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题名	环境和生物基质中二烷基次膦酸铝的分析方法、水解行为与分布研究
作者	牛宇敏
学位类别	博士
答辩日期	2015-05
授予单位	中国科学院研究生院
授予地点	北京
导师	刘景富
关键词	二烷基次膦酸铝，二烷基次膦酸，分析方法，水解行为，分布特征，Aluminum dialkyl phosphinates, dialkyl phosphinate acids, analytical methods, hydrolyzation, distribution
其他题名	Aluminum dialkyl phosphinate in environmental and biological matrices: analytical methods, hydrolyzation and distribution
学位专业	分析化学
中文摘要	二烷基次膦酸铝（Aluminum dialkyl phosphinates, ADPs）是近年来发展起来的一种新型阻燃剂。由于具有产品密度低、阻燃剂用量少、机械性能较好、色泽较佳、烟密度较低、相对漏电起痕指数值较高等优点，其在电子电气工业中具有广泛的应用前景，被认为是替代溴代阻燃剂的最有前途的产品之一，目前已在多个国家用于电子元器件等产品。同其他阻燃剂一样，ADPs 有可能在生产或使用过程中有意或无意地排放进入环境，对环境以及人类健康造成潜在危害。然而，目前关于这类物质的研究只集中在各种新型化合物的合成、性能优化以及应用领域的推广等方面，关于其环境行为研究和生物安全性的评价仍处于空白阶段。     本文旨在研究ADPs 在环境及生物基质中的分布和转化，这就需要分析测定环境及生物基质中ADPs 的含量。ADPs 的理化性质较为特殊，既难溶于有机溶剂，也很难溶解于水，因此为分析检测带来了困难。所以本研究选择了目前应用较多的三种ADPs，分别为甲基乙基次膦酸铝（Aluminum methylethyl phosphinate，AMEP）、二乙基次膦酸铝（Aluminum diethylphosphinate，ADEP）和甲基环己基次膦酸铝（Aluminum methylcyclohexyl phosphinate，AMHP），从其溶解性着手进行研究，基于离子色谱串联质谱（IC-MS）和液相色谱串联质谱（LC-MS）检测方法，建立了环境水体、土壤和沉积物以及生物基质中三种ADPs 及其水解产物二烷基次膦酸（Dialkyl phosphinate acids，DPAs）的分析测定方法，并对典型区域土壤和沉积物中ADPs 的分布特征和迁移转化规律进行了初步研究。最后采用 体外实验和体内实验结合的方法，研究了ADPs 在体内的分布和转化情况。论文主要包括以下几个部分：     首先，分别建立了IC、LC-MS 和IC-MS 分析测定ADPs 的方法。三种方法各有优缺点：带有电导检测器的离子色谱的方法定量限为100 μg/L，该方法操作简单，对仪器要求不高，适用于基质较为简单的样品中ADPs 含量较高时的测定；IC-MS 法可以实现对三种ADPs 的同时监测，该方法灵敏度高，其定量限为0.01 μg/L，可以实现复杂基体中痕量ADPs 的测定；LC-MS 法同样也可以实现对三种ADPs 的同时监测，方法的定量限为1.0 μg/L。相比IC-MS 法，LCMS法对质谱检测器的污染和伤害更小，同时也更简单、高效，分离时间显著降 低，可以实现复杂基体中痕量ADPs 的快速测定。     其次，本文对ADPs 的溶解性能进行了初步探索。ADPs 很难溶于有机溶剂，也很难溶于水，可溶解于酸或碱。利用LC-MS、核磁共振（NMR）以及电感耦合等离子体质谱（ICP-MS）等多种技术手段，发现ADPs 一旦溶解于水，就会水解为DPAs，因此采用上述三种不同分析方法测定的ADPs 均是以DPAs 的形式。此外，ADPs 的水解与pH 值有显著关系，ADPs 在pH 值为中性条件下，溶解度最低，随着pH 值的升高或降低，ADPs 的水解程度都会升高。当溶液中Ca2+和Mg2+含量较高时，也会促进ADPs 的水解。为了更好地研究ADPs 在环境中的转化，本实验利用已有的ADPs 标准品，合成了纯度均在90%以上的三种对应的水解产物DPAs。     由于ADPs 一旦溶解于水，就会水解为DPAs，所以本研究认为环境水体中ADPs 主要以DPAs 的形式存在。因此建立了环境水体中三种相应的DPAs 的测定方法，分别为甲基乙基次膦酸（Methylethyl phosphinatea acid，MEPA），二乙基次膦酸（ Diethyl phosphinate acid ， DEPA ） 和甲基环己基次膦酸（Methylcyclohexyl phosphinate acid，MHPA）。采用MAX 固相萃取柱对环境水体样品进行净化和富集，IC-MS 进行测定，该方法通过大体积进样的方式以及柱后添加甲醇和氨水的方式进一步提高灵敏度。方法的定量限（Limits of quantification，LOQs）为3.5−9.3 ng/L，该方法灵敏度高，重现性好，适用于自来水、河水、污水处理厂进水及出水等不同环境水体样品中DPAs 的测定。     本文建立了土壤和底泥中痕量ADPs 及其水解产物DPAs 的测定方法，并利用该方法研究了某生产厂周边的土壤和底泥样品中ADPs 和DPAs 的污染水平与迁移转化。所建立的测定方法为：（1）样品提取，以75 mM 氨水为提取溶剂，同时提取ADPs 和DPAs；以甲酸-水-甲醇（5:5:90，v/v/v）为提取溶剂，仅提取DPAs；（2）样品净化方法，采用MAX 强阴离子交换固相萃取柱分别对上述提取液进行净化；（3）通过LC-MS/MS 分别测定上述净化液中ADPs 和DPAs 总和的含量以及单DPAs 的含量，两者之差即为ADPs 的含量。本方法的定量限为0.9−1.0 μg/kg，在三个水平的加标回收率为69.0−112.4%，相对标准偏差RSD <21%。应用本方法，对某生产厂周边的土壤和沉积物样品进行了分析，结果发现，在表层土壤中，ADPs 和DPAs 的含量分别为3.9−1279.3 μg/kg 和1.0−448 μg/kg。在废液废渣倾倒处和排污口的所有土壤和沉积物泥芯样品中均检测到ADPs，浓度范围为30.8−4628.0 μg/kg；与此同时，DPAs 的检出率> 90%，浓度范围为1.1−374.6 μg/kg。ADPs 和DPAs 在泥芯中的垂直分布与TOC 无关，而与样品的pH 值高度相关。此外，本研究表明土壤和沉积物中的DPAs 来源于ADPs 的水解。ADPs 的高度水解（高达49.6%）表明ADPs 一旦被排放，会与其水解产物共存于环境中。因此，为了评估ADPs 的环境风，有必要同时对ADPs 和DPAs的环境行为以及毒性进行进一步研究。     最后，本文建立了动物组织中痕量AMHP及其水解产物MHPA的测定方法，并初步研究了它们对雄性SD 大鼠暴露后在体内的分布与转化。对于固体组织样品，所建立的分析方法的定量限为19−2000 μg/kg；对于血清和尿液，方法的定量限为0.95−1900 μg/L，该方法只能对动物组织中AMHP 和MHPA 的总量加以测定。体外实验表明，AMHP 在人工模拟胃肠液中的水解率分别为46.6%和42.5%。由于本章方法不能区分组织中AMHP 和MHPA，为探索AMHP 在体内的转化，我们以MHPA 暴露组作为对照，研究了AMHP 对雄性SD 大鼠暴露后，AMHP 和MHPA 在组织、体液以及排泄物中的分布特征。AMHP 和MHPA的总量在AMHP 暴露组和MHPA 暴露组的血清、脑组织、肾脏、肝脏、尿液和粪便中的分布无显著差异，证实了AMHP 在体内会大部分水解为MHPA，并以MHPA 的形式吸收进入血液，然后到达各个组织。MHPA 主要通过粪便（75.0−79.2%）排出体外，其次为尿液（21.7−28.1%）。尽管超过95%的MHPA可以通过粪便和尿液排出体外，其在肝脏和肾脏中仍然有一定的富集。此外，在脑组织中检测到MHPA 提示这类物质可能产生类胆碱毒性效应，需要进一步研究。尽管通过病理学切片在肝脏组织观察到炎症反应， 但丙二醛（Malondialdehyde，MDA）水平、超氧化物歧化酶（Superoxide dismutase，SOD）活性和过氧化氢酶（Catalase，CAT）活性并没有改变，说明AMHP 及其水解产物MHPA 对大鼠仅具有轻微毒性。
英文摘要	Aluminum dialkyl phosphinates (ADPs) are one class of emerging phosphorus flame retardants due to their superior characteristics such as the low density, small dosage as flame retardant, high mechanical performance, fine color and luster, low smoke density, and high comparative tracking index. ADPs shows extensive application prospects and are considered one of the most promising flame retardant substitutes for the brominated flame retardants (BFRs). They have been used in the electronic and electrical industry components in many countries. With their increasing production and wide applications, ADPs will inevitably be introduced into environments. However, their analytical method, and occurrence and fates in environment and organism have not been reported yet.     This work focuses on the distribution and transformation of ADPs in the environmental and biological matrices, which strongly relys on the determination of trace level ADPs in these matrices. The analytical method of ADPs was urgently needed. Unlike the other common environmental polllutants, however, ADPs are hardly dissolved in the organic solvent and water, making the analysis of ADPs a great challenge. Three ADPs with the relatively high production, including aluminum methylethyl phosphinate (AMEP), aluminum diethylphosphinate (ADEP) and aluminum methylcyclohexyl phosphinate (AMHP), were selected as the target ADPs for this study. After testing theirsolubility in various solvents, we developed a series of analytical methods for the determination of ADPs and their hydrolysates (dialkyl phosphinate acids, DPAs) in water, soil and sediment, and biological tissue samples. With these analytical methods, we studied the distribution and transformation of ADPs in soil and sediment and organisms. This dissertation consists of the following parts:      Firstly, analytical methods based on ion chromatography (IC), ion chromatography tandem mass spectrometry (IC-MS) and liquid chromatography tandem mass spectrometry (LC-MS), respectively, were developed to measure the three target ADPs. The IC method is suitable for direct determination of ADPs without any sample purification, but it has low sensitivity as the limits of quantitation (LOQs) are about 100 μg/L. The IC-MS method is appropriate for simultaneous determinaiton of the three ADPs in complex matrix at trace levels, with LOQs of 0.01 μg/L. The LC-MS method is also suited for simultaneous analysis of ADPs with LOQs of 1.0 μg/L. Compared with IC-MS, the LC-MS method has less chances of contaminating mass spectrometry,higher efficiency and simplicity, and the separation time is greatly shortened. Therefore,LC-MS is preferred in determination of trace ADPs in complex environmental and biological matrices.     Secondly, the dissolubility of ADPs in various solvents were preliminary studied.We found that ADPs are hardly dissolved in water and organic solvents, but are dissolved in the acid and alkali waters. Comprehensively using LC-MS, nuclear magnetic resonance (NMR) and inductively coupled plasma mass spectrometry (ICPMS), we found that once dissolved in water, ADPs were hydrolyzed to DPAs. Therefore, what we determined in the above methods are actually the corresponding DPAs, the hydrolysis counterparts of ADPs. All the ADPs exhibite strongly pH dependent solubility, which shows the minimum value at pH 6.8. and dramatically increased at lower and higher pH values. It was also found that the presence of Ca2+/Mg2+ enhanced the solubility of ADPs in aqueous solutions. Additionally, to better understand the transformation of ADPs, DPAs were synthesized and characterized by NMR.     Given that once dissolved in water, ADPs are completely hydrolyzed and exist as DPAs, it is of interests to measure ADPs in environmental waters. However, there has no report on the determination of DPAs in environmental water samples. For the first time, we developed a method for analysis of trace DPAs, including methylethyl phosphinatea acid (MEPA), diethyl phosphinate acid (DEPA) and methylcyclohexyl phosphinate acid (MHPA) in different environmental water samples such astap water, river water, influent and effluent water of waste water treatment plant (WWTP). In this proposed method, MAX cartridges were employed for the purification, and IC-MS method with large volume injection (200 μL) and postcolumn addition of methanol and NH3·H2O were employed to enhance the method sensitivity. The limits of the method were 3.5–9.3 ng/L for DPAs in environmental water samples. The method is reliable,showing high sensitivity, good recovery and repeatability.     We further developed a method for the analysis of trace ADPs and their hydrolysates, and studied their occurrences and fates in soils and sediments. ADPs and DPAs in soil and sediment samples were determined by (i) extracting both ADPs and DPAs with 75 mM NH3·H2O, and selectively extract DPAs only with formic acid-watermethanol (5:5:90, v/v/v); (ii) quantifying the total content of ADPs and DPAs, and DPAs by LC-MS/MS analysis of the DPA contents in the former and the latter extract, respectively; and (iii) calculating ADPs from the content difference between the former and the latter extracts. The LOQs of the proposed method were 0.9−1.0 μg/kg, and the mean recoveries ranged from 69.0% to 112.4% with relative standard deviations ≤ 21% (n = 6). In soil and sediment samples around a manufacturing plant, ADPs and DPAs were detected in surface soils in the ranges of 3.9−1279.3 and 1.0−448.6 μg/kg, respectively.  While ADPs were found in all the samples of the soil and sediment cores from the drain outlet and the waste residue treatment site at levels ranging from 30.8 to 4628.0 μg/kg, DPAs were found in more than 90% of these samples with concentrations in the range of 1.1−374.6 μg/kg. The occurrences of ADPs and DPAs are not in correlation with the total organic carbon, whereas the occurrences of DPAs are highly correlated with the sample pH. Our study also suggests that the DPAs in the samples sourced from the hydrolysis of ADPs. The high hydrolysis degrees of ADPs (up to 49.6%) suggest that once released into the environment, ADPs are likely to coexist with their hyrolysates. Thus, to evaluate the environmental safety of ADPs, the environmental behavior and toxicity of both ADPs and DPAs should be considered.     At last, we developed a method for the analysis of trace AMHP and its hydrolysates MHPA in various biological tissues, serum and excretions, and preliminarily studied distribution and biotransformation of AMHP in male SD rats. The LOQs of the proposed method were 19−2000 μg/kg for solid samples and 0.95−1900 μg/L for serum and urine samples. With the developed analytical methods, we found that about 46.6% and 42.5% AMHP hydrolyzed to MHPA in simulated gastric juice and intestinal fluid, respectively. Since the proposed method could not distinguish AMHP and MHPA in bio tissues and excretions, we compared the distribution and clearance of total AMHP and MHPA after gavage of AMHP, with MHPA as control, to Sprague Dawley (SD) rats. The non-significant difference between the two exposure group in all the determined organs and excretions, including serum, brain, kidney, liver, urine and feces suggested that most of AMHP was hydrolyzed to MHPA in vivo, and then uptake into serum and distributed to different tissues in the form of MHPA. Fecal and urinary excretion eliminated 75.0−79.2% and 21.7−28.1% MHPA, respectively. Although > 95% MHPA could be excreted, enrichment of MHPA in liver and kidney occurred as the concentration in these organs were much higher than that in serum. MHPA was also detected in brain, which calls for further study on its cholinergic toxicity. The histology and biochemical indexes analysis showed slight inflammatory reaction in hepatic portal area in all exposure group, but the level of malondialdehyde (MDA) and the activities of superoxide dismutase (SOD) and catalase (CAT) in liver showed no obvious changes compared with the control rats, suggesting the toxicity of AMHP and MHPA to rat was mild.
内容类型	学位论文
源URL	[http://ir.rcees.ac.cn/handle/311016/34361]
专题	生态环境研究中心_环境化学与生态毒理学国家重点实验室
推荐引用方式 GB/T 7714	牛宇敏. 环境和生物基质中二烷基次膦酸铝的分析方法、水解行为与分布研究[D]. 北京. 中国科学院研究生院. 2015.

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