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1、沈阳师范大学 硕士学位论文 化肥磷酸二铵对花背蟾蜍蝌蚪的毒性效应研究 姓名：李贞 申请学位级别：硕士 专业：细胞生物学 指导教师：陆宇燕 20100301 化肥磷酸二铵对花背蟾蜍蝌蚪的毒性效应研究 I 化肥磷酸二铵对花背蟾蜍蝌蚪的毒性效应研究 摘 要 摘 要 近年来，随着环境污染的日益加重，两栖类动物物种在全球范围内正急剧衰 减或消失，已有研究证明此现象发生与化肥污染有关。本文从个体、细胞和组织 的不同水平研究了磷酸二铵 （Diammonium Phosphate） 对花背蟾蜍 (Bufo. Raddei) 蝌蚪的毒性效应，实验结果如下： （一） 由于有卵胶膜的存在，磷酸二铵对蝌蚪孵化期前的影
2、响较小，孵化所需 时间基本相同，孵化率均在90%以上。 （二） 蝌蚪经磷酸二铵处理后，体重和体长均小于对照组，且随浓度升高和染 毒时间延长而降低。被处理的蝌蚪表现出腹部膨大且透明，个别出现头部膨大的 现象，23期处理组鳃丝发育明显小于对照组等畸形现象。 （三） 磷酸二铵处理对蝌蚪具有明显的致死效应，且染毒时间及磷酸二铵浓度与 死亡率具有明显的相关性。 （四）磷酸二铵对蝌蚪肝脏的结构损伤主要表现在：首先，肝细胞表现出明显的 膨胀现象，由于充血造成了肝血窦而显著扩增；而色素沉积由块状分布变为指突 状分布。 且在高浓度处理组中， 肝脏内出现由于淋巴细胞浸润而形成的块状坏死； 第二，肝细胞出现脂肪样变
3、性；由于核内异染色质呈凝集状态，故胞核整体呈淡 染现象。最明显特征是24期处理组中，随浓度升高枯否氏细胞数量也呈增多的趋 势。第三，电镜观察，随着磷酸二铵浓度的升高及染毒延长，肝细胞胞质中脂滴 数量和大小也之增多和增大；糖原颗粒数量增多，并出现了多聚糖原；线粒体双 层膜结构基本完好，但嵴结构大部分结构模糊不清；粗面内质网上核糖体出现普 遍的脱颗粒现象，且在高浓度处理下集聚在胞膜内侧分布，且在胞质内可观察到 大量髓样结构。 （五）磷酸二铵引起蝌蚪肝脏内糖原含量变化，低浓度染毒（0.18g/l,0.24g/l） 蝌蚪的糖原含量均较对照组高，且随浓度增大而升高。 （六）磷酸二铵对蝌蚪肝脏内酶系酶活也
4、有影响，低浓度染毒可以诱导24期和42 期蝌蚪过氧化氢酶活性升高，抑制31期和46期酶活性；低浓度染毒可以诱导蝌蚪 超氧化物歧化酶活性升高，高浓度染毒（1.4g/l）引起蝌蚪酶活随染毒时间呈先 降后升变化；低浓度染毒（0.18g/l,0.24g/l）可以诱导31期和42期蝌蚪乳酸脱 氢酶酶活性成高，抑制24期和46期酶活性。 化肥磷酸二铵对花背蟾蜍蝌蚪的毒性效应研究 II （七）低浓度染毒，蝌蚪微核率及核异常率均随磷酸二铵浓度升高而上升；高浓 度染毒，蝌蚪微核率随染毒时间呈先升后降变化，核异常率则保持上升。 关键词：磷酸二铵，花背蟾蜍蝌蚪，生长发育，微核率，核异常率 化肥磷酸二铵对花背蟾蜍蝌蚪
5、的毒性效应研究 III The Toxicological Effects of Fertilizers( The Toxicological Effects of Fertilizers(DAP ) on the Larvae of ) on the Larvae of Bufo. Raddei Abstract In recent years, as pollution increased, amphibian species of worldwide, is rapidly attenuated or disappeared, studies have shown that this p
6、henomenon has been occurring with the fertilizer pollution. In this paper,we studied the toxic effects of fertilizer DAP (Diammonium Phosphate) on raddei (Bufo. Raddei) in different levels(tissues, cells and individuals ).The results as follows: The impact of DAP on the tadpoles hatching was not obv
7、ious,the time of final incubation were same and hatching rate were above 90%. The weight and body length of tadpole in treatment group is small than those in the control group with the concentration and exposure time-dependent decrease.DAP treated tadpoles showed abdominal swelling and transparent,
8、appeared head enlargement , the gill filament of phenomenon 23 in treatment group developed smaller than control group , appeared reduction in surface characteristics of pigment . DAP have lethal effects on the tadpoles and the mortality increased with prolonged exposure. In treatment group,the tadp
9、ole liver micro-structural damage manifested in: Showed fatty degeneration of the liver, liver sinusoid widened, the liver cell nucleus pyknosis state, the distribution of pigment from the block into a sudden-like distribution of means.The most obvious feature is the 24 treatment group appeared libr
10、aries in cell and the numbe increased with concentration;High concentrations occured within treatment group in the liver due to the formation of massive lymphocytic infiltration and necrosis;Ultrastructural features: The cytoplasm appeared a large number of lipid droplets, glycogen particles increas
11、ed, the distribution of mitochondria reduced and the structure of crest is not clear, endoplasmic reticulum structure was normal but the distribution is reduced, hepatic blood cell distribution has increased 化肥磷酸二铵对花背蟾蜍蝌蚪的毒性效应研究 IV sinus base and its distribution within the cytoplasm of a large numb
12、er of lysosomal granules, the most obvious feature is the degranulation of endoplasmic reticulum, the marrow-like structure appeared of the cytoplasm in high concentration group. DAP caused changes of glycogen content in liver . In low-concentration exposure group(0.18g / l, 0.24g / l), the glycogen
13、 content was higher compared with control group, and increased with the concentration. DAP also affect the activity of liver enzymes, low-concentration exposure could induce CAT increased in 24 and 42 tadpoles and decreased in31 and 46 ; low concentrations could induce tadpoles exposed to SOD activi
14、ty increased;high concentration exposure (1.4g / l) caused SOD activity increasing the first then falling down; low concentration exposure (0.18g / l, 0.24g / l) could induce 31 period and 42 tadpoles of LDH activity into high, inhibited 24 and 46 tadpoles . Low-concentration exposure, tadpoles micr
15、onucleus and nuclear abnormalities rate increased with the rise in the concentration of DAP, high-concentration exposure, tadpoles exposed to time with the micronucleus rate was first increased and then decreased change, nuclear anomaly rate remained increased. Keywords: DAP, Bufo.Raddei tadpole, gr
16、owth and development, micro-nuclear rate, nuclear anomalies rate 学位论文独创性声明 本人所呈交的学位论文是在导师的指导下取得的研究成果。 据我 所知，除文中已经注明引用的内容外，本论文不包含其他个人已经发 表或撰写过的研究成果。对本文的研究做出重要贡献的个人和集体， 均已在文中作了明确说明并表示了谢意。 作者签名： 日期： 学位论文使用授权声明 本人授权沈阳师范大学研究生处， 将本人硕士学位论文的全部或 部分内容编入有关数据库进行检索； 有权保留学位论文并向国家主管 部门或其指定机构送交论文的电子版和纸质版， 允许论文被查阅和借 阅； 有权可以采用影印、 缩印或扫描等复制手段保存、 汇编学位论文。 保密的学位论文在解密后适用本规定。 作者签名： 日期： 化肥磷酸二铵对花背蟾蜍蝌蚪的毒性效应研究 1 第一部分 引言 第一部分 引言 一水环境污染介绍 世界的水资源为 13.5 亿 mk3，其中 9.65%是海水，28%是淡水，扣除冰川和 冰山后，可利用的淡
