DETERMINATION OF COENZYME A AND ACETYL COA IN TISSUE EXTRACTS

A sensitive and specific method for the determination of coenzyme A and acetyl CoA in tissue extracts is described. CoA is recycled through a coupled enzyme system in which the rate of formation of NADH is proportional to CoA concentration when the necessary enzymes and substrates are present in excess. The same system can be used to measure acetyl CoA in tissue extracts by first binding free CoA irreversibly with N-ethylmaleimide. The method is convenient in that all of the required reagents are commercially available. The rate of NADH production can be followed either spectrophotometrically or, with greater sensitivity, fluorometrically. With the latter, concentrations of less than 40 picomoles per milliliter can be measured. This method was used to determine the CoA and acetyl CoA content of perchloric acid extracts of various rat tissues. Close agreement was found with literature values for liver acetyl CoA but CoA values were generally much lower than those previously reported, apparently because of increased specificity. © 1969.