使用生物测试方法评价水质毒性的研究进展

doi:10.11894/iwt.2021-0345

摘要/Abstract

摘要：

生物测试方法包括体外生物测试与体内生物测试，是水质毒性评价中的重要手段。综述了常用于水质毒性评价的体外和体内生物测试方法。针对体外生物测试，介绍了细胞毒性与遗传毒性实验的方法及应用现状，并着重论述了重组受体报告基因法的生物活性终点种类和效应触发值；针对体内生物测试方法，阐述了体内实验的受试生物种类、毒性测试方法与毒性终点判断。在此基础上，比较了体外生物测试和体内生物测试的优缺点，并对生物测试应用于水质毒性评价的未来研究方法提出了建议，旨在为今后生物测试方法在水质毒性评价中的应用及相关标准制定提供参考。

关键词: 体外生物测试, 体内生物测试, 细胞毒性, 遗传毒性, 报告基因实验

Abstract:

Bioassays, including in vitro and in vivo bioassays, are important methods for aquatic toxicity evaluation. The in vitro and in vivo bioassay methods commonly used in aquatic toxicity evaluation were reviewed. In regard to in vitro bioassays, the methods and the application of cytotoxicity and genotoxicity assays were briefly introduced and the bioactivity endpoints and effect-based trigger values of reporter gene assays were expounded detailedly. For in vivo bioassays, the subject organisms, methods of toxicity assays and toxicity endpoints were discussed. Based on these, the merits and demerits of in vitro and in vivo bioassays were compared and the future development of bioassays were suggested, aiming to provide reference for the future research and standard setting on aquatic toxicity evalation by bioassays.

Key words: in vitro bioassays, in vivo bioassays, cytotoxicity, genotoxicity, reporter gene assay

中图分类号:

X703

胡贤达,耿金菊,于清淼,许柯,张徐祥,任洪强. 使用生物测试方法评价水质毒性的研究进展[J]. 工业水处理, 2021, 41(6): 14-24.

Xianda Hu,Jinju Geng,Qingmiao Yu,Ke Xu,Xuxiang Zhang,Hongqiang Ren. Research progress of aquatic toxicity evaluation using bioassays[J]. Industrial Water Treatment, 2021, 41(6): 14-24.

https://www.iwt.cn/CN/Y2021/V41/I6/14

图/表 4

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受体名称	英文简称	生物活性	参考文献
雌激素受体	ER	（抗）雌激素效应^a	〔44-45〕
雄激素受体	AR	（抗）雄激素效应^a	〔44-45〕
孕激素受体	PR	（抗）孕激素效应^a	〔44-45〕
糖皮质激素受体	GR	（抗）糖皮质激素效应^a	〔42, 45〕
盐皮质激素受体	MR	（抗）盐皮质激素效应^a	〔42, 45〕
芳烃受体	AhR	平面芳烃生物反应，调节异生素代谢酶如CYP 1A1^b	〔6, 46〕
孕烷X受体	PXR	异生素代谢反应，调节CYP450	〔6, 46〕
过氧化物酶体增殖物激活受体	PPARγ	控制细胞内多种代谢过程，尤其在脂质代谢中起重要作用	〔6, 46〕
甲状腺受体	TR	（抗）甲状腺激素效应^a	〔47〕

受体名称	英文简称	生物活性	参考文献
雌激素受体	ER	（抗）雌激素效应^a	〔44-45〕
雄激素受体	AR	（抗）雄激素效应^a	〔44-45〕
孕激素受体	PR	（抗）孕激素效应^a	〔44-45〕
糖皮质激素受体	GR	（抗）糖皮质激素效应^a	〔42, 45〕
盐皮质激素受体	MR	（抗）盐皮质激素效应^a	〔42, 45〕
芳烃受体	AhR	平面芳烃生物反应，调节异生素代谢酶如CYP 1A1^b	〔6, 46〕
孕烷X受体	PXR	异生素代谢反应，调节CYP450	〔6, 46〕
过氧化物酶体增殖物激活受体	PPARγ	控制细胞内多种代谢过程，尤其在脂质代谢中起重要作用	〔6, 46〕
甲状腺受体	TR	（抗）甲状腺激素效应^a	〔47〕

活性终点	生物实验类别^a	EBT值^b	注释	参考文献
ER活性	ERα-CALUX	0.28 ng/L EEQ	斑马鱼胚胎体内效应	〔82〕
	MELN	0.56 ng/L EEQ
	ER-GeneBLAzer	0.24 ng/L EEQ
	Hela-9903	0.18 ng/L EEQ
	YES	0.50 ng/L EEQ
	ER-CALUX	0.5 ng/L EEQ	由最低有影响浓度推导	〔83〕
	YES、ER-CALUX、MELN、MVLN	0.1~0.4 ng/L EEQ	由多篇文献总结得出，位于上行的是长期暴露EBT值，下行为短期暴露	〔80〕
	YES、ER-CALUX、MELN、MVLN	0.5~2.0 ng/L EEQ
	ER-CALUX	3.8 ng/L EEQ	针对饮用水得出	〔84〕
	YES	12 ng/L EEQ	针对饮用水得出	〔85〕
	ER-CALUX	0.2 ng/L EEQ
AR活性	AR-GeneBLAzer	14 ng/L TTEQ	针对饮用水得出	〔85〕
	AR-CALUX	11 ng/L DHTEQ	针对饮用水得出	〔84〕
PR活性	PR-CALUX	560 ng/L LevoEQ	针对饮用水得出	〔84〕
GR活性	GR-CALUX	21~150 ng/L DexaEQ	针对饮用水得出	〔84~85〕
抗-AR活性	antiAR-CALUX	14.4 μg/L FEQ	初步应用于地表水评估	〔16〕
抗-PR活性	antiPR-CALUX	13 ng/L REQ	初步应用于地表水评估	〔16〕

活性终点	生物实验类别^a	EBT值^b	注释	参考文献
ER活性	ERα-CALUX	0.28 ng/L EEQ	斑马鱼胚胎体内效应	〔82〕
	MELN	0.56 ng/L EEQ
	ER-GeneBLAzer	0.24 ng/L EEQ
	Hela-9903	0.18 ng/L EEQ
	YES	0.50 ng/L EEQ
	ER-CALUX	0.5 ng/L EEQ	由最低有影响浓度推导	〔83〕
	YES、ER-CALUX、MELN、MVLN	0.1~0.4 ng/L EEQ	由多篇文献总结得出，位于上行的是长期暴露EBT值，下行为短期暴露	〔80〕
	YES、ER-CALUX、MELN、MVLN	0.5~2.0 ng/L EEQ
	ER-CALUX	3.8 ng/L EEQ	针对饮用水得出	〔84〕
	YES	12 ng/L EEQ	针对饮用水得出	〔85〕
	ER-CALUX	0.2 ng/L EEQ
AR活性	AR-GeneBLAzer	14 ng/L TTEQ	针对饮用水得出	〔85〕
	AR-CALUX	11 ng/L DHTEQ	针对饮用水得出	〔84〕
PR活性	PR-CALUX	560 ng/L LevoEQ	针对饮用水得出	〔84〕
GR活性	GR-CALUX	21~150 ng/L DexaEQ	针对饮用水得出	〔84~85〕
抗-AR活性	antiAR-CALUX	14.4 μg/L FEQ	初步应用于地表水评估	〔16〕
抗-PR活性	antiPR-CALUX	13 ng/L REQ	初步应用于地表水评估	〔16〕

受试生物	水样类型	毒性终点	暴露时间	注释	参考文献
藻类（S. obliquus）	市政污水处理厂进出水	生长抑制、叶绿素a浓度、SOD活性、细胞膜完整性	72 h	能够检出部分污水厂出水毒性没有削减，叶绿素a浓度、SOD活性相对更敏感	〔87〕
藻类（S. obliquus）	地表水	光合作用二阶段（PSII）抑制	4.5 h	39个地点只有一处有抑制作用，化学分析表明可能主要归因于除草剂利奴隆	〔12〕
四种无脊椎动物（Lumbriculus variegatus，Chironomus riparius，Potamopyrgus antipodarum，Daphnia magna）	污水处理厂臭氧化出水	存活率、干生物量、羽化时间、胚胎数量、个体数	7~28 d	臭氧化出水对无脊椎动物产生一定抑制，夹杂带丝蚓（L. variegatus）表现最敏感	〔88〕
斑马鱼（Danio rerio）胚胎	地表水	死亡率、孵化率、发育异常（畸形、心包水肿、卵黄囊水肿、血凝、色素沉着、眼缺陷等）	24~96 h	对水样进行了浓缩	〔14〕
斑马鱼（Danio rerio）胚胎	市政污水处理厂出水	死亡率、亚致死率、遗传毒性（彗星实验）、ABC转运蛋白活性（反应细胞损伤）	24~48 h	斑马鱼胚胎实验是废水毒性评估的灵敏有效工具，均测出与对照组显著差异	〔89〕
黑头呆鱼（Fathead minnow）	回用水	成年雄性鱼血浆卵黄蛋白原含量（反映雌激素活性的生物标志物）	21 d	实验组为对照组的0.5~3.2倍，表明影响较小	〔90〕
斑马鱼（Danio rerio）	污水处理厂臭氧化出水	成年雄性鱼卵黄蛋白原基因表达，繁殖成功率、游泳行为	21 d	几个终点均观察到相比对照组异常	〔91〕
褐鳟鱼（Salmo trutta f. fario）和虹鳟鱼（Oncorhynchus mykiss）	臭氧化、活性炭深度处理前后水样	肝脏单加氧酶和EROD酶（反映二噁英类物质对鱼类胁迫的生物标志物）	40~100 d	该研究支持臭氧活性炭深度处理，有利于改善鱼类健康	〔92〕
鳌虾（Astacus leptodactylus）	市政污水处理厂出水	过氧化氢酶（CAT）、超氧化物歧化酶（SOD）活性、脂质过氧化（TBARS）和谷胱甘肽（GSH）水平（氧化应激生物标志物）	24~96 h	是评价水样引起氧化应激的灵敏生物标志物	〔93〕
欧洲鲈鱼（Dicentrarchus labrax）	毒性实验配制水样	乙酰胆碱酶活性（AchE）（反应神经毒性的生物标志物）	96 h	—	〔94〕

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