| Popis: |
A highly sensitive method for determination of the activity of epoxide hydrolase (EH) (EC 3.3.2.3) in the mammalian lymphocytes was developed. 9,10-Epoxy-9,10-dihydrophenanthrene was used as substrate. For elimination of the enzyme latency the lymphocytes were solubilized with lubrol PX, a nonionic detergent. 9,10-Hydroxy-9,10-dihydrophenanthrene was estimated quantitatively by the method of the internal standard (1-naphthol) after reversed phase HPLC. The time of chromatographic matographic separation was 2.5 minutes. By the pH optimum EH in the lymphocytes differed from microsomal EH. However, the structure of their active sites was similar. Among the species studied (mice, rats, guinea-pigs, man), the lowest activity of EH was detected in the lymphocytes of mice and the highest activity in the lymphocytes of guinea-pigs. The activity of EH in the lymphocytes of man was 10-20 times higher than the sensitivity limit of the method (5 pkmol/mg/min). By its sensitivity and reproducibility and the level of the activity determined the method is much more superior to the methods described in the literature. |
