Amperometric determination of oxalate in plasma and urine by liquid chromatography with immobilized oxalate oxidase.
A detection system for plasma and urinary oxalate involving a postcolumn enzymatic reaction and electrochemical detection is described. Oxalate oxidase was immobilized on AF-Tresyl Toyopearl 650 gel. The immobilized oxalate oxidase was packed into a stainless-steel column (10 x 4 mm I.D.) and used on-line as an immobilized enzyme reactor (IMER). The hydrogen peroxide produced by enzymatic reaction was detected amperometrically at a platinum electrode maintained at +0.5 V vs. Ag/AgCl. The HPLC separation of oxalate was carried out using a Capcell Pak C8 column and an isocratic mobile phase containing 80 mM KH2PO4 and 5 mM tetra-n-butylmammonium phosphate as an ion-pair reagent. The IMER was active and stable in the mobile phase employed. The plot of peak height against concentration of oxalate was linear in the range 0.1-1.6 mumol/ml with a correlation coefficient of 0.9989. The detection limit was 10 nmol/ml at a signal-to-noise ratio of 3. The within-day and between-day coefficients of variation were 5.3 and 7.9%, respectively.
Department of Analytical Chemistry, Niigata College of Pharmacy, Japan.
S
S
Yamato
H
H
Wakabayashi
M
M
Nakajima
K
K
Shimada
D002851:Chromatography, High Pressure Liquid; D004563:Electrochemistry; D004800:Enzymes, Immobilized; D006801:Humans; D006861:Hydrogen Peroxide; D006863:Hydrogen-Ion Concentration; D010070:Oxalates / Q000032:analysis* / Q000097:blood / Q000652:urine; D010088:Oxidoreductases; D012996:Solutions
D004800:Enzymes, Immobilized; D010070:Oxalates; D012996:Solutions; D006861:Hydrogen Peroxide; D010088:Oxidoreductases; C020598:oxalate oxidase