采用多次叠加污染的方式模拟菲(Phenanthrene, Phe)在土壤中逐步累积的过程，分析在不同老化阶段(1、7、14、28和56 d)土壤Phe的生物有效性特征，及其对蚯蚓体腔细胞酶活性、染色体和溶酶体膜的毒性效应。结果表明：随着土壤培养时间的延长，土壤中Phe有效含量、蚯蚓体内Phe富集量、蚯蚓体腔细胞超氧化物歧化酶(Superoxide dismutase, SOD)和过氧化物酶(Peroxidase, POD)活性、丙二醛(Malondialdehyde, MDA)含量以及微核率均呈前期（1~28 d）下降较快，后期（29~56 d）下降速率逐渐减小的趋势，中性红保留时间呈逐渐升高的趋势，表明随着培养时间的延长，Phe对蚯蚓的毒性效应逐渐减小。土壤Phe有效含量与蚯蚓体内Phe富集量、SOD和POD酶活性、MDA含量、微核率和中性红保留时间均呈极显著相关（p<0.01），决定系数（R2）分别为0.887 9、0.893 4、0.870 9、0.930 5、0.893 3和0.885 1。在1~56 d老化时间内，多次叠加污染土壤中Phe有效含量和蚯蚓体内Phe富集量分别较一次污染低23.19%~50.15%和12.64%~57.89%；蚯蚓体腔细胞SOD和POD酶活性分别较一次污染低3.40%~47.24%和6.49%~38.02%；MDA含量比一次污染低1.30%~57.82%；微核率较一次污染低12.48%~22.99%；中性红保留时间较一次污染高1.35%~13.97%，这表明土壤Phe多次叠加污染对蚯蚓体腔细胞的毒性效应低于一次污染。
Phenanthrene (Phe), a typical persistent PAHs (polycyclic aromatic hydrocarbons) type organic pollutant, is accumulating step by step in low dosage, posing a potential threat to the soil environment. As one-time Phe pollution of the soil differs in process from gradual accumulation of Phe pollution in low dosage, it is very likely to overestimate environmental risk of the Phe entering the soil in the first manner. Phe entering the soil does not diffuse or translocate in the soil. Therefore, most Phe stays in aqueous phase in the soil due to short soil-Phe contact time. In this paper, a multiple-time addition method was used to simulate of the process of Phe accumulation in the soil. With the multiple-time addition approach, Phe was added into the soil in low dosage 12 times, whereas with the one-time addition method, the same quantity of Phe was added into the soil all at once. Then, bioavailability and toxic effects on enzyme activity in earthworm coelomocytes, chromosome and lysosome membrane of the earthworms exposed to the pollutant at different aging phases (1, 7, 14, 28, and 56 d) were analyzed. It was found that with soil incubation going on, content of available Phe, Phe accumulation in earthworm, activity of SOD and POD in earthworm coelomocytes, content of MDA and micronucleus rates (MNR) all dropped rapidly in the early period (1~28 d) and slowed down their drops gradually in the late period (29~56 d), but neutral red retention time (NRRT) kept rising on steadily, indicating that Phe toxicity to earthworm declined with soil incubation going on. Under the condition of multiple-time overlying pollution and one-time pollution, soil available Phe decreased by 1.12 µg kg-1 d-1 and 3.08 µg kg-1 d-1, respectively, during the early stage (1~28 d) of incubation and by 0.17 µg kg-1 d-1 and 0.39 µg kg-1 d-1, respectively, during the late stage (29~56 d) and Phe accumulation in earthworms did by 7.59 µg kg-1 d-1 and 21.46 µg kg-1 d-1, respectively, during the early stage, and by 0.63 µg kg-1 d-1 and 1.91 µg kg-1 d-1, respectively, during the late stage. Soil available Phe (Tenax-TA extractable Phe) was in highly significant positive relationship (R2=0.887 9, p<0.01) with Phe accumulation in earthworms and in highly significant relationship, too, with SOD and POD activities, MDA content, micronucleus rate and NRRT in earthworm coelomocytes with the coefficient of determination being 0.893 4, 0.870 9, 0.930 5, 0.893 3 and 0.885 1, respectively. During the 56 days of aging, soil available Phe and Phe accumulation in earthworms was 23.19%~50.15% and 12.64%~57.89% lower, respectively; SOD and POD activities in earthworm coelomocytes 3.40%~47.24% and 6.49%~38.02%, respectively; MDA content 21.30%~57.82% lower; and MNR in earthworm coelomocytes 2.48%~22.99% lower, while NRRT in earthworm coelomocytes was 1.35%~13.97% higher in the soil subjected to multiple-time overlying pollution than in the soil subjected to one-time pollution, which indicates that Phe entering into the soil through multiple-time overlying pollution is lower than that through one-time pollution in toxicity to earthworms. This finding can be cited as theoretic basis for assessment of eco-toxicity of PAHs-type organic pollutants in soil.
马静静,钱新春,张 伟,郑 彬,葛高飞,郜红建.土壤菲多次叠加污染对蚯蚓的毒性效应[J].土壤学报,2015,52(6):1374-1382. DOI:10.11766/trxb201501230053 Ma Jingjing, Qian Xinchun, Zhang Wei, Zheng Bin, Ge Gaofei, Gao Hongjian. Toxic effect of multiple-time overlying pollution of Phe in soil on eisenia fetida[J]. Acta Pedologica Sinica,2015,52(6):1374-1382.复制