Low-pH method for the enzymatic assay of D-glucaric acid in urine.
The enzymatic methods for measuring D-glucaric acid in urine are based on the conversion of D-glucaric acid into its 1,4-lactone and measurement of inhibition of 1,4-lactone against beta-glucuronidase at pH 5.0. All the enzymatic methods described suffer from the disadvantage of a procedure that is complicated and inherently inaccurate, because the nature of glucaric acid/1,4-lactone equilibrium has not been properly considered in the development of such methods. After elucidating the factors influencing glucaric acid/1,4 lactone equilibrium in more detail, a low-pH enzymatic method has been developed in which the 1,4-lactone is formed in the urine sample by acid boiling at pH 3.8 and assayed at the same pH using beta-glucuronidase from Limpets. This procedure allows the acid/lactone equilibrium to remain stable during both the lactonization step and the enzymatic assay. The coefficient of variation for the proposed method (within-run and between-day precision) was from 4.2 to 8.7. The analytical recovery varied from 92-108%.
D005937:Glucaric Acid / Q000652:urine*; D005966:Glucuronidase / Q000037:antagonists & inhibitors*; D006358:Hot Temperature; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007783:Lactones / Q000494:pharmacology / Q000652:urine; D008722:Methods; D008974:Mollusca / Q000201:enzymology; D013400:Sugar Acids / Q000652:urine*
D007783:Lactones; D013400:Sugar Acids; D005966:Glucuronidase; D005937:Glucaric Acid