典型PhACs在城市水系统中的迁移分布规律及其在人工湿地中的去除研究
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摘要
由于人类对PhACs(pharmaceutically active compounds)的过分使用甚至是滥用，导致这类化合物及其代谢产物被持续不断地排入地表水、地下水及土壤等各种环境介质中，PhACs在环境中的迁移、转化、生态毒理及其控制处理已成为环境科技界日益关注的热点问题。而在我国，还鲜有此方面的研究报道。重庆地处长江和嘉陵江汇合处，饮用水源水质的优劣涉及到长江中下游上亿人民的饮水安全，近年来，重庆经济规模不断扩大，人口迅速增长，PhACs消费量巨大，而关于水环境中PhACs的行为与归趋的研究还未见报道。本研究采用高效液相色谱串联质谱法，系统地研究了典型PhACs在重庆水环境中的迁移分布规律；以CWS工艺作为PhACs进入环境的屏障技术，研究了目标PhACs在湿地中的行为与去除机理。具体的研究内容如下：
①复杂基质中PhACs的检测分析方法：
利用固相萃取技术富集水环境样品中的目标PhACs，利用超声辅助萃取提取、固相萃取净化污泥介质样品中的PhACs，通过优化各种前处理条件及色谱质谱的仪器条件，建立了基于HPLC-MS/MS的检测环境中痕量的21种目标PhACs(包括氟喹诺酮类抗生素、大环内酯类抗生素、磺胺类抗生素、止痛剂、调血脂药、降高血压药、他汀类药物与抗癫痫类药等八大类药物)残留的方法平台，目标PhACs在河水、污水厂的进水、出水样品及脱水污泥样品中的回收率分别在66.4~114.9%、62.5~133.2%、64.5%~111.9%和59.5%~139.2%之间，相对标准偏差小于16%；目标PhACs在河水、污水厂的进水、出水样品及脱水污泥样品中的LOQs分别为0.03~3.4ng/L、0.2~17.5ng/L,0.2~5.6ng/L和0.2~5.8μg/kg。
②典型PhACs在水环境中的含量、消除机制及风险评估：
对重庆市四家典型污水处理厂（鸡冠石污水厂、唐家沱污水厂、李家沱污水厂和井口污水厂）各工艺阶段的进水和出水以及各污水厂初沉池和二沉池的污泥进行采样检测分析，结果表明，21种目标PhACs均能在污水样品中检测出，除吉非罗齐、对乙酰氨基酚（ACM）与布洛芬外其余18种目标PhACs也能在污泥中检测出。目标PhACs在污水中的检测浓度范围在ng/L~μg/L，在污泥中的检出浓度在μg/kg(以干重计)水平；ACM具有最高的进水浓度（1.2~7.1μg/L），其次为磺胺甲噁唑（SMZ）、阿奇霉素（AZM）、罗红霉素（ROX）、氧氟沙星（OFX），AZM在污泥中具有最高的残留浓度（549.70μg/kg）。
除ACM外，其它目标PhACs并不能在污水处理工艺中完全去除。卡马西平（CBZ）、美托洛尔（MTP）和降固醇酸（CA）甚至出现了负增长。通过质量平衡计算分析发现，微生物降解作用是目标PhACs在污水处理过程中得以去除的主要机制，大部分目标PhACs通过污泥的吸附去除的量可忽略不计。不过，由于喹诺酮类抗生素及AZM含有带正电荷的氮原子或含带正电荷的二甲氨基组，由于静电引力的作用，污泥吸附也是其从水相中去除的相关途径之一。
基于PhACs在污水处理厂进水中的浓度水平及其在人体内的代谢动力学参数，建立了PhACs的消费量反演模型。通过该模型，能从城市水循环末端推算有关PhdACs消费量的数据，可以为药物监管相关部门提供科学的数据参考。
根据检测出的PhACs环境浓度(MEC)和预测无效应浓度(PNEC)两个重要数值获得风险表征比(MEC/PNEC)的风险评价方法,进行了污水处理厂出水、受纳水体及污泥中21种PhACs的环境风险分析。结果表明磺胺嘧啶（SDZ）、SMZ、OFX与红霉素（ERY）四种抗生素在污水处理厂出水及其污泥中综合评价因子均大于1。在受纳水体中所有目标PhACs的RQ值均小于1，但SDZ、OFX、ERY及CA的RQ值大于0.1小于1，可能会对环境产生中等程度的危害，其余药物均小于0.1。21种目标PhACs在三种不同的环境下（污水厂出水、污泥及受纳水体）的RQ(sum)均大于1，因此环境中痕量PhACs的残留会对周围生态环境产生不同程度的危害。
重庆地区饮用水源水中检测除了痕量浓度（ng/L水平）的PhACs，常规处理工艺并不能完全去除原水中含有的PhACs。大多数PhACs的去除主要发生在砂滤阶段，对于砂滤池出水中含有的PhACs，加氯消毒阶段几乎无影响。
③人工湿地系统（CWS）对PhACs的去除研究：
通过对备选填料（陶粒、碎石、石英砂和沸石）物化性质及去污效果的考察，筛选出了理想的湿地填料，在此基础上以陶粒为主填料建立了潜流CWS，考察了PhACs在CWS的去除效果，从生物学和植物学角度解析PhACs胁迫下，湿地微生物和湿地植物（旱伞草）的响应。结果表明：CWS能有效的去除目标PhACs，PhACs在0~500μg/L的浓度水平下，植物体内SOD、POD和CAT能通过相互协调作用，有效地消除植物过氧化所产生的活性氧化物，防御细胞膜过氧化；在PhACs胁迫下，旱伞草叶片光合作用受到一定影响，叶片有衰老的迹象，但这种影响会随CWS运行时间的增长而降低，最终不会对植物的生长状况产生较大影响。通过对指代微生物种群的PLFAs种类进行鉴别、聚类和主成分分析发现，进水PhACs浓度在0~500μg/L下，各CWS中主要功能菌群种类未发生改变，但是其顺序会有所变化，与对照CWS相比（Planted (0)），均可检测出新的PLFAs。进水PhACs浓度在0~500μg/L水平下，CWS中基质酶活性（过氧化氢酶、脱氢酶和脲酶）未受到抑制，在10μg/L与30μg/L下，土壤基质酶活性反而升高。
Pharmaceutically active compounds (PhACs) are widely used for the treatment ofbacterial diseases in humans, and some are applied to livestock and fish for diseaseprophylaxis and treatment or for growth promotion.Large amounts of PhACs and theirmetabolites are continuously released into environmental compartments such as rivers,lakes, even ground water and soil, because all kinds of drugs were misused seriously forlack of scientific direction. Thetransportation, transformation, ecological toxicology andcontrolof PhACs have therefore provoked considerably scientific attention around theworld over the past decade. However, only a few studies about the situation in Chinahave been reported. The Chongqing region is located at the confluence of YangtzeRiver and Jialing River, water quality of drinking water source influences the safety ofdrinking water of the middle and lower Yangtze River.During the past two decades,Chongqing has become one of the fastest growing economies and most denselyurbanized areas in the world, and pharmaceutical consumption of the region is large.However, there are no studies reporting the behavior and fate of PhACs in the aquaticenvironment of Chongqing.Therefore, the occurrence, behavior and environmental fateof PhACs in the aquatic environment of this regionwere studied using ahigh-performance liquid chromatography coupled to a tandem massspectrometer.Besides, we researched the behavior and removal mechanismPhACs in theconstructed wetland systems (CWS) which are the last defenses forPhACs entering intoenvironment. The results are as follows:
The methods for determination of selected PhACs in the environment
A sensitiveanalyticalmethodwas developed fordeterminingthe selected PhACs(including several antibiotics, analgesics, antiepileptics, antilipidemics andantihypersensitives) in the water and sludge. The wastewater samples were extracted byOasis HLB cartridges (6mL,500mg),while the solid samples (sludge and suspendedsolid matter) were extracted by ultrasonic-assisted extraction with solvents followed byan enrichment and clean-up step with solid-phase extraction using HLB cartridges. Theextracted PhACs were analyzed using high-performanceliquidchromatography-electrospray ionization tandem massspectrometry(HPLC-MS/MS) with multiple reactions monitoring(MRM).The recoveries achieved forthe target PhACs ranged from66.4~114.9%for the river water, from62.5~133.2%for the influent, from64.5%~111.9%for the effluent, and from59.5%~139.2%for thesludge and their relative standard deviation was below16%. Limits of quantification(LOQs)(signal-to-noise ratio10) ranged from0.03ng/L to3.4ng/L for the surfacewater, from0.2ng/L to17.5ng/L for influent wastewater, from0.2ng/L to5.6ng/L foreffluent wastewater, and from0.17μg/kg to5.83μg/kg for sludge, respectively.
Occurrence, transportation and ecotoxicological assessmentof the selectedPhACs in the aquatic environment
Twenty-one target PhACs from8therapeutic classes covering a wide range ofphysicochemical properties and biological activities, were analyzed at four full-scalewastewater treatment plants (WWTPs) in Chongqing. All the21analyzed PhACs weredetected in wastewater at concentrations ranging from low ng/L to a few μg/L.Of all21target PhACs,18were present in sludge and most PhACs were found at ng/g dw levels.Acetaminophen predominated in all analyzed influent samples at concentrations rangingbetween1.2-7.7μg/L, followed by sulfamethoxazole, azithromycin, roxithromycin andofloxacin
The elimination of PhACs except acetaminophen is incomplete,and carbamazepine,clofibric acid and metoprololshowed negative removal.Based on the mass balanceanalysis,biodegradation is believed to be the primary removal mechanism, and for mostof the selected pharmaceutical residues, the portion wasted with the treated sludge intothe environment was negligible. However, because all the target quinoloneantibioticscontain nitrogen as positively-charged moiety and azithromycin possessespositively charged dimethylamino group, quinolone antibiotics and azithromycin wouldhave high sorption potentials through electrostatic interactions between their positivelycharged locations and the negatively charged sludge.
Here, based on measured pharmaceutical concentrations ininfluent of the WWTPand the pharmacokinetics of PhACs in humans,we created a back-calculatedmodel.Theback-calculation can supply more consumption information and,if necessary,can work as a reference for improving current statutoryregulation on pharmaceuticalconsumption.
Based on the result of the risk assessment for the effluent, receiving water andsludge, the environment concentrations of single compounds (including sulfadiazine,sulfamethoxazole, ofloxacin, azithromycin and erythromycin-H2O) in effluent andsludge, as well as the mixture of the21detected PhACs in effluent, sludge andreceiving water had a significant ecotoxicological risk to algae. The concentration addition model was used to estimate the toxicity of the mixture of the21PhACs. Theoverall RQ(sum)for algae was more than unity in the effluent, receiving water and sludge.Thus, the mixture can lead to a high risk in the Chongqing water environment, whichmust not be ignored.
Of all21target PhACs,14PhACs were detected in source water withazithromycinhaving the highest concentration (about20ng/L). Conventional watertreatment processes cannot remove these PhACscompletely, and there were some targetPhAC were detected in treated water,11in DWTP A and6DWTP B, respectively.Theobtained results showed that the individual treatment steps in the DWTPs mostmarkedly contributing to removal of most target substances were sand filtration, and theremoval of most PhACs in the disinfection processes was negligible.
The removal of PhACs in CWS
Thesorption capacity of four frequently-used support medium (zeolite, gravel,LightExpandedClayAggregate(LECA)andquartz sand) weighed against selectedpharmaceuticals (metropolol, carbamazepine, clofibric acid, sulfamethoxazole,norfloxacin)was measured, and their physical, chemical andmineralogicalcharacterization waspursued. Gathering the information from these approaches helpedin selecting adsorbent materials for CWs—LECA. Then we built the CWS and studiedthe removal efficiencies of the selected PhACs (SMZ, OFX, ROX and CBZ) in theCWS, and the response of microbes and microphytes to the PhACs exposure in CWS atlow concentrations. Results illustrated that the CWS can effectively remove the PhACsand the removal effect of the selected PhACs in the our CWS were higher than thetraditional WWTPs. At different pharmaceutical concentration levels (0~500μg/L), thesuperoxide dismutase (SOD), catalase (CAT) and peroxidase (POD) are mutuallyinteractive to remove the reactive oxygen species (ROS), and the photosynthesis of theplant was influenced, but the influence was weakened as time went by and the plantgrowth was not influenced eventually.
Furtherinvestigationswereconductedusingphospholipidfattyacids(PLFAs)profilestofollowtheresponseofthe soilmicrobial communitywiththepurposetoillustratethe removal mechanismof PhACs in theCWS. The functional bacteria in different CWS were not changed but their orderindifferent treatment was altered. In comparison with the control (Planted (0)), some newPLFAs were found in the CWS at the concentrations of PhACsranging from10μg/L to500μg/L. The soil enzyme activities (catalase,urease and dehydrogenase activity) were promoted at concentrationsof the target PhACs between10μg/Land30μg/L, and werenot inhibited with the target pharmaceutical concentrations ranging from100μg/L to500μg/L.
①复杂基质中PhACs的检测分析方法：
利用固相萃取技术富集水环境样品中的目标PhACs，利用超声辅助萃取提取、固相萃取净化污泥介质样品中的PhACs，通过优化各种前处理条件及色谱质谱的仪器条件，建立了基于HPLC-MS/MS的检测环境中痕量的21种目标PhACs(包括氟喹诺酮类抗生素、大环内酯类抗生素、磺胺类抗生素、止痛剂、调血脂药、降高血压药、他汀类药物与抗癫痫类药等八大类药物)残留的方法平台，目标PhACs在河水、污水厂的进水、出水样品及脱水污泥样品中的回收率分别在66.4~114.9%、62.5~133.2%、64.5%~111.9%和59.5%~139.2%之间，相对标准偏差小于16%；目标PhACs在河水、污水厂的进水、出水样品及脱水污泥样品中的LOQs分别为0.03~3.4ng/L、0.2~17.5ng/L,0.2~5.6ng/L和0.2~5.8μg/kg。
②典型PhACs在水环境中的含量、消除机制及风险评估：
对重庆市四家典型污水处理厂（鸡冠石污水厂、唐家沱污水厂、李家沱污水厂和井口污水厂）各工艺阶段的进水和出水以及各污水厂初沉池和二沉池的污泥进行采样检测分析，结果表明，21种目标PhACs均能在污水样品中检测出，除吉非罗齐、对乙酰氨基酚（ACM）与布洛芬外其余18种目标PhACs也能在污泥中检测出。目标PhACs在污水中的检测浓度范围在ng/L~μg/L，在污泥中的检出浓度在μg/kg(以干重计)水平；ACM具有最高的进水浓度（1.2~7.1μg/L），其次为磺胺甲噁唑（SMZ）、阿奇霉素（AZM）、罗红霉素（ROX）、氧氟沙星（OFX），AZM在污泥中具有最高的残留浓度（549.70μg/kg）。
除ACM外，其它目标PhACs并不能在污水处理工艺中完全去除。卡马西平（CBZ）、美托洛尔（MTP）和降固醇酸（CA）甚至出现了负增长。通过质量平衡计算分析发现，微生物降解作用是目标PhACs在污水处理过程中得以去除的主要机制，大部分目标PhACs通过污泥的吸附去除的量可忽略不计。不过，由于喹诺酮类抗生素及AZM含有带正电荷的氮原子或含带正电荷的二甲氨基组，由于静电引力的作用，污泥吸附也是其从水相中去除的相关途径之一。
基于PhACs在污水处理厂进水中的浓度水平及其在人体内的代谢动力学参数，建立了PhACs的消费量反演模型。通过该模型，能从城市水循环末端推算有关PhdACs消费量的数据，可以为药物监管相关部门提供科学的数据参考。
根据检测出的PhACs环境浓度(MEC)和预测无效应浓度(PNEC)两个重要数值获得风险表征比(MEC/PNEC)的风险评价方法,进行了污水处理厂出水、受纳水体及污泥中21种PhACs的环境风险分析。结果表明磺胺嘧啶（SDZ）、SMZ、OFX与红霉素（ERY）四种抗生素在污水处理厂出水及其污泥中综合评价因子均大于1。在受纳水体中所有目标PhACs的RQ值均小于1，但SDZ、OFX、ERY及CA的RQ值大于0.1小于1，可能会对环境产生中等程度的危害，其余药物均小于0.1。21种目标PhACs在三种不同的环境下（污水厂出水、污泥及受纳水体）的RQ(sum)均大于1，因此环境中痕量PhACs的残留会对周围生态环境产生不同程度的危害。
重庆地区饮用水源水中检测除了痕量浓度（ng/L水平）的PhACs，常规处理工艺并不能完全去除原水中含有的PhACs。大多数PhACs的去除主要发生在砂滤阶段，对于砂滤池出水中含有的PhACs，加氯消毒阶段几乎无影响。
③人工湿地系统（CWS）对PhACs的去除研究：
通过对备选填料（陶粒、碎石、石英砂和沸石）物化性质及去污效果的考察，筛选出了理想的湿地填料，在此基础上以陶粒为主填料建立了潜流CWS，考察了PhACs在CWS的去除效果，从生物学和植物学角度解析PhACs胁迫下，湿地微生物和湿地植物（旱伞草）的响应。结果表明：CWS能有效的去除目标PhACs，PhACs在0~500μg/L的浓度水平下，植物体内SOD、POD和CAT能通过相互协调作用，有效地消除植物过氧化所产生的活性氧化物，防御细胞膜过氧化；在PhACs胁迫下，旱伞草叶片光合作用受到一定影响，叶片有衰老的迹象，但这种影响会随CWS运行时间的增长而降低，最终不会对植物的生长状况产生较大影响。通过对指代微生物种群的PLFAs种类进行鉴别、聚类和主成分分析发现，进水PhACs浓度在0~500μg/L下，各CWS中主要功能菌群种类未发生改变，但是其顺序会有所变化，与对照CWS相比（Planted (0)），均可检测出新的PLFAs。进水PhACs浓度在0~500μg/L水平下，CWS中基质酶活性（过氧化氢酶、脱氢酶和脲酶）未受到抑制，在10μg/L与30μg/L下，土壤基质酶活性反而升高。
Pharmaceutically active compounds (PhACs) are widely used for the treatment ofbacterial diseases in humans, and some are applied to livestock and fish for diseaseprophylaxis and treatment or for growth promotion.Large amounts of PhACs and theirmetabolites are continuously released into environmental compartments such as rivers,lakes, even ground water and soil, because all kinds of drugs were misused seriously forlack of scientific direction. Thetransportation, transformation, ecological toxicology andcontrolof PhACs have therefore provoked considerably scientific attention around theworld over the past decade. However, only a few studies about the situation in Chinahave been reported. The Chongqing region is located at the confluence of YangtzeRiver and Jialing River, water quality of drinking water source influences the safety ofdrinking water of the middle and lower Yangtze River.During the past two decades,Chongqing has become one of the fastest growing economies and most denselyurbanized areas in the world, and pharmaceutical consumption of the region is large.However, there are no studies reporting the behavior and fate of PhACs in the aquaticenvironment of Chongqing.Therefore, the occurrence, behavior and environmental fateof PhACs in the aquatic environment of this regionwere studied using ahigh-performance liquid chromatography coupled to a tandem massspectrometer.Besides, we researched the behavior and removal mechanismPhACs in theconstructed wetland systems (CWS) which are the last defenses forPhACs entering intoenvironment. The results are as follows:
The methods for determination of selected PhACs in the environment
A sensitiveanalyticalmethodwas developed fordeterminingthe selected PhACs(including several antibiotics, analgesics, antiepileptics, antilipidemics andantihypersensitives) in the water and sludge. The wastewater samples were extracted byOasis HLB cartridges (6mL,500mg),while the solid samples (sludge and suspendedsolid matter) were extracted by ultrasonic-assisted extraction with solvents followed byan enrichment and clean-up step with solid-phase extraction using HLB cartridges. Theextracted PhACs were analyzed using high-performanceliquidchromatography-electrospray ionization tandem massspectrometry(HPLC-MS/MS) with multiple reactions monitoring(MRM).The recoveries achieved forthe target PhACs ranged from66.4~114.9%for the river water, from62.5~133.2%for the influent, from64.5%~111.9%for the effluent, and from59.5%~139.2%for thesludge and their relative standard deviation was below16%. Limits of quantification(LOQs)(signal-to-noise ratio10) ranged from0.03ng/L to3.4ng/L for the surfacewater, from0.2ng/L to17.5ng/L for influent wastewater, from0.2ng/L to5.6ng/L foreffluent wastewater, and from0.17μg/kg to5.83μg/kg for sludge, respectively.
Occurrence, transportation and ecotoxicological assessmentof the selectedPhACs in the aquatic environment
Twenty-one target PhACs from8therapeutic classes covering a wide range ofphysicochemical properties and biological activities, were analyzed at four full-scalewastewater treatment plants (WWTPs) in Chongqing. All the21analyzed PhACs weredetected in wastewater at concentrations ranging from low ng/L to a few μg/L.Of all21target PhACs,18were present in sludge and most PhACs were found at ng/g dw levels.Acetaminophen predominated in all analyzed influent samples at concentrations rangingbetween1.2-7.7μg/L, followed by sulfamethoxazole, azithromycin, roxithromycin andofloxacin
The elimination of PhACs except acetaminophen is incomplete,and carbamazepine,clofibric acid and metoprololshowed negative removal.Based on the mass balanceanalysis,biodegradation is believed to be the primary removal mechanism, and for mostof the selected pharmaceutical residues, the portion wasted with the treated sludge intothe environment was negligible. However, because all the target quinoloneantibioticscontain nitrogen as positively-charged moiety and azithromycin possessespositively charged dimethylamino group, quinolone antibiotics and azithromycin wouldhave high sorption potentials through electrostatic interactions between their positivelycharged locations and the negatively charged sludge.
Here, based on measured pharmaceutical concentrations ininfluent of the WWTPand the pharmacokinetics of PhACs in humans,we created a back-calculatedmodel.Theback-calculation can supply more consumption information and,if necessary,can work as a reference for improving current statutoryregulation on pharmaceuticalconsumption.
Based on the result of the risk assessment for the effluent, receiving water andsludge, the environment concentrations of single compounds (including sulfadiazine,sulfamethoxazole, ofloxacin, azithromycin and erythromycin-H2O) in effluent andsludge, as well as the mixture of the21detected PhACs in effluent, sludge andreceiving water had a significant ecotoxicological risk to algae. The concentration addition model was used to estimate the toxicity of the mixture of the21PhACs. Theoverall RQ(sum)for algae was more than unity in the effluent, receiving water and sludge.Thus, the mixture can lead to a high risk in the Chongqing water environment, whichmust not be ignored.
Of all21target PhACs,14PhACs were detected in source water withazithromycinhaving the highest concentration (about20ng/L). Conventional watertreatment processes cannot remove these PhACscompletely, and there were some targetPhAC were detected in treated water,11in DWTP A and6DWTP B, respectively.Theobtained results showed that the individual treatment steps in the DWTPs mostmarkedly contributing to removal of most target substances were sand filtration, and theremoval of most PhACs in the disinfection processes was negligible.
The removal of PhACs in CWS
Thesorption capacity of four frequently-used support medium (zeolite, gravel,LightExpandedClayAggregate(LECA)andquartz sand) weighed against selectedpharmaceuticals (metropolol, carbamazepine, clofibric acid, sulfamethoxazole,norfloxacin)was measured, and their physical, chemical andmineralogicalcharacterization waspursued. Gathering the information from these approaches helpedin selecting adsorbent materials for CWs—LECA. Then we built the CWS and studiedthe removal efficiencies of the selected PhACs (SMZ, OFX, ROX and CBZ) in theCWS, and the response of microbes and microphytes to the PhACs exposure in CWS atlow concentrations. Results illustrated that the CWS can effectively remove the PhACsand the removal effect of the selected PhACs in the our CWS were higher than thetraditional WWTPs. At different pharmaceutical concentration levels (0~500μg/L), thesuperoxide dismutase (SOD), catalase (CAT) and peroxidase (POD) are mutuallyinteractive to remove the reactive oxygen species (ROS), and the photosynthesis of theplant was influenced, but the influence was weakened as time went by and the plantgrowth was not influenced eventually.
Furtherinvestigationswereconductedusingphospholipidfattyacids(PLFAs)profilestofollowtheresponseofthe soilmicrobial communitywiththepurposetoillustratethe removal mechanismof PhACs in theCWS. The functional bacteria in different CWS were not changed but their orderindifferent treatment was altered. In comparison with the control (Planted (0)), some newPLFAs were found in the CWS at the concentrations of PhACsranging from10μg/L to500μg/L. The soil enzyme activities (catalase,urease and dehydrogenase activity) were promoted at concentrationsof the target PhACs between10μg/Land30μg/L, and werenot inhibited with the target pharmaceutical concentrations ranging from100μg/L to500μg/L.
引文
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