thiobarbituric acid, giving rise to red condensation products having an absorption maximum at 545-550 nm. Certain workers have used the thiobarbituric acid test without previous oxidation of the unsaturated oligo-D-galactosiduronates with periodic acid.3-29,239,260 Even here, the products of reaction with thiobarbituric acid have an absorption maximum at 550 nm. 

Hyaluronate lyase purified from Streptomyces hyalurolyticus has been used to provide a specific assay for hyaluronic acid, its action resulting in the generation of 8-D-gluco-4-ene-pyranosyluronic acid groups. The assay is based on the long established periodate oxidation of such groups to inter alia j3-formylpyruvic acid which specifically gives a chromophore with thiobarbituric acid. 

The other widely employed colorimetric assay is the thiobarbituric acid procedure which is useful for free but not bound sialic acid. Proposed independently by Warren (1959) and Aminoff (1959), the method is based on periodate oxidation of sialic acid to )8-formylpyruvic acid (Figure 11) which then reacts with two moles of 2-thiobarbituric acid to form a chromophore (XLIX) with absorption maximum at 549 nm. The identity of this chromophore was established through synthesis... 

In the Warren (1959) procedure NANA and NGNA had molar extinction coefficients of 57,000 and 46,000, respectively, while the Aminoff (1959) method showed a somewhat greater spread between these two. Since the same chromophore is produced for both sialic acids, this difference probably reflects unequal yields of /S-formylpyruvic acid. The high extinction coefficients make this one of the more sensitive methods, as little as 3 tg being detectable (Warren, 1963). O-Acetylated sialic acids respond differently depending on the position of substitution color yield is reduced only slightly with an acetyl on the C4-hydroxyl, more so with acetyl at Cg or C9, and no color forms when the O-acetyl is at C7 (Aminoff, 1%1 Warren, 1964 Schauer and Faillard, 1968). N-Glycolyl-8-O-methylneuraminic acid produced only 13% of the... 

The TBA assay for 3-deoxyglyc-2-ulosonic acids, used by Weissbach and Hurwitz11 to determine 3-deoxy-D-arabino-2-heptulosonic acid, was subsequently applied by Warren13 and by Aminoff14 to the analysis ofN-acetylneuraminic acid (NeuAc, 5 see Fig. 4). The formation of the 3-formylpyruvate-derived chromophore 4 in the periodate... 

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