MEASUREMENT OF ACETYL COENZYME A IN RAT LIVER

The estimation of acetyl CoA in rat liver samples by enzymic methods has been investigated with particular reference to the techniques used for obtaining the samples. In the assay it was found necessary to measure the NADH2 produced in the coupled enzyme reaction fluorometrically and to add internal standards of acetyl CoA to the cuvet at the start of the assay. Details of the instrumentation and methods used are given. For the estimation of acetyl CoA in the liver, speed in stopping the metabolism of the liver is of great importance, particularly in the fed rat. Here it is essential to use the quick-freezing technique (17,18) with the cooled tongs applied to the liver in situ. Interruption of the hepatic circulation for a few seconds before freezing leads to an increase in the acetyl CoA concentration in the liver of the fed rat. The quick-freezing technique is most easily done on the anesthetized rat and ether is considered the anesthetic of choice. A 24 hr fast gives a 4-fold increase in the acetyl CoA concentration in the liver. The larger amounts of acetyl CoA present permit rather more latitude in the methods that can be used to obtain the samples. Speed in freezing the liver is, however, still necessary and ether is again the anesthetic of choice. © 1969.