中国科学院海岸带环境过程与生态修复重点实验室知识库

随着社会经济的快速发展，我国土壤污染态势愈加严峻。重金属是一类分布广泛、毒性较强、受到重点管控的土壤污染物。矿山开采、金属冶炼、大气沉降以及农用化学品的频繁施用是土壤重金属的主要来源。为保护土壤环境质量安全，我国以重金属总量为限定值分别制定了农用地土壤和建设用地土壤环境质量风险管控值。然而，重金属的生物毒性不仅与总量有关，而在更大程度上由生物有效性决定，而化学形态又在很大程度上由其化学形态决定。以生物有效性为基础的土壤环境质量基准和标准研究是现阶段我国土壤环境质量管理的重要研究方向之一。土壤中重金属的化学形态可以通过具有不同溶解性和提取性的单一的和连续逐级组合的化学试剂来区分。筛选具有普适性的重金属有效态提取方法，并表征不同形态对污染土壤重金属生物有效性的影响是制定以生物有效性为基础的重金属污染限值的关键。

铅锌矿是我国主要有色金属矿之一。铅锌矿区及周边土壤重金属复合污染突出。铜是铜矿区、果园土壤和农药波尔多液（硫酸铜）生产企业及其周边土壤主要重金属污染物。本研究选择湖南省铅锌矿区砷（As）、镉（Cd）、铅（Pb）复合污染土壤和铜（Cu）污染模拟农田土壤为研究对象，针对不同区域土壤中主要生态受体、暴露途径和保护目标，探究重金属的化学形态、生物有效性及其相互关系。主要研究内容有：针对铅锌矿区重金属复合污染土壤，利用体外胃肠模拟提取法，分析人体通过口腔暴露复合污染土壤中As、Cd和Pb的生物可给性；结合小鼠模型测得的相对生物有效性，探讨体外胃肠模拟提取法评估重金属复合污染土壤人体生物有效性的适用性；评估土壤理化性质对重金属化学形态及人体生物有效性的影响；针对不同性质的人工模拟Cu污染农田土壤，采用具有不同提取能力的化学试剂提取黑土、潮褐土、脱潜水稻土中的有效态Cu；通过暴露实验，分析Cu在生菜和蚯蚓等敏感生物中的富集特征及毒性效应，阐明其与有效态Cu的关系，并进一步推导以化学提取有效态为基础的Cu生态毒性阈值。本研究得到的主要结果如下：

（1）供研究的铅锌矿区复合污染土壤中重金属总量、有机碳、无定形Fe、Al含量等是影响该区域土壤重金属化学形态分配的主要因素。

（2）铅锌矿区复合污染土壤UBM（Unified Bioaccessibility Research Group Europe Method）体外胃肠模拟提取法胃相As、Cd、Pb的生物可给性与小鼠模型测得的相对生物有效性相关性较好（R2 >0.67）,可以表征复合污染土壤重金属的人体生物有效性。

（3）铅锌矿区复合污染土壤中As、Cd、Pb的有效性受重金属总量、土壤铁锰铝及氧化物含量等基本理化性质和金属化学形态的影响。醋酸提取态、氧化结合态和有机结合态重金属是人体生物可利用的主要形态。

（4）三种农田土壤中不同提取能力的化学试剂提取的效果不同。酸溶性（HNO3）（41.38%）和络合性（EDTA-Na2）（56.81%）提取剂对3种土壤中Cu的平均提取效率显著高于交换性（NH4OAc）（0.12%）和弱交换性（CaCl2）（8.70%）提取剂。生菜Cu富集量及其毒性效应与CaCl2提取态Cu含量相关性最好，蚯蚓Cu富集量及其死亡率与HNO3提取态Cu含量相关性最好，

（5）选用物种最敏感指标推导了3种农田土壤Cu有效态毒性阈值的EC 20（20%抑制浓度）和EC 50（50%抑制浓度）。基于不同化学提取态Cu含量对生菜的毒性阈值EC 20范围分别是90.45~170.10 mg/kg（HNO3提取），102.78~195.31 mg/kg（EDTA-Na2提取），3.97~20.06 mg/kg（NH4OAc提取），和0.21~8.68 mg/kg（CaCl2提取）；EC 50范围分别是110.48~187.60 mg/kg（HNO3提取），118.63~230.49 mg/kg（EDTA-Na2提取），5.69~32.23 mg/kg（NH4OAc提取）和0.26~9.62 mg/kg（CaCl2提取）。基于不同化学提取态Cu含量对赤子爱胜蚓死亡率的毒性阈值EC 20范围分别是138.26~193.16 mg/kg（HNO3提取），107.80~225.88 mg/kg（EDTA-Na2提取），8.92~11.58 mg/kg（NH4OAc提取），和0.36~10.57 mg/kg（CaCl2提取）；EC 50范围分别是183.07~221.23 mg/kg（HNO3提取），180.38~331.09 mg/kg（EDTA-Na2提取），13.06~18.30 mg/kg（NH4OAc提取）和0.54~13.21 mg/kg（CaCl2提取）。

（6）不同农田土壤中，基于化学提取有效态Cu浓度的毒性EC 20 和EC 50值存在较大差异。潮褐土中Cu对生菜和蚯蚓的毒性EC值较低，黑土中较高。土壤中的大量共存阳离子及溶解性有机质等对Cu的生物毒性起到了缓解作用。

本研究对基于重金属化学形态和生物有效性的土壤环境风险管控具有重要意义。

The rapid development of social economy has resulted in serious soils pollution in China, among which heavy metals are a kind of soil pollutants with wide distribution, high toxicity and should be controlled primarily. Mining, smelting, atmospheric deposition and frequent agricultural chemicals application are main sources of heavy metals in soils. China has formulated the environmental quality standards based on heavy metals total contents for agricultural and construction lands, respectively, to protect their quality and safety. However, the bioavailability of heavy metals is not only related to their total contents, but also to their chemical speciations to a greater extent. At present, the revision of environmental quality standards for soils based on bioavailability is one of the most important research directions of soil environmental quality management in China. The chemical speciations of heavy metals in soils can be distinguished by single and successive chemical reagents with different solubility and extractability. Therefore, selecting the universal bioavailability evaluation methods to characterize different heavy metals speciations on bioavailability is a key step for developing limits of heavy metals based on bioavailability in contaminated soils.
Lead-zinc mine is one of the main non-ferrous metal mines in China. Pollution of multiple heavy metals is prominent in soils surrounding lead-zinc mining areas. Copper is the main heavy metal pollutant in copper mining areas, orchard soils, pesticide Bordeaux liquid (copper sulfate) production enterprises and their surrounding soils. In this study, heavy metals chemical speciations, bioavailability and their relationships were studied in As, Cd and Pb co-contaminated soils in lead-zinc mining areas of Hunan province and three farmland copper contaminated soils, based on different biological receptors, exposure pathways and protection targets in soils of different regions. Firstly, the human oral bioaccessibility of As, Cd and Pb in lead-zinc mining areas co-contaminated soils were measured by the UBM (Unified Bioaccessibility Research Group Europe Method), and then compared to relative bioavailability in the mouse model, with the aim to evaluate the applicability of UBM in co-contaminated soils. The effects of soil physico-chemical properties and heavy metal chemical speciations on human bioavailability were further studied. Then the available Cu in three farmland soils (black soils, meadow cinnamon soils andunsubmerged paddy soil) were extracted by 4 chemical reagents with different extraction capacities, and the relationships between Cu enrichment as well as toxic effects in lettuce and earthworm and chemically extracted Cu were established. Furthermore, the ecotoxicity thresholds of Cu were derived based on chemically extracted Cu. The main results of this study were summarized as follows:
(1) Heavy metals total contents, organic carbon, and amorphous Fe and Al oxides were important factors affecting the chemical fractions of As, Cd and Pb in 12 co-contaminated soil of lead-zinc mining areas.
(2) UBM (Unified Bioaccessibility Research Group Europe Method) gastric phase bioaccessibility was highly correlated with the relative bioavailability of As, Cd and Pb in mice (R2 > 0.67), and used to characterize the bioavailability of heavy metals in co-contaminated soil.
(3) The bioavailability of As, Cd and Pb in co-contaminated soils of mining areas was affected by physical and chemical properties, such as heavy metals total contents, Fe/Mn/Al oxides and their chemical speciations. The acetic acid extracted fraction, oxidized bound fraction and organic bound fraction of heavy metals were the main bioavailable fractions for human.
(4) The extraction effects of chemical reagents with different extraction capacity in the three farmland soils were different. The average extraction efficiency of Cu by acid-soluble (HNO3) (41.38%) and complex (EDTA-Na2) (56.81%) reagents was significantly higher than that of the exchangeable (NH4OAc) (0.12%) and weakly exchangeable (CaCl2) (8.70%) reagents in the three soils. Cu accumulated and Cu toxic effect in lettuce correlated best with CaCl2 extracted Cu, while Cu accumulated and mortality in earthworms correlated best with HNO3 extracted Cu.
(5) The EC 20 (20% inhibitory concentration) and EC 50 (50% inhibitory concentration) based on chemically extracted Cu in three soils were derived by selecting the most sensitive indicator of the species. The range of ecotoxicity thresholds EC 20 for lettuce based on different chemically extracted Cu contents was 90.45~170.10 mg/kg (HNO3 extraction), 102.78~195.31 mg/kg (EDTA-Na2 extraction), 3.97~20.06 mg/kg (NH4OAc extraction), and 0.21~8.68 mg/kg (CaCl2 extraction); the range of EC 50 was 110.48~187.60 mg/kg (HNO3 extraction), 118.63~230.49 mg/kg (EDTA-Na2 extraction), 5.69~32.23 mg/kg (NH4OAc extraction) and 0.26~9.62 mg/kg (CaCl2 extraction). The range of ecotoxicity thresholds EC 20 for earthworm Eisenia foetida based on different chemicallyextracted Cu contents was 138.26~193.16 mg/kg (HNO3 extraction), 107.80~225.88 mg/kg (EDTA-Na2 extraction), 8.92~11.58 mg/kg (NH4OAc extraction), and 0.36~10.57 mg/kg (CaCl2 extraction), respectively; the range of EC 50 was 187.07~221.23 mg/kg (HNO3 extraction), 180.38~331.09 mg/kg (EDTA-Na2 extraction), 13.06~18.30 mg/kg (NH4OAc extraction) and 0.54~13.21 mg/kg (CaCl2 extraction).
(6) The ecotoxicity thresholds values of Cu EC 20 and EC 50 based on chemically extracted Cu were significantly different in different farmland soils. In meadow cinnamon soils, the ecotoxicity thresholds values of EC 20 and EC 50 for lettuce and earthworms were higher than that of in black soils. This may due to the higher amounts of coexisting cations and dissolved organic matter in black soils, which have a certain alleviation effect on the biological toxicity of Cu.
This study is of great significance for soil environmental risk management based on chemical heavy metals chemical speciation and bioavailability.

第1章绪论............................................................................................................1
1.1 我国土壤重金属污染现状...........................................................................1
1.1.1 矿区周边土壤重金属污染................................................................1
1.1.2 农田土壤重金属污染........................................................................2
1.1.3 城市及工业场地土壤重金属污染....................................................2
1.2 土壤中重金属的化学形态...........................................................................2
1.3 土壤中重金属的生物有效性.......................................................................3
1.4 土壤中重金属生物有效性的影响因素.......................................................5
1.4.1 重金属来源........................................................................................5
1.4.2 土壤理化性质....................................................................................6
1.4.3 重金属形态........................................................................................7
1.4.4 生物受体............................................................................................8
1.5 土壤重金属生物有效性表征方法...............................................................8
1.5.1 化学提取法........................................................................................8
1.5.2 生物评价法......................................................................................11
1.6 生物有效性在土壤重金属毒性阈值研究和质量标准制定中的应用.....15
1.7 本研究拟解决的科学问题和研究目标、研究内容及技术路线.............15
1.7.1 科学问题..........................................................................................15
1.7.2 研究目标..........................................................................................16
1.7.3 研究内容..........................................................................................16
1.7.4 技术路线..........................................................................................17
第2章铅锌矿区复合污染土壤中砷、镉和铅的化学形态..............18
2.1 前言.............................................................................................................18
2.2 研究材料与方法.........................................................................................18
2.2.1 采样区域概况..................................................................................18
2.2.2 土壤样品采集..................................................................................19
2.2.3 土壤基本理化性质分析..................................................................19
2.2.4 土壤中重金属总量及化学形态分析..............................................20
2.2.5 质量控制和数据处理......................................................................22

2.3 结果与讨论.................................................................................................22
2.3.1 矿区复合污染土壤理化性质及重金属总量..................................22
2.3.2 矿区复合污染土壤中砷的化学形态..............................................25
2.3.3 矿区复合污染土壤中镉的化学形态..............................................26
2.3.4 矿区复合污染土壤中铅的化学形态..............................................28
2.3.5 矿区复合污染土壤砷、镉和铅化学形态的影响因素..................30
2.4 小结.............................................................................................................33
第3章铅锌矿区复合污染土壤中砷、镉和铅的人体生物可给性和相对生物有效性..................................................................................34
3.1 前言.............................................................................................................34
3.2 研究方法.....................................................................................................34
3.2.1 土壤中砷、镉和铅的人体生物可给性测定..................................34
3.2.2 土壤中砷、镉和铅的相对人体生物有效性测定..........................36
3.2.3 数据处理..........................................................................................37
3.3 结果与讨论.................................................................................................37
3.3.1 矿区复合污染土壤中砷、镉和铅的生物可给性..........................37
3.3.2 矿区复合污染土壤中砷、镉和铅人体生物可给性的影响因素..40
3.3.3 矿区复合污染土壤中砷、镉和铅的相对人体生物有效性..........42
3.3.4 矿区复合污染土壤中砷、镉和铅人体生物可给性与相对人体生物有效性关系..................................................................................48
3.4 小结.............................................................................................................51
第4章不同性质农田土壤-生菜系统中铜的化学形态与生物有效性......................................................................................................................................52
4.1 前言.............................................................................................................52
4.2 研究材料与方法.........................................................................................52
4.2.1 土壤样品采集与基本理化性质测定..............................................52
4.2.2 外源铜添加和老化..........................................................................53
4.2.3 土壤中铜的化学形态分析..............................................................53
4.2.4 生菜盆栽实验..................................................................................54
4.2.5 生菜毒性阈值..................................................................................54
4.2.6 数据统计..........................................................................................55
4.3 结果与讨论.................................................................................................55
4.3.1 农田土壤中铜的化学形态..............................................................55

4.3.2 农田土壤中生菜毒性与铜化学形态关系......................................57
4.3.3 农田土壤中生菜可食部位累积量与铜化学形态关系..................59
4.3.4 基于生物有效性的农田土壤中铜的生菜毒性阈值......................60
4.4 小结.............................................................................................................62
第5章不同性质农田土壤-蚯蚓系统中铜的化学形态与生物有效性...................................................................................................................64
5.1 前言.............................................................................................................64
5.2 研究材料与方法.........................................................................................64
5.2.1 试验材料..........................................................................................64
5.2.2 蚯蚓急性毒性实验..........................................................................64
5.2.3 蚯蚓慢性毒性实验..........................................................................64
5.2.4 蚯蚓生态毒性阈值..........................................................................65
5.2.5 数据处理..........................................................................................65
5.3 结果与讨论.................................................................................................65
5.3.1 农田土壤中蚯蚓死亡率与铜化学形态关系..................................65
5.3.2 农田土壤中蚯蚓累积铜含量与铜化学形态关系..........................69
5.3.3 基于生物有效性的农田土壤中铜的蚯蚓毒性阈值......................69
5.4 小结.............................................................................................................71
第6章总讨论与总结论..................................................................................72
6.1 总讨论.........................................................................................................72
6.1.1 土壤理化性质对重金属化学形态的影响......................................72
6.1.2 重金属化学形态与生物有效性关系..............................................72
6.1.3 基于生物有效性的土壤重金属毒性阈值和土壤环境质量基准..73
6.2 总结论.........................................................................................................74
6.3 论文创新点.................................................................................................74
6.4 论文不足与研究展望.................................................................................75
6.4.1 论文不足..........................................................................................75
6.4.2 研究展望..........................................................................................75
参考文献...................................................................................................................77
附录........................................................................................................................91
英文缩略对照表.....................................................................................................91
致谢........................................................................................................................93

作者简历及攻读学位期间发表的学术论文与研究成果..........................95