好气条件下不同形态外源砷在土壤中的转化

在35%的田间持水量下,通过模拟试验研究了外源二甲基砷酸盐（DMA）、一甲基砷酸盐（MMA）、五价无机砷［As(V)］在土壤中的形态转化.结果表明: 外源砷进入土壤后,其含量均有随时间推延而逐渐下降的趋势,两种不同形态的有机砷DMA和MMA在土壤中主要发生脱甲基化过程,经150 d的恒温恒湿培养,其在土壤中主要转化为As(V),DMA处理仅在120 d时检测到少量MMA,而MMA处理则在7～60 d内均有少量的DMA生成.培养结束时土壤中DMA和MMA含量均显著降低(P<0.01),降幅分别为99.5%、94.3%,而两者的主要转化产物As(V)的含量则分别显著增加了4.61和5.15倍.表明外源有机态砷在土壤中基本上被转化为无机形态;与有机态外源砷相比,外源As(V)进入土壤后其形态基本上没有发生转化.     

外源二甲基砷在土壤中的转化

利用盆栽试验,研究了外源二甲基砷（DMA）进入土壤后的形态及其与土壤胶体结合态的变化.结果表明：土壤中的DMA主要转化为砷酸根［As(V)］,并伴有少量的一甲基砷（MMA）生成;当外源DMA添加浓度为30 mg·kg^-1时,DMA的转化率最高,其在培养时间为10、15、30、40 d时的转化率分别为6.71%、8.11%、11.33%、19.32%;随着外源DMA添加浓度的增加土壤中易溶性砷（AE-As）浓度呈逐渐增加的趋势,但随着培养时间的增加,AE-As浓度不断降低;与不加外源DMA的对照相比,添加不同量的DMA使土壤中铝型砷（Al-As）、铁型砷（Fe-As）、钙型砷（Ca-As）浓度均有所增加,DMA及其转化产物与土壤中铝、铁、钙的氧化物或氢氧化物间的吸附或固定可能是其主要原因.     

外源二甲基砷对油菜生长及土壤中砷生物有效性的影响

通过盆栽试验研究了土壤中添加外源二甲基砷(DMA)对油菜(Brassica campestris)生长及土壤中砷生物有效性的影响.结果表明:随着外源DMA添加量的增加,油菜的出苗率和生物量均在一定程度上表现出了低浓度促进而高浓度抑制的现象.当外源DMA添加量达到90 mg·kg-1时,第2季盆栽油菜的出苗率和生物量与对照相比分别下降了9.5%和57.0%,表明DMA对油菜生长的影响具有长期性;随着外源DMA浓度的增加,土壤中有效态砷及油菜体内的砷含量均表现出增加的趋势,相关性分析表明,该三者间具有极显著相关关系;添加入土壤中的DMA主要发生去甲基化作用,产物主要为As(V)及少量的As(Ⅲ),且随着外源DMA添加量的增加,As(V)和As(Ⅲ)的浓度均表现出增加的趋势.     

砷形态对黑藻和竹叶眼子菜有机酸含量的影响

采用室内溶液培养法,运用气相色谱-质谱联用(Gas chromatography-mass spectrometry,GC-MS)技术测定在As(Ⅲ)、As(Ⅴ)和二甲基胂酸(Dimethylarsinic acid,DMA)3种形态砷(浓度0—5.0 mg/L)胁迫下,砷超富集植物黑藻(Hydrilla verticillata(Linn.f.)Royle.)和非砷超富集植物竹叶眼子菜(Potamogeton malaianus Miq.)体内生成和体外分泌的7种有机酸的含量。结果表明,黑藻体内草酸、丙二酸、柠檬酸、苹果酸和棕榈酸都显著高于竹叶眼子菜(P<0.05)。在As(Ⅲ)处理时,黑藻体内柠檬酸、苹果酸、棕榈酸、亚麻酸和总有机酸显著下降,竹叶眼子菜棕榈酸显著下降;在As(Ⅴ)处理时,黑藻棕榈酸和总有机酸显著下降,竹叶眼子菜棕榈酸显著下降;在DMA处理时,黑藻草酸、柠檬酸和苹果酸显著下降,竹叶眼子菜棕榈酸、亚麻酸和总有机酸显著下降。黑藻在As(Ⅲ)处理下棕榈酸、亚麻酸、总有机酸和As(Ⅴ)处理下草酸含量与体内As含量呈显著负相关(P<0.05),而竹叶眼子菜在As(Ⅲ)或DMA处理下棕榈酸、亚麻酸、总有机酸含量以及在DMA处理下柠檬酸含量与体内As含量均呈显著负相关(P<0.05)。两种植物根系分泌物中均检测出草酸、丙二酸、琥珀酸和棕榈酸,但只有在As(Ⅲ)或As(Ⅴ)处理时黑藻分泌的草酸与体内As含量呈显著正相关。本研究表明As(Ⅲ)或As(Ⅴ)胁迫下黑藻向体外分泌草酸是其超富集砷的一种重要机制。     

淹水-控温模式下砷污染水稻土溶液中砷形态的变化

以水稻种植区的砷污染土壤为研究对象,采用高效液相色谱(HPLC)-电感耦合等离子体质谱(ICP-MS)联用分析测试系统,研究了不同培养温度(5、27和50℃)对灭菌和不灭菌的土壤淹水后其溶液中砷赋存形态变化的影响.结果表明:在土壤溶液中检测到的砷形态只有无机三价砷(AsⅢ)、无机五价砷(Asv)和有机的二甲基砷(DMAv),未检测到单甲基砷(MMAv)的存在;在不同控温条件下随淹水时间的延长,AsⅢ逐渐转变为砷的主要赋存形态,平均比例约为64％;Asv次之,约占35％,DMAv的含量相对最低,约占1％;土壤灭菌与否对土壤溶液中五价砷的水平没有明显影响,但明显影响了五价砷的还原和促进了无机三价砷的甲基化,并且灭菌的促进效果随着淹水及培养时间的延长而逐渐降低;50℃、淹水23 d时,灭菌土壤溶液中DMAv浓度最高,为23.7 ng·mL-1,这说明灭菌土壤中残留的某些嗜热微生物成为优势菌群并促进了土壤溶液中砷的甲基化.结合水稻生长的实际环境条件对该研究结果进行分析,培养温度27℃淹水23 d后不灭菌的自然土壤溶液中砷浓度处于较低水平,因此在砷污染的水稻种植区建议采用短周期干湿交替的水分管理模式,在保障产量的情况下可尽量降低土壤溶液中砷的水平.     

外源As(Ⅲ)在不同母质发育土壤中的老化过程

应用模拟试验方法,研究了外源As(Ⅲ)在第四纪红土、紫色砂页岩和花岗岩3种母质发育土壤中的老化过程,并分析了该过程中砷的有效性、结合态等的变化.结果表明: 经过120 d老化后,3种土壤中均只能检测到As(Ⅴ)的存在;伴随老化过程的进行,土壤中有效态砷的含量均呈下降趋势,且下降幅度为:紫色砂页岩发育土壤(RS₂)>花岗岩发育土壤(RS₃)>第四纪红土发育土壤(RS₁).应用准二级动力学方程可以较好地模拟3种土壤中有效态砷含量的变化(P＜0.05),土壤pH、有机质及铁铝锰氧化物是影响砷老化的主要因素,且锰氧化物的影响大于铁铝氧化物(P＜0.05).相关分析表明,非专性吸附态砷和专性吸附态砷是构成土壤有效砷的主要形态.     

单一与复合砷锑处理对波士顿蕨富集与转化砷和锑的作用

通过水培试验,研究了单一和复合砷锑处理对波士顿蕨(Nephrolepis exaltata L.)植株各部位及其亚细胞砷锑富集和形态转化的影响,以期查明植物体内砷与锑之间可能的相互作用。结果表明,与对照组相比,单一和复合处理条件下,波士顿蕨各部位砷锑含量均有显著的增加,砷含量表现为根叶柄羽叶,锑含量表现为根羽叶叶柄。与20 mg·L-1单一Sb(III)处理相比,复合处理条件下,随着砷的同时加入(10 mg·L-1As)植株各部位锑含量出现下降,同时根部细胞器中的锑出现向细胞壁与胞液转移、而羽叶与叶柄出现由细胞壁向其他两个亚细胞组分转化的现象;与10 mg·L-1单一As(III)处理相比,复合处理条件下,锑的加入(20 mg·L-1Sb)显著降低了波士顿蕨各部位砷的含量,同时促进了根部细胞壁中的砷向胞液与细胞器转移,以及羽叶与叶柄胞液处砷向细胞壁与细胞器的转移。因此,波士顿蕨体内,砷锑之间具有拮抗作用。单一As(III)处理下,波士顿蕨根部以As(V)为主、羽叶主要以DMA为主,而Sb(III)的同时加入则能够促进As(III)更多地向As(V)和DMA转化;单一Sb(III)处理下,波士顿蕨体内以Sb(III)为主导形态,随着As(III)的同时加入,在波士顿蕨根与叶柄处,As(III)强烈地抑制Sb(III)向Sb(V)转化,相反在羽叶处却会促进Sb(III)向Sb(V)转化。     

无机砷在植物体内的吸收和代谢机制

砷污染已成为全球非常突出且急需解决的环境问题,严重威胁人类健康和环境安全.在自然环境和土壤系统中,砷的存在形态相当复杂,但植物砷毒害主要源于As（V）和As（Ⅲ）暴露.As（V）通过Pi的吸收通道被植物根系吸收,并在还原酶（AR）作用下被快速还原为As（Ⅲ）.As（Ⅲ）通过NIP蛋白通道进入植物体内,在砷甲基转移酶(ArsM)的作用下转化为甲基化砷或与谷胱甘肽（GSH）、植物螯合肽（PC）等多肽的巯基螯合封存在根部液泡或转运到地上部分,从而起到砷解毒的作用.同时,植物吸收的一部分砷也可外排到外部介质.本文以农作物尤其是水稻为主线,详述了As（V）和As（Ⅲ）吸收、外排及As（V）还原、As（Ⅲ）甲基化、螯合作用的最新研究进展,并提出了今后的研究重点.     

紫色非硫细菌的砷代谢机制

[目的]系统阐述紫色非硫细菌(PNSB)砷代谢机制和砷代谢基因簇的进化关系.[方法]通过生物信息学方法分析了PNSB砷代谢基因簇的分布、组成、排布方式.采用UV-Vis和HPLC-ICP-MS方法,研究了3个PNSB种类对砷的抗性、砷形态及价态的转化、砷在细胞中的积累和分布以及磷酸盐对As细胞毒性的影响.[结果]砷基因簇分析表明:已公布全基因组序列的17个PNSB菌株基因组中均含有以ars operon为核心的砷代谢基因簇,由1-4个操纵子组成,主要含有与细胞质砷还原和砷甲基化代谢相关的基因,但基因的组成和排列方式因种和菌株而异,尤其是arsM和两类进化来源不同的arsC.实验结果表明:光照厌氧条件下,3个PNSB种类对As(V)和As(Ⅲ)均具有抗性,As(V)和As(Ⅲ)均能进入细胞 ;在胞内As(V)能够还原为As(Ⅲ)并被排出胞外,但不能将As(Ⅲ)氧化为As(V),也未检测到甲基砷化物 ;磷酸盐浓度升高,能够抑制As(V)进入细胞,降低As(V)对细胞的毒性,而不能抑制As(Ⅲ)进入细胞.[结论]PNSB砷代谢机制主体为细胞质As(V)还原,也还有砷甲基化途径.通过对砷代谢基因簇结构多样性特点和进化方式分析,提出了与Rosen不同的ars operon进化途径.这对深入开展PNSB砷代谢和基因之间的相互作用研究奠定基础.     

超富集植物蜈蚣草中砷化学形态的EXAFS研究

采用同步辐射扩展X射线吸收精细结构(SREXAFS)技术研究了超富集植物蜈蚣草(PterisvittataL.)中As的化学形态及其在转运过程中的变化。结果表明,蜈蚣草中的As主要以As(Ⅲ)与O配位的形态存在。As(V)被植物吸收后,很快转化为As(Ⅲ),其转化过程主要发生在根部。As(Ⅲ)向地上部转运的过程中价态基本不变。在植物的根部和部分叶柄中存在少量与As-GSH相似的As-S结合方式,但是在As含量最高的羽叶中基本上未发现这种结合方式。与需要提取和分离过程的化学方法相比,采用EXAFS方法研究植物中的砷形态不需经过预分离或化学预处理就可以直接测定植物样品中元素的化学形态,因此可以避免样品预处理过程对As形态的干扰,并获得可靠的砷化学形态方面的信息。     

控制性分根区交替滴灌对苹果幼树形态特征与根系水分传导的影响

采用固定滴灌(根区一侧固定供水)、控制性分根区交替滴灌(根区两侧交替供水)和常规滴灌(紧贴幼树基部供水)3种灌水方式和3种灌水定额(固定滴灌和交替滴灌均为10、20和30 mm,常规滴灌为20、30和40 mm),对比研究了控制性分根区交替滴灌对苹果幼树形态特征与根系水分传导的影响.结果表明： 交替滴灌的根区两侧土壤出现反复干湿交替过程,常规滴灌的根区两侧土壤含水率差异不显著.在灌水定额相同时,灌水侧的土壤含水率在3种灌水方式间差异不显著.与常规滴灌和固定滴灌相比,交替滴灌显著增加了苹果幼树的根冠比、壮苗指数和根系水分传导,在30 mm灌水定额处理下,交替滴灌的根冠比分别增加31.6%和47.1%,壮苗指数增加34.2%和53.6%,根系水分传导增加9.0%和11.0%.3种灌水方式下,根干质量和叶面积均与根系水分传导呈显著线性正相关.控制性分根区交替滴灌增强了苹果幼树根系水分传导的补偿效应,促进了根系对水分的吸收利用,有利于干物质向各个器官均衡分配,显著提高了根冠比和壮苗指数.     

A simple and fast detection technique for arsenic speciation based on high‐efficiency photooxidation and gas‐phase chemiluminescence detection

High‐efficiency photooxidation (HEPO) and gas phase chemiluminescence detection (CL) combined with high‐performance liquid chromatography (HPLC) and hydride generation were developed for speciation of As(III), As(V), monomethylarsonic acid (MMA) and dimethylarsinic acid (DMA). After chromatography separation, the arsenic species were passed through HEPO which performed efficient photooxidation and converted MMA and DMA to As(V) in several seconds. Then the reaction of ozone and arsine upon hydride generation produced a CL signal as the analytical parameter. The total analytical process was completed within 10 min. The effects of operational parameters such as the concentrations of hydrochloric acid and NaBH₄ solution, carrier gas flow and air gas flow for ozone generation were investigated. Detection limits were 3.7, 10.3, 10.2 and 10.0 µg/L for As(III), As(V), MMA and DMA, respectively. The recoveries of the four arsenic species in human urine sample ranged from 87 to 94%. Copyright © 2009 John Wiley & Sons, Ltd.     

Arsenic speciation in turnip as affected by application of chicken manure bearing roxarsone and its metabolites

Roxarsone (ROX) is widely used as a feed additive in intensive animal production. While animals are fed with ROX, the most commonly detectable As forms in fresh manures include ROX and small quantities of its metabolites such as arsenate (As(V)), arsenite (As(III)), monomethylarsonic acid (MMA) and dimethylarsinic acid (DMA). A pot experiment was conducted to investigate the uptake, translocation and distribution of ROX, As(V), As(III), MMA and DMA in turnips, with the soil amended with 2% and 3% (w/w) chicken manure (CM) bearing ROX and its metabolites. Soil without any fertilizer was the control. The results show that only As(V) and As(III) were detected in turnip control samples. As(V), As(III) and DMA were found in all CM applied samples, but not ROX or MMA. This implies that turnip cannot take up ROX directly and accumulate MMA at detectable levels. The contents of DMA in tubers and the three As species in shoots increased with the CM rate in contrast to reduced levels of As(V) and As(III) in tubers. Increased CM rate enhanced the translocation of the three As species, especially for DMA, from tubers to shoots. DMA was the major form (42.9–61.4% in tubers and 38.1–76.3% in shoots), followed by As(III), in turnip plants fertilized with CM. The results indicate that ROX and its metabolites in animal manures can be introduced into human food chain by the way ROX → animal → manure → soil → crop.     

Methylated arsenic species in plants originate from soil microorganisms

? Inorganic arsenic (iAs) is a ubiquitous human carcinogen, and rice (Oryza sativa) is the main contributor to iAs in the diet. Methylated pentavalent As species are less toxic and are routinely found in plants; however, it is currently unknown whether plants are able to methylate As. ? Rice, tomato (Solanum lycopersicum) and red clover (Trifolium pratense) were exposed to iAs, monomethylarsonic acid (MMA(V)), or dimethylarsinic acid (DMA(V)), under axenic conditions. Rice seedlings were also grown in two soils under nonsterile flooded conditions, and rice plants exposed to arsenite or DMA(V) were grown to maturity in nonsterile hydroponic culture. Arsenic speciation in samples was determined by HPLC-ICP-MS. ? Methylated arsenicals were not found in the three plant species exposed to iAs under axenic conditions. Axenically grown rice was able to take up MMA(V) or DMA(V), and reduce MMA(V) to MMA(III) but not convert it to DMA(V). Methylated As was detected in the shoots of soil-grown rice, and in rice grain from nonsterile hydroponic culture. GeoChip analysis of microbial genes in a Bangladeshi paddy soil showed the presence of the microbial As methyltransferase gene arsM. ? Our results suggest that plants are unable to methylate iAs, and instead take up methylated As produced by microorganisms.     

淹水-控温模式下砷污染水稻土溶液中砷形态的变化

以水稻种植区的砷污染土壤为研究对象,采用高效液相色谱(HPLC)-电感耦合等离子体质谱(ICP-MS)联用分析测试系统,研究了不同培养温度(5、27和50 ℃)对灭菌和不灭菌的土壤淹水后其溶液中砷赋存形态变化的影响.结果表明： 在土壤溶液中检测到的砷形态只有无机三价砷(As^Ⅲ)、无机五价砷(As^V)和有机的二甲基砷(DMA^V),未检测到单甲基砷(MMA^V)的存在;在不同控温条件下随淹水时间的延长,As^Ⅲ逐渐转变为砷的主要赋存形态,平均比例约为64%;As^V次之,约占35%,DMA^V的含量相对最低,约占1%;土壤灭菌与否对土壤溶液中五价砷的水平没有明显影响,但明显影响了五价砷的还原和促进了无机三价砷的甲基化,并且灭菌的促进效果随着淹水及培养时间的延长而逐渐降低;50 ℃、淹水23 d时,灭菌土壤溶液中DMA^V浓度最高,为23.7 ng·mL^-1,这说明灭菌土壤中残留的某些嗜热微生物成为优势菌群并促进了土壤溶液中砷的甲基化.结合水稻生长的实际环境条件对该研究结果进行分析,培养温度27 ℃淹水23 d后不灭菌的自然土壤溶液中砷浓度处于较低水平,因此在砷污染的水稻种植区建议采用短周期干湿交替的水分管理模式,在保障产量的情况下可尽量降低土壤溶液中砷的水平.     

Exceptions in patterns of arsenic compounds in urine of acute promyelocytic leukaemia patients treated with As<Subscript>2</Subscript>O<Subscript>3</Subscript>

Arsenic trioxide (As(III) in solution) has been shown to be the most active single agent in combating acute promyelocytic leukemia (APL). It is metabolized and excreted via urine as monomethylarsonic acid (MMA), dimethylarsinic acid (DMA) and As(V), along with excess As(III). In our study eight APL patients were treated (intravenously) with 0.15 mg As₂O₃/kg/day. During the therapy As(III) and its metabolites were followed in pre- and post-infusion urine using HPLC for separation followed by on-line detection using hydride generation-atomic fluorescence spectrometry. Five patients had a normal excretion pattern of residual arsenic compounds in morning pre-infusion urine, with 15–25 % of As(III), 35–55 % of DMA, 25–30 % of MMA and 1–5 % of As(V), while three patients showed unexpected exceptions from typical excretion patterns of arsenic compounds (i) a high DMA/MMA ratio (factor 5.3), (ii) severe As(III) oxidation (10.2 % As(III) converted to As(V)) or (iii) the presence of an excessive amount of As(III) (average 30.4 % of total arsenic). Intriguing was the occurrence of post-infusion oxidation of As(III) to As(V) observed in almost all patients and being especially high (>40 %) in patient with increased residual As(V). Results indicate that arsenic metabolites patterns can be unpredictable. Observed high levels of un-metabolised As(III) are a warning signal for side effects and for routine determination of arsenic metabolites during first days of treatment. High or low percentages of MMA or DMA did not show any observable effect on treatment results, while clear presence of post-infusion As(V) supports theoretical claims of in vivo oxidation (detoxification) of As(III) to As(V) associated with various metabolic processes.     

Trivalent arsenicals induce lipid peroxidation, protein carbonylation, and oxidative DNA damage in human urothelial cells

Drinking arsenic-contaminated water is associated with an increased risk of bladder cancer. Arsenate (iAs(V)), arsenite (iAs(III)), monomethylarsonous acid (MMA(III)), monomethylarsonic acid (MMA(V)), dimethylarsinous acid (DMA(III)), and dimethylarsinic acid (DMA(V)) have all been detected in the urine of people who drink arsenic-contaminated water. The aim of this research was to investigate which of these arsenicals are more hazardous to human urothelial cells. The results indicate that iAs(III), MMA(III), and DMA(III) were more potent in inducing cytotoxicity, lipid peroxidation, protein carbonylation, oxidative DNA damage, nitric oxide, superoxide, hydrogen peroxide, and cellular free iron than MMA(V), DMA(V), and iAs(V) in human urothelial carcinoma and transformed cells. However, the results did not show convincingly that the trivalent arsenicals were more potent than pentavalent arsenicals in decreasing the intracellular contents of total thiol, protein thiol, and reduced glutathione. Induction of oxidative DNA damage was observed with 0.2 microM of iAs(III), MMA(III), or DMA(III) as early as 1h. Because of its high oxidative damage, higher proportion in urine, and lower cytotoxicity, DMA(III) may be the most hazardous arsenical to human urothelial cells.     

Alternative splicing variants of human arsenic (+3 oxidation state) methyltransferase

Arsenic (+3 oxidation state) methyltransferase (As3MT) catalyzes the methylation of trivalent arsenic (As(III)) to monomethylarsonate (MMA(V)) and dimethylarsinic acid (DMA(V)), and plays an important role in the detoxification of arsenicals. Here, we report the identification of two splicing variants of the human As3MT gene. One splicing variant was an exon-3 skipping (Δ3) form which produced a premature stop codon, and the other was an exon-4 and -5 skipping (Δ4,5) form which produced a 31.1 kDa As3MT protein. In addition to the full-length mRNA of As3MT, Δ4,5 mRNAs were detected in HepG2, A549, HL60, K562, and HEK293 cells. The methyltransferase activity of the recombinant Δ4,5 As3MT and wild-type As3MT proteins purified from Escherichia coli was determined. Speciation analysis by HPLC–ICP-MS showed a clear peak of MMA(V) after incubation of As(III) with the wild-type As3MT protein, but not with the Δ4,5 As3MT protein. In addition, COS-7 cells transfected with Δ4,5 As3MT cDNA did not convert As(III) to MMA(V) or DMA(V). The lack of methyltransferase activity of Δ4,5 As3MT seems to be related to the deletion of an S-adenosylmethionine-binding site and a critical cysteine residue. These data suggest that the expression pattern of splicing variants of the As3MT gene may affect the capacity for arsenic methylation in cells.