[Establishment of method for cytochrome P450 of earthworms (Eisenia fetida) as a biomarker].

Earthworm (Eisenia fetida) was chosen as test soil fauna to establish the method for determination of cytochrome P450 content. By means of thoroughly washing with salt solution, changing centrifugal acceleration and dissolving microsomal membranes of earthworms with sodium cholate, the determination of cytochrome P450 was performed. Base on the method, dynamic relationships of dose-response for cytochrome P450 contents in earthworm and phenanthrene concentrations were conducted by contact filter paper and soil contaminated with phenanthrene. Results indicate that cytochrome P450 are covered up by interferential material in earthworm, which makes the classic peak lag and appear at 455 - 457nm in the CO difference spectrum with a spectrophotometer. Through increasing centrifugal acceleration and adding reagent of solubilization, the interference is removed mostly and the classic peak of P450 returns to appear at 450nm +/- 1nm. The exposure tests of phenanthrene with contact filter paper (10(-6), 10(-5), 10(-4), 10(-3) and 10(-2) mg x mL(-1)) and soil (1, 2, 4 and 8 mg x kg(-1)) reveal that the dose-response relationships exist between the pollutant and cytochrome P450. Soil tests of different exposure durations (1, 3, 7, 14 and 28d) show the effects of different phenanthrene concentration on earthworm cytochrome P450 contents are to activate (7d) and to inhibit (14d and 28d), and the content of P450 is 0.99 - 1.41 and 0.77 - 0.88 (p < 0.05) times of control respectively. From this study, cytochrome P450 content of earthworm has the merit of simpleness, speediness and economy for determination, and it could be used as a sensitive biomarker for monitoring the exposure of sublethal pollution in terrestrial ecosystem.