Toxic effects of soil benzo(a)pyrene multi-time superimposed pollution on antioxidant enzymes activities of Eisenia fetida coelomocytes

Benzo(a)pyrene (BaP), one of the most persistent organic pollutants (POPs), accumulates in soil and thereby poses a serious threat to soil environment quality. There have been frequent overestimations of environmental risks of BaP due to the over-evaluated toxic effects of available fractions with one-time imposed pollution method. In this paper, we proposed a new multi-time addition method aimed at reliable simulation of the process of BaP infiltration into soils. Furthermore, soil available BaP contents, accumulated BaP in earthworms (Eisenia fetida) and the toxic effects on superoxide dismutase (SOD) and peroxidase (POD) activities in earthworms coelomocytes were investigated by using superimposed multi-time and one-time pollution methods. The results suggested that the contents of available BaP and its accumulated amounts in earthworms sharply declined in 1 28 d and then slowly decreased in the following 28 56 d of incubation time under both superimposed multi-time and one-time pollution methods. The rates of decrease of soil available BaP were 2.37 μg·kg^-1·d^-1 and 3.35 μg·kg^-1·d^-1 in 1 28 d and 0.24 μg·kg^-1·d^-1 and 0.53 μg·kg^-1·d^-1 in 28 56 d for the superimposed multi-time and one-time pollution methods, respectively. The accumulated BaP in earthworms declined at the rates of 6.94 μg·kg^-1·d^-1 and 14.84 μg·kg^-1·d^-1 in 1 28 d, and 0.73 μg·kg^-1·d^-1 and 1.64 μg·kg^-1·d^-1 in 28 56 d under the superimposed multi-time and one-time pollution methods, respectively. The highly significant positive correlation (R² = 0.914 7,P < 0.01) was noted between accumulated amounts of BaP in earthworms and Tenax extracted contents of BaP from soils. Moreover, SOD and POD activities in earthworms were positive correlated with soil available BaP contents, with best-fit regression equations of y = 0.118 6x + 3.595 and y = 0.114x + 17.727 (where y is enzyme activity and x is the available BaP content in soil) and correlation coefficients R² = 0.754 3 and 0.829 6, respectively. Also SOD and POD activities in earthworms were positively correlated with accumulated amounts of BaP in earthworms, with best-fit regression equations of y = 0.028 9x + 4.524 8 and y = 0.026 9x + 18.803 (where y is enzyme activity and x is accumulated quantity in earthworm) and correlation coefficients R² = 0.704 0 and 0.727 1, respectively. Although soil available BaP content was significantly (P < 0.05) correlated with enriched BaP amount in earthworms under the superimposed multi-time pollution method, the correlation coefficient was lower for superimposed multi-time pollution than for one-time pollution assay. Under superimposed multi-time pollution, soil available BaP content and accumulated amounts of BaP in earthworms were 17.1% 38.6% (P < 0.05) and 22.6% 46.8% (P < 0.05) lower than those under one-time pollution, respectively. The ranges of SOD and POD activities in earthworm coelomocytes under superimposed multi-time pollution were respectively 49.6% 82.7% and 75.5% 109.6% lower than those under one-time pollution method during the entire incubation time. The results suggested that soil available BaP content and its toxicity to earthworms under superimposed multi-time pollution were lower than those under one-time pollution.