DETERMINATION OF SUBSTRATES USING POLY(ETHYLENE GLYCOL)-STABILIZED DEHYDROGENASE ENZYMES BY MICROLITER PER MINUTE FLOW-INJECTION

Flow injection (FI), at a flow rate of mu l min(-1), is an effective m ethod for enzymic substrate determinations using low concentrations of poly(ethylene glycol) (PEG)-stabilized soluble enzymes. PEG stabilize s dehydrogenase enzymes for at least several days by promoting sub-uni t association. Band broadening of knitted open tubular reactors is red uced as flow rate decreases below 300 mu l min(-1) and a small tubing diameter is important for a faster rate of absorbance signal increase with residence time. Small (0.5 mu l) sample injections also ensure na rrow FI peaks. The determination of several substrates such as pyruvat e, lactate, and cortisone using appropriate PEG-stabilized enzymes is demonstrated with this FI instrument at 25 or 50 mu l min(-1) with sam ple throughputs of the order of 2-3 min per sample. The determination of lactate in serum samples is also possible. The advantage of this me thod, sample throughput, is not sacrificed but enzyme consumption is c onsiderably less, compared to standard mi min(-1) FI.