How do you determine the micromoles of NADH consumed in an enzyme activity test?

To determine the micromoles of NADH consumed in an enzyme activity test, the process involves quantifying the change in absorbance as the reaction proceeds. NADH, a coenzyme that can absorb light at specific wavelengths, changes in concentration during the reaction, resulting in measurable changes in absorbance.

The correct method involves utilizing the change in absorbance in conjunction with the molar absorptivity coefficient, which represents how strongly a chemical species absorbs light at a given wavelength. By dividing the observed change in absorbance by the molar absorptivity coefficient, one can derive the concentration of NADH that has been utilized in the reaction. This calculation directly leads to determining the micromoles of NADH consumed based on the reaction conditions established in the test.

Understanding this process is crucial, as it allows for accurate quantitation of enzyme activity, which is often expressed in terms of substrate (here, NADH) consumption over time. This method is a fundamental principle in enzymatic assays, as it links spectrophotometric measurements to the biochemical activity being studied.