Modified method for analysis of glyoxylate derivatives in biological materials

✍ Scribed by J. Nirmala; K.Sivarama Sastry

• Publisher: Elsevier Science
• Year: 1972
• Tongue: English
• Weight: 479 KB
• Volume: 47
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(72)90295-3

✦ Synopsis

A highly sensitive method for assaying glyoxylate is based on the Rimini-Schryver react,ion. This involves the formation of glyoxylate phenylhydrazone and its subsequent oxidation, in strongly acid medium, to the intensely red 1,5-diphenylformazan, absorbing at, 520 nm. This reaction was first applied for glyoxylate determination by Fosse and Hieulle (1) and by Fosse and Bossuyt (2). Many variations of the basic procedure have been reported since then, but the recently described method of Vogels and van der Drift (3) is among the most suited for t.he differential analysis of allantoin, allantoate, ureidoglycolate, and glyoxylate; these authors have developed conditions for the selective hydrolysis of such glyoxylate derivat.ives to glyoxylate, which is then assayed.

During our studies on allantoin metabolism, we found that such analysis is not possible in protein-rich tissue homogenates since pronounced turbidities were obtained during analysis of these extracts due to the presence of concentrated acid in the terminal stages of the assay. Routine deproteinization procedures, as treatment with trichloroacetic acid, were inadvisable in view of the nature of the analytical procedure and also the lability of allantoate and ureidoglycolate; also, for instance, trichloroacetate itself interferes in the assay (see below). We present,, in this paper, a modified procedure hy which analysis of glyoxylate (and glyoxylate derivatives) can be performed dircrtly without deproteinization. Here, the coupling of glyoxylate with phenylhydrazine and its oxidation to the formazan derivative are conducted in the presence of protein and the colored product i, ': selectively extracted into a mixture of chloroform/isoamyl alcohol and quantitated in this ljhaxe. Some factors that, influence this procedure are also discussed.