Rapid spectrophotometric determination of p-nitrophenylpropionate esterase activity in rat tissues

✍ Scribed by J.P. Liberti

• Publisher: Elsevier Science
• Year: 1968
• Tongue: English
• Weight: 430 KB
• Volume: 23
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(68)90008-0

✦ Synopsis

The detection of nonspecific e&erase activity in various tissues was the subject of several reports in the 1930's (l-3). The disadvantages of these methods which employ stalagmometric, titrimetric, and manometric measurements have been underscored by Huggins and Lapides (4)) who have developed a more sensitive procedure. Their method utilizes the development of color from the hydrolysis of colorless esters of p-nitrophenol.

Using various acyl esters of pNP-OH1 in conjunction with certain inhibitors, these authors, as well as Gomori (5), were able to distinguish three types of e&erase activity in animal tissues. Aldridge (6, 7) has classified these activities as A-, B-, or C-type esterases according to their solubility characteristics and their response to certain inhibitors. Using pNPP as the substrate, Niemi et al. ( 8) recently reported a B-type esterase of rat testis which is remarkably affected by hypophysectomy.

These workers employed the calorimetric method of Huggins and Lapides (4) for the enzyme assays.

Our laboratory has been interested in the effects of bovine growth hormone on certain hepatic esterases. We have employed in a preliminary way the method of Huggins and Lapides using pNPP as the enzyme substrate to measure the enzyme activity in rat liver extracts. However, because of the large number of tubes required for the determination and because of the wide variation in enzyme activities commonly obtained, it was necessary to modify greatly the conditions of the assay in order to satisfy these objections. The results of these modifications form the basis of this report. In brief, the method to be described involves the continuous spectrophotometric measurement of the reaction product, pNP-OH. The average time required for one determination is about 30 sec.