oa Interim : Interdisciplinary Journal - Evaluation of a spectrophotometric method to follow hydrolysis of acetates with whole yeast cells
The rate limiting factor when screening for hydrolysis of esters by whole yeast cells or monitoring these reactions, involves the analysis of numerous samples by TLC, GC, and / or HPLC. Ester hydrolysis can be monitored with pH indicators such as para-nitro-phenol (p-NP) because when the indicator and reaction buffer have the same pKa, the absorbance change is directly proportional to ester hydrolysis. Such systems are used to monitor hydrolysis reactions catalysed by isolated enzymes. However, the use of such systems to monitor hydrolysis using whole microbial cells is complicated. We are evaluating a spectrophotometric method to monitor acetate hydrolysis by whole yeast cells in microtiter plates. In order for the method to work, very low cell and buffer concentrations must be used. This means that the method requires high hydrolytic activity. Conditions must still be optimised to correlate the values obtained from the microtiter plate reader with GC results.
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