New ways of enzymatic two-substrate determinations in flow injection systems Original Research Article

This contribution presents two new conceptional strategies for the simultaneous determination of two substrates in flow injection systems. In most biotechnological processes, the substrate consumed as well as the product produced are of special interest for supervision and control. In most cases, substrate and product are interrelated via a redox reaction chain or the law of mass action, respectively. Their enzymatic analysis using dehydrogenases needs often in most cases the use of the cosubstrate in opposite redox states (NAD+ or NADH). The consequence in a simultaneous two-substrate analysis is a more complex experimental setup. This paper describes two principal new concepts, using only one cosubstrate solution and in situ generation of the cosubstrate in the opposite redox state or a procedure using equilibrium constants, respectively. So opposite configuration requirements, or two substrates which are interrelated as substrate and product of the enzymatic analysis can be determined simultaneously, keeping the analysis system minimised. The determination of ethanol/acetate and the analysis of glucose/fructose serve as models for the proposed methods. A comparison of the results with established standard analysis kits is given.