黑碳对沉积物中疏水性有机物的生物富集、降解与基因毒性的作用机制
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摘要
沉积物对疏水性有机物的吸附是影响其在环境中迁移转化,生物可利用性和生态效应的主要因素之一。黑碳作为沉积物有机质中一种特殊组分,对一些疏水性有机物(如多环芳烃,多氯联苯等)有很强的吸附能力,其吸附能力可达沉积物中其他形式有机质吸附能力的10-1000倍。因此研究黑碳对沉积物中疏水性有机物的生物可利用性(如生物富集,微生物降解,亚急性毒性)的作用机制有助于准确评价疏水性污染物的生态风险,为沉积物环境标准的制定提供理论依据。
本研究首先采用热氧化法从杭州西湖沉积物中提取黑碳,以多环芳烃物质-菲和芘,五氯苯酚以及拟除虫菊酯农药-氯菊酯为目标污染物,采用静态吸附实验和迭代计算法估算黑碳对沉积物中疏水性有机物吸附的贡献。再选取生物碳(biochar),木碳(charcoal)和单壁碳纳米管(single-walled carbon nanotube)三种黑碳材料,从生物富集,微生物降解,亚急性毒性(基因毒性)三个方面阐述黑碳对沉积物中上述四种疏水性有机物生物可利用性的作用。通过Tenax(多孔的高聚膜材料)解吸,固相微萃取-Polydimethylsiloxane (PDMS)纤维以及彗星实验等仿生手段表征生物可利用性,并结合模型构建,多元线性回归等数学统计手段对黑碳影响生物可利用性的机制进一步探讨。主要结论如下：
采用热氧化法提取的沉积物黑碳对四种疏水性有机物的吸附能力差异明显。黑碳对菲,芘和五氯苯酚的吸附能力强于沉积物中其他形式的有机碳,KBC分别为KTOC的21.9,34.7,17.0倍。黑碳对氯菊酯的吸附能力与沉积物中其他有机碳的吸附能力类似,KBC仅为KTOC的1.2倍。随着污染物液相浓度的增加,黑碳表面吸附点位趋于饱和,其对沉积物中疏水性有机物吸附的贡献量下降。在液相浓度小于8.9μg/l和0.8μg/l时,黑碳对菲和芘的吸附贡献量高达50%,最大贡献量分别为84%和63%。黑碳对五氯苯酚的吸附呈相似趋势,最大吸附量为30%。黑碳对氯菊酯吸附的贡献量平均值为7.5±1.1%,说明黑碳对氯菊酯的吸附作用并没有明显强于沉积物中其他形式的有机碳。
添加黑碳后,菲在沉积物中的解吸和生物富集趋势被显著抑制,而对氯菊酯却无显著影响。Tenax解吸实验表明添加黑碳可以显著减慢菲在沉积物中的解吸速率,解吸312小时后可被解吸的菲由未添加黑碳时的67.4%减低至28.6%,由三项动力学模型拟合得到的快速解吸组分(Frapid)也由未添加黑碳时的0.265减少至0.131。添加黑碳对氯菊酯的解吸速率并无显著影响。添加黑碳后,菲在底栖生物-摇蚊体内的生物富集趋势被显著抑制,较未添加黑碳时,生物富集系数下降了72%,而添加黑碳对氯菊酯在摇蚊体内的生物富集并无显著影响。通过构建模型和多元线性回归分析表明除快速解吸组分以外,存在于慢速解吸组分中的部分化合物也可被摇蚊富集,也说明与黑碳结合的化合物可被摇蚊主动吸收富集。
沉积物中添加黑碳后,菲的微生物降解速率被减慢。其中新型黑碳材料-单壁碳纳米管(SWCNT)对菲的微生物降解的抑制作用强于传统型黑碳-生物碳和木碳。沉积物中添加生物碳和木碳后,固相微萃取法-PDMS纤维检测到的菲的自由溶解态浓度(Cfree)相对未添加组下降了10%-60%,添加SWCNT后,Cfree下降85-95%。说明黑碳对菲的强吸附能力降低菲的液相浓度,进而减缓微生物降解速率；并且SWCNT对菲的吸附能力强于生物碳和木碳。解吸-微生物降解耦合模型拟合结果表明降解菌可以直接降解与SWCNT结合的菲,这主要是因为SWCNT较生物碳和木碳有更大的表面积,表面可以吸附更多的降解菌。而且SWCNT颗粒较小,较易跨越降解菌的细胞膜,有助于降解菌直接利用与SWCNT结合的菲。SWCNT吸附可溶解性有机质后,表面积和微孔孔容下降,表面极性官能团增加,SWCNT对菲的吸附能力减弱,因此对菲的微生物降解速率的抑制作用减弱。
沉积物不同粒径大小的各组分(沙粒,淤泥,粘土)中,黑碳也是决定疏水性有机物分布,微生物降解速率的主要因素。芘在沉积物各粒径组分上的分布与各组分中的黑碳含量,总有机碳含量显著正相关。芘在各个粒径组分上的微生物降解速率与黑碳含量,总有机碳含量,颗粒表面积均显著负相关。通过Tenax解吸-微生物降解耦合模型分析发现,降解菌可以直接降解和淤泥,粘土结合的芘,主要是因为淤泥和粘土组分中分布了较多的芘,降解菌因化学趋向性而较易富集在颗粒表面,从而直接降解结合态的芘。并且该两种组分的表面积较大,可吸附较多的降解菌,促进降解菌直接利用与固体颗粒结合的芘。
沉积物中添加黑碳类物质-生物碳后,彗星实验结果表明芘和五氯苯酚对赤子爱胜蚓的基因毒性显著减弱。生物碳浓度从0%增加至5%时,芘／五氯苯酚的基因毒性分别减弱50%和80%。当生物碳含量增加为10%时,芘／五氯苯酚的基因毒性较5%时有所增加。赤子爱胜蚓活体暴露于添加生物碳的沉积物(未添加污染物)结果表明,含有10%生物碳的沉积物也可诱导显著的DNA损伤,说明生物碳自身也可能具有基因毒性,但仍需进一步实验研究证实。
Sorption of hydrophobic organic compounds (HOCs) by sediment plays an important role in their transportation, bioavailability and ecological effects. Black carbon, a special component in sediment organic matter (SOM), is considered as supersorbent for HOCs, and the sorptive capacity of black carbon could be 10-1000 times higher than those of other forms of SOM. Therefore, the study about the effect of black carbon on bioavailability (e.g. bioaccumulation, biodegradation, and subleathal toxicity) of HOCs in sediment will help us to make an accurate prediction for HOCs risk assessment as well as reasonable sediment quality criteria.
In the first part of this research, black carbon was isolated by thermal oxidation method from West Lake sediment, and four typical HOCs, including two PAHs (phenanthrene (PHE) and pyrene (PYE)), pentachlorophenol (PCP), and one pyrethroid (permethrin (PM)), were selected as the target contaminants. Equilibrium batch sorption experiment and a three-step iterative calculation were used to evaluate the contribution from black carbon to the sorption of HOCs by sediment. In the second part of this research, the effect of black carbon (biochar, charcoal, and single-walled carbon nanotube) on the bioavailability of HOCs has been investigated from three aspects of bioaccumulation, biodegradation, and sublethal toxicity (genotoxicity). Some mimic methods, i.e. Tenax desorption, solid phase micro-extraction (SPME)-disposal Polydimethylsiloxane (PDMS) fiber, and comet assay, were employed to quantify the bioavailability. Some models also have been modified and some statistical methods (such as multivariable linear regression) were adopted, which would provide further understanding about the underlying mechanism.
The sorption capacity of black carbon toward PHE, PYE, and PCP was much stronger than those of other forms of sediment organic carbon, and the values of KBC were 21.9,34.7, and 17.0 times of KTOC values. The sorption capacity of black carbon toward PM was comparable to that of other forms of sediment organic carbon, and the value of KBC was only 1.2 times of KTOC value. As the surface sorption sites of black carbon became saturated, the contribution from black carbon to the sorption of HOCs in sediment decreased with the increase of aqueous concentrations of sorbates. When the aqueous concentrations of PHE and PYE were less than 8.9μg/l and 0.8μg/l, the contribution from black carbon to sorption were higher than 50%, with the highest values of 84% and 63%, respectively. Sorption toward PCP showed a similar trend with the highest contribution of 30%. The averaged contribution from black carbon to PM sorption was 7.5±1.1%, indicating the sorption capacity of black carbon toward PM was not so significant as that toward PAH compounds.
The desorption rate of PHE in the Tenax desorption test was significantly reduced when the sediment was amended with black carbon. The desorbed fraction of PHE decreased from 67.4% to 28.6% after black carbon amendment, and the rapid desorption pool (Frapid) derived from three-phase kinetic model also dropped from 0.265 to 0.131. In contrast, there is no significant effect on PM desorption rate after black carbon addition. The bioaccumulation of PHE in Chironomus tentans was decreased after black carbon addition, and the biota-sediment accumulation factor (BSAF) decreased 72%, but there is no significant influence observed on the PM bioaccumulation. A multivariate linear regression model was developed, and model analysis result showed that, besides chemicals (PHE and PM) in the rapid desorption pool, part of chemicals in slow desorption pool were also available to Chironomus tentans, which meant chemicals, even associated with black carbon particles, could also be accumulated by Chironomus tentans.
The biodegradation of PHE by Mycobacterium vanbaalenii PYR-1 was inhibited by addition of black carbons (single-walled carbon nanotube (SWCNT), biochar, and charcoal), and the inhibitory effect of SWCNT was stronger than those of biochar and charcoal. The inhibitory effect of black carbons observed was partly due to their suppression on the bioavailability of PHE, as revealed by measurement of the freely dissolved concentration (Cfree) in the sediment using solid-phase microextraction (disposal PDMS fiber). The Cfree dropped 10%-60% after the addition of biochar and charcoal, and about 85%-95% decrease was observed when SWCNT was amended into sediment. A desorption-biodegradation coupled model was used to investigate the microbial availability of black carbon-associated PHE. Model analysis showed that the PHE sorbed with SWCNT could be directly utilized by PYR-1, which could be attributed to the relatively larger surface area and smaller particle size of SWCNT. The surface area and micropore volume of SWCNT were decreased when coated with dissolved organic matters, which also introduced polar fuctional groups onto SWCNT surface, resulting in the reduction of sorption capacity toward PHE, and the inhibitory effect on the biodegradation rate of PHE.
On the sediment particle-size scale (sand, silt, and clay), the distributions of PYE and biodegradation rates of PYE were mainly driven by black carbon contents. The distributions of PYE were significantly positively correlated with black carbon and total organic carbon content in various sized fractions. The biodegradation rate of PYE were significantly negatively correlated with black carbon content, total organic carbon content, and surface areas of various sized fractions. A biodegradation model was modified by imbedding a two-phase desorption relationship describing sequential Tenax extractions. Model analysis showed that PYE sorbed on silt and clay aggregates was directly utilized by the degrading bacteria. The enhanced bioavailability was attributed to the higher PYE concentration, or larger surface area in the silt and clay fractions, which appeared to overcome the reduced bioavailability of PYE due to sorption, making PYE on the silt and clay particles readily available to degrading microbes.
The genotoxicity induced by PYE and PCP in Eisenia fetida, which was quatified by comet assay, was inhibited after addition of black carbon-biochar. The genotoxicity of PYE and PCP was decreased 50% and 80%, respectively, when the biochar content increased from 0% to 5%. However, the 10% biochar treatment had a higher genotoxic effect than that in the 5% biochar-added exposure. This may have been caused by the genotoxic compounds and high alkalinity contained in the biochar, which means that the biochar probably could induce genotoxic effect.
本研究首先采用热氧化法从杭州西湖沉积物中提取黑碳,以多环芳烃物质-菲和芘,五氯苯酚以及拟除虫菊酯农药-氯菊酯为目标污染物,采用静态吸附实验和迭代计算法估算黑碳对沉积物中疏水性有机物吸附的贡献。再选取生物碳(biochar),木碳(charcoal)和单壁碳纳米管(single-walled carbon nanotube)三种黑碳材料,从生物富集,微生物降解,亚急性毒性(基因毒性)三个方面阐述黑碳对沉积物中上述四种疏水性有机物生物可利用性的作用。通过Tenax(多孔的高聚膜材料)解吸,固相微萃取-Polydimethylsiloxane (PDMS)纤维以及彗星实验等仿生手段表征生物可利用性,并结合模型构建,多元线性回归等数学统计手段对黑碳影响生物可利用性的机制进一步探讨。主要结论如下：
采用热氧化法提取的沉积物黑碳对四种疏水性有机物的吸附能力差异明显。黑碳对菲,芘和五氯苯酚的吸附能力强于沉积物中其他形式的有机碳,KBC分别为KTOC的21.9,34.7,17.0倍。黑碳对氯菊酯的吸附能力与沉积物中其他有机碳的吸附能力类似,KBC仅为KTOC的1.2倍。随着污染物液相浓度的增加,黑碳表面吸附点位趋于饱和,其对沉积物中疏水性有机物吸附的贡献量下降。在液相浓度小于8.9μg/l和0.8μg/l时,黑碳对菲和芘的吸附贡献量高达50%,最大贡献量分别为84%和63%。黑碳对五氯苯酚的吸附呈相似趋势,最大吸附量为30%。黑碳对氯菊酯吸附的贡献量平均值为7.5±1.1%,说明黑碳对氯菊酯的吸附作用并没有明显强于沉积物中其他形式的有机碳。
添加黑碳后,菲在沉积物中的解吸和生物富集趋势被显著抑制,而对氯菊酯却无显著影响。Tenax解吸实验表明添加黑碳可以显著减慢菲在沉积物中的解吸速率,解吸312小时后可被解吸的菲由未添加黑碳时的67.4%减低至28.6%,由三项动力学模型拟合得到的快速解吸组分(Frapid)也由未添加黑碳时的0.265减少至0.131。添加黑碳对氯菊酯的解吸速率并无显著影响。添加黑碳后,菲在底栖生物-摇蚊体内的生物富集趋势被显著抑制,较未添加黑碳时,生物富集系数下降了72%,而添加黑碳对氯菊酯在摇蚊体内的生物富集并无显著影响。通过构建模型和多元线性回归分析表明除快速解吸组分以外,存在于慢速解吸组分中的部分化合物也可被摇蚊富集,也说明与黑碳结合的化合物可被摇蚊主动吸收富集。
沉积物中添加黑碳后,菲的微生物降解速率被减慢。其中新型黑碳材料-单壁碳纳米管(SWCNT)对菲的微生物降解的抑制作用强于传统型黑碳-生物碳和木碳。沉积物中添加生物碳和木碳后,固相微萃取法-PDMS纤维检测到的菲的自由溶解态浓度(Cfree)相对未添加组下降了10%-60%,添加SWCNT后,Cfree下降85-95%。说明黑碳对菲的强吸附能力降低菲的液相浓度,进而减缓微生物降解速率；并且SWCNT对菲的吸附能力强于生物碳和木碳。解吸-微生物降解耦合模型拟合结果表明降解菌可以直接降解与SWCNT结合的菲,这主要是因为SWCNT较生物碳和木碳有更大的表面积,表面可以吸附更多的降解菌。而且SWCNT颗粒较小,较易跨越降解菌的细胞膜,有助于降解菌直接利用与SWCNT结合的菲。SWCNT吸附可溶解性有机质后,表面积和微孔孔容下降,表面极性官能团增加,SWCNT对菲的吸附能力减弱,因此对菲的微生物降解速率的抑制作用减弱。
沉积物不同粒径大小的各组分(沙粒,淤泥,粘土)中,黑碳也是决定疏水性有机物分布,微生物降解速率的主要因素。芘在沉积物各粒径组分上的分布与各组分中的黑碳含量,总有机碳含量显著正相关。芘在各个粒径组分上的微生物降解速率与黑碳含量,总有机碳含量,颗粒表面积均显著负相关。通过Tenax解吸-微生物降解耦合模型分析发现,降解菌可以直接降解和淤泥,粘土结合的芘,主要是因为淤泥和粘土组分中分布了较多的芘,降解菌因化学趋向性而较易富集在颗粒表面,从而直接降解结合态的芘。并且该两种组分的表面积较大,可吸附较多的降解菌,促进降解菌直接利用与固体颗粒结合的芘。
沉积物中添加黑碳类物质-生物碳后,彗星实验结果表明芘和五氯苯酚对赤子爱胜蚓的基因毒性显著减弱。生物碳浓度从0%增加至5%时,芘／五氯苯酚的基因毒性分别减弱50%和80%。当生物碳含量增加为10%时,芘／五氯苯酚的基因毒性较5%时有所增加。赤子爱胜蚓活体暴露于添加生物碳的沉积物(未添加污染物)结果表明,含有10%生物碳的沉积物也可诱导显著的DNA损伤,说明生物碳自身也可能具有基因毒性,但仍需进一步实验研究证实。
Sorption of hydrophobic organic compounds (HOCs) by sediment plays an important role in their transportation, bioavailability and ecological effects. Black carbon, a special component in sediment organic matter (SOM), is considered as supersorbent for HOCs, and the sorptive capacity of black carbon could be 10-1000 times higher than those of other forms of SOM. Therefore, the study about the effect of black carbon on bioavailability (e.g. bioaccumulation, biodegradation, and subleathal toxicity) of HOCs in sediment will help us to make an accurate prediction for HOCs risk assessment as well as reasonable sediment quality criteria.
In the first part of this research, black carbon was isolated by thermal oxidation method from West Lake sediment, and four typical HOCs, including two PAHs (phenanthrene (PHE) and pyrene (PYE)), pentachlorophenol (PCP), and one pyrethroid (permethrin (PM)), were selected as the target contaminants. Equilibrium batch sorption experiment and a three-step iterative calculation were used to evaluate the contribution from black carbon to the sorption of HOCs by sediment. In the second part of this research, the effect of black carbon (biochar, charcoal, and single-walled carbon nanotube) on the bioavailability of HOCs has been investigated from three aspects of bioaccumulation, biodegradation, and sublethal toxicity (genotoxicity). Some mimic methods, i.e. Tenax desorption, solid phase micro-extraction (SPME)-disposal Polydimethylsiloxane (PDMS) fiber, and comet assay, were employed to quantify the bioavailability. Some models also have been modified and some statistical methods (such as multivariable linear regression) were adopted, which would provide further understanding about the underlying mechanism.
The sorption capacity of black carbon toward PHE, PYE, and PCP was much stronger than those of other forms of sediment organic carbon, and the values of KBC were 21.9,34.7, and 17.0 times of KTOC values. The sorption capacity of black carbon toward PM was comparable to that of other forms of sediment organic carbon, and the value of KBC was only 1.2 times of KTOC value. As the surface sorption sites of black carbon became saturated, the contribution from black carbon to the sorption of HOCs in sediment decreased with the increase of aqueous concentrations of sorbates. When the aqueous concentrations of PHE and PYE were less than 8.9μg/l and 0.8μg/l, the contribution from black carbon to sorption were higher than 50%, with the highest values of 84% and 63%, respectively. Sorption toward PCP showed a similar trend with the highest contribution of 30%. The averaged contribution from black carbon to PM sorption was 7.5±1.1%, indicating the sorption capacity of black carbon toward PM was not so significant as that toward PAH compounds.
The desorption rate of PHE in the Tenax desorption test was significantly reduced when the sediment was amended with black carbon. The desorbed fraction of PHE decreased from 67.4% to 28.6% after black carbon amendment, and the rapid desorption pool (Frapid) derived from three-phase kinetic model also dropped from 0.265 to 0.131. In contrast, there is no significant effect on PM desorption rate after black carbon addition. The bioaccumulation of PHE in Chironomus tentans was decreased after black carbon addition, and the biota-sediment accumulation factor (BSAF) decreased 72%, but there is no significant influence observed on the PM bioaccumulation. A multivariate linear regression model was developed, and model analysis result showed that, besides chemicals (PHE and PM) in the rapid desorption pool, part of chemicals in slow desorption pool were also available to Chironomus tentans, which meant chemicals, even associated with black carbon particles, could also be accumulated by Chironomus tentans.
The biodegradation of PHE by Mycobacterium vanbaalenii PYR-1 was inhibited by addition of black carbons (single-walled carbon nanotube (SWCNT), biochar, and charcoal), and the inhibitory effect of SWCNT was stronger than those of biochar and charcoal. The inhibitory effect of black carbons observed was partly due to their suppression on the bioavailability of PHE, as revealed by measurement of the freely dissolved concentration (Cfree) in the sediment using solid-phase microextraction (disposal PDMS fiber). The Cfree dropped 10%-60% after the addition of biochar and charcoal, and about 85%-95% decrease was observed when SWCNT was amended into sediment. A desorption-biodegradation coupled model was used to investigate the microbial availability of black carbon-associated PHE. Model analysis showed that the PHE sorbed with SWCNT could be directly utilized by PYR-1, which could be attributed to the relatively larger surface area and smaller particle size of SWCNT. The surface area and micropore volume of SWCNT were decreased when coated with dissolved organic matters, which also introduced polar fuctional groups onto SWCNT surface, resulting in the reduction of sorption capacity toward PHE, and the inhibitory effect on the biodegradation rate of PHE.
On the sediment particle-size scale (sand, silt, and clay), the distributions of PYE and biodegradation rates of PYE were mainly driven by black carbon contents. The distributions of PYE were significantly positively correlated with black carbon and total organic carbon content in various sized fractions. The biodegradation rate of PYE were significantly negatively correlated with black carbon content, total organic carbon content, and surface areas of various sized fractions. A biodegradation model was modified by imbedding a two-phase desorption relationship describing sequential Tenax extractions. Model analysis showed that PYE sorbed on silt and clay aggregates was directly utilized by the degrading bacteria. The enhanced bioavailability was attributed to the higher PYE concentration, or larger surface area in the silt and clay fractions, which appeared to overcome the reduced bioavailability of PYE due to sorption, making PYE on the silt and clay particles readily available to degrading microbes.
The genotoxicity induced by PYE and PCP in Eisenia fetida, which was quatified by comet assay, was inhibited after addition of black carbon-biochar. The genotoxicity of PYE and PCP was decreased 50% and 80%, respectively, when the biochar content increased from 0% to 5%. However, the 10% biochar treatment had a higher genotoxic effect than that in the 5% biochar-added exposure. This may have been caused by the genotoxic compounds and high alkalinity contained in the biochar, which means that the biochar probably could induce genotoxic effect.
引文
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