A BIOELECTROCHEMICAL METHOD FOR THE DETERMINATION OF ACETATE WITH IMMOBILIZED ACETATE KINASE

Flow injection determinations of acetate were carried out using immobi lized acetate kinase, pyruvate kinase and lactic dehydrogenase with an amperometric method. Two acetate kinases from E. coli and B. stearoth ermophilus were tested. It was found that the immobilized acetate kina se from B. stearothermophilus was more stable than that from E. coli., but it is much more expensive and less available. Acetate kinase coup ling at pH 7.4 using CPG aminopropyl and glutaraldehyde seems to be su perior to other immobilization methods. A high immobilization yield ca n be obtained by immobilization of the three enzymes separately giving high conversions of all the three. Plots of current versus concentrat ion show a useful operating range from 0.3 to 2 mM acetate with a line ar response. The detection limit was 0.2 mM at a now rate of 0.3 ml/mi n with 200 mu l injections. The method is therefore well suited for mo nitoring of the level of acetate in fermentations with acetate as the carbon source.