A Rapid High-Performance Ion-Chromatographic Procedure for the Estimation of Glycolate in Plasma and Urine
The authors describe a direct HPIC procedure for the determination of glycolate in urine and plasma. Plasma (1.0 ml) was mixed with HC1 (0.5 ml of 75 mM), heated to 100°C for five min, and centrifuged (1). Urine was diluted 1:2 with purified water. The samples were subsequently filtered (0.22-μ filter) and an aliquot (50 μl of urine, 200 μl of plasma) was injected into the HPIC which was equipped with an AG6 guard column, an AS6 analytical column, and an anion micro-membrane suppressor column (Dionex Corporation, Sunnyvale, California, USA). The flow rate of the elution solution was 1.0 ml/min, programmed by a solvent select valve: 0–25 min with 12-mM sodium borate, 25–30 min with 25-mM sodium carbonate, 30–40 min with 50-mM sodium borate, and 40–50 min with 25-mM sodium borate. The flow rate of the regenerant solution (12-mM H2SO4) was 3.0 ml/min. All reagents were of analar grade or equivalent and solutions were prepared with purified water. The total run time was 50 min. The specificity of the procedure was confirmed by treatment of an aliquot of the sample with glycolate oxidase, noting the complete absence of detector response at the elution time for glycolate. The recovery of added 1- 14C-labelled glycolate to plasma (determined by collection of the eluting glycolate peak) was 100.9±1.75(±SD)% (n=6). The recovery of added glycolate to urine samples was 98.8±2% (n=9). Intra-assay precision in urine and plasma was 2.5% (n=20) and 7.8% (n=11), respectively. The sensitivity was 20 ng injected onto the column. The urinary glycolate concentrations are comparable to those determined by previous authors (2, 3). The normal plasma glycolate levels (5.3–9.1 ymol/1) as determined by this technique are considerably lower than 0.10–0.30 mmol/1 (4) and 0.12–0.33 mmol/1 (5), but agree with the concentrations determined by J. Vrbanac and C. Sweeley (unpublished data) quoted by Tolbert (6). These results are more in keeping with recently-published plasma-oxalate concentrations (0.8–1.5 ymol/1 (7) and 1.3–3.1 ymol/1 (8), and the method which we have developed for plasma glyoxylate which has yielded preliminary values of less than 250 nmol/1 (Hughes, Hagen, and Sutton, unpublished observations).
KeywordsUrine Sample Unpublished Observation Detector Response Previous Author Elution Time
- 5.PM Zarembski, PhD Thesis, University of Leeds (1968).Google Scholar