Studies on an on-line reaction of micro lactate dehydrogenase in capillaryelectrophoresis using electrochemical detection

The method for an on-line reaction of micro lactate dehydrogenase in capill ary electrophoresis using electrophoretically mediated micro-assay(EMMA) co upled with electrochemical detection(EC) was developed. The running electro lyte(pH 9,2) was composed of 50 mmol/L Tris-HCl, 1,0 mmol/L nicotinamide ad enine dinuceotide(NAD(+)), 5,0 mmol/L lithium lactate and 0, 2 mmol/L cetyl pridinium bromide (CPB). LDH was injected into the capillary, As it migrate d electrophoretically in the capillary, LDH catalyzed the reaction between lactate and NAD+. The product NADH of the enzyme-catalyzed reaction was mon itored by a bundle of carbon fiber electrode(150 mum diameter) held at +0.8 V vs, SCE, The detection limits for LDH activity using constant-voltage mo de and zero-voltage mode in EMMA were determined to be 1.1 X 10(-9) U and 6 ,0 X 10(-10) U, respectively. The formulae concerning the width or the heig ht of the plateau in the electropherogram for EMMA were deduced, and some s ignificant conclusions were drawn from these formulae, The developed method has been used for the determination of ultramicro LDH activity in serum wi th satisfactory results.