Use of Liquid Chromatography Mass Spectrometry for Measurement of Microsomal UGT Activity

Sagebrush ( Artemisia tridentate ) produces a large number of toxic chemicals to protect itself from herbivores. ^4,5 Nevertheless, it is a major part of the diet of sage-grouse ( Centrocercus urophasianus ) to the extent that over the winter, sagebrush constitutes around 90% of the sage grouse’s diet. ⁵ In order to understand how sage-grouse can be proficient at metabolizing sagebrush toxic compounds, we designed a protocol to analyze the UDP-glucuronosyltransferase (UGT) enzymes activity in vitro using mice microsomes. UGT enzymes add a glucuronic acid group to substances through a condensation reaction. ^1,2,3 4-methylumbelliferone ² was used as a substrate for the enzyme. Liquid chromatography mass spectrometry (LCMS) was used to demonstrate that the amount of 4-methylumbelliferone decreased over time, and a product, 4-methylumbelliferone glucuronide appeared correspondingly. This protocol will be used to investigate the transformation of sagebrush compounds by sage-grouse liver UGTs.