The amino acid fermentation by Corynebacterium glutamicum was monitored with a new technique that uses the first derivative of the NAD(P)H fluorescence signal. The rate of change of NAD(P)H pools is indicative of intracellular redox balance variations that correspond to metabolic changes. The profile of this signal showed several characteristics that coincided with major metabolic events during fermentation. We show here that the derivative fluorescence signal can accurately estimate points of threonine depletion, viable cell count, and the end of amino acid formation. Furthermore, on-line optimization strategies can be developed by using the derivative fluorescence signal.