Acute Toxicity And Estrogenic Effects Of Nonylphenol To Rana Chensinensis Tadpoles

With the rapid development of industry and agriculture, environmental contamination became seriously. A well known group of organic compound is the alkylphenol polyethoxylates (APEs), which are widely used in industry and daily life. APEs undergo a biodegradation process to give derivatives such as nonylphenol (NP) and octylphenol (OP), which have estrogenic effects. It brought an extreme threat on survival and reproduction of vertebrates. In this study, Rana chensinensis tadpoles were exposed to NP in acute toxicity tests and chronic aqueous exposure tests, and acute toxicity, development and sexual differentiation were examined after that period to evaluate the effects and the mechanisms of estrogenic effects of NP. The main results and conclusions as followings:1. A standard method was used to evaluate the acute toxicity of nonylphenol on Rana chensinensis tadpoles at the water temperature of 21±1℃.The results showed that the regress equation between the death probability of Rana chensinensis tadpoles and the concentration logarithm of NP were y=8.4087x+10.202, y=9.5104x+11.745, y=10.284x+12.498 and y=10.619x+13.095 respectively for 24, 48, 72, 96h. The mean LC₅₀ values were 0.24, 0.20, 0.19 and 0.17 mg·L^-1 respectively. The mean SC(safe concentration) values was 0.017 mg/l, The mean 96h LCo values was 0.14 mg · L^-1.2. Rana chensinensis tadpoles were continuously exposed to 100, 60, 30, 10μg ·L ^-1 NP and 3, 0. 3μg·L^-1 E2 (positive control). Exposure occurred throughout the stage 26 to stage 66, and weight, total length were examined at day20, day40, complete metamorphosis respectivel.Time of metamorphosis were examined at period of metamorphosis. The results showed that NP did not affect mortality of Rana chensinensis tadpoles. 100 and 10μg ·L^-1 NP had similar effects induced by 3μg·L^-1 E₂, reduced the weight of froglets (metamorphs) and delayed the time of metamorphosis. It suggested that NP did not directly damage the organism but disrupted the endocrine system to affect the development of Rana chensinensis tadpoles under 100μg·L^-1 NP.3. The aim of this study was to evaluate both estrogenic effects in directly NP-exposed wild Rana chensinensis tadpoles and possible transgeneration effects in the offspring of exposed mature Rana chensinensis. Analysis of post-metamorphic sex ratio,indicated as followings: (1) wild tadpoles were exposed to NP, beween lOOjag ? L ' and 3Ofjg ? L"1, female ratio was elevated. A tendency for a dose-dependent increase in the female ratio was observed in the NP expose groups. It suggested NP induced feminization in wild Rana chensinensis tadpoles. The necessary concentration of NP for inducing feminization was at least 102 times higher than that of E2. (2) Only female mature exposed to NP would had transgeneration effects on the offspring (3) Female progeny were exoposed to NP. Their maximal female ratio was 85. 2%, towered above the wild female ratio about 40%. This study demonstrated sub-lethiferous NP concentrations in water affected gonadal development and sexual differentiation in amphibian, and induce sex ratio deviating and reproductive obstacle. It resulted in population delines in amphibian species.4. To understand the effects of NP on the level of immunoreation of testosterone (T), aromatase (ARO), androgen receptor (AR) and estrogen receptor (ER) by immunocytochemisry in gonad in froglets of NP-exposed wild tadpoles. It indicated as followings: (1) The level of immunoreactive T was reduced in the gonad. (2) NP was in agreement with the effects of E2. An increase in the level of immunoreactive ARO was found in the gonad. (3) NP had similar effects induced by E2, reduced the level of immunoreactiveAR in the gonad. (4) The level of immunoreactiveER was just reduced in the testis of froglets of wild tadpoles were exposed to lOOug ? L"1 NP. (5) There was a similar effect on the level of immunoreactiveT, ARO, AR, ER in testis and ovary of froglets of NP-exposed wild tadpoles. Our results further suggested that NP increased the activity of ARO, and converted testosterone to estrogen.lt induced an increase in the E2 relative content in gonad. E2 inhibited the expression of AR, and inhibited androgen binding of AR, At last, NP induced feminization in gonad of froglets from NP-exposed wild tadpoles.