Urinary glycolate measured by use of (S)-2-hydroxy-acid oxidase

Abstract

Glycolate can be determined in urine by using (S)-2-hydroxy-acid oxidase (EC 1.1.3.15; formerly called "glycolate oxidase"), either immobilized in a continuous-flow system or in a semiautomated procedure for the centrifugal analyzer. In the presence of peroxidase (EC 1.11.1.7), the hydrogen peroxide formed from glycolate is detected by use of a peroxide indicator reaction. Before the analysis, urine must be treated with charcoal to remove reducing substances such as ascorbic acid, which interfere with the assay by decreasing the color of the indicator reaction. Lactate also interferes with the determination of glycolate because it also is a substrate for this oxidase; thus a correction has to be made for the lactate content of urine. The system with (S)-2-hydroxy-acid oxidase immobilized to the inner surface of nylon tubing is accurate, precise, and sensitive but unsuitable for routine use because, even immobilized, the oxidase is unstable and can only be used for 12 days. We have used the semiautomated assay routinely: it has a mean analytical recovery of 96% (SD 4.2%), a within-batch CV less than 2%, and a between-batch CV less than 5%. The normal reference interval for urinary excretion of glycolate so measured is 0.13 to 1.31 mmol per day (n = 55).