Biotic ligand model, a flexible tool for developing site-specific water quality guidelines for metals

The biotic ligand model (BLM) is a mechanistic approach that greatly improves our ability to generate site-specific ambient water quality criteria (AWQC)for metals in the natural environment relative to conventional relationships based only on hardness. The model is flexible; all aspects of water chemistry that affect toxicity can be included, so the BLM integrates the concept of bioavailability into AWQC--in essence the computational equivalent of water effect ratio (WER) testing. The theory of the BLM evolved from the gill surface interaction model (GSIM) and the free ion activity model (FIAM). Using an equilibrium geochemical modeling framework, the BLM incorporates the competition of the free metal ion with other naturally occurring cations (e.g., Ca2+, Na+, Mg2-, H+), togetherwith complexation by abiotic ligands [e.g., DOM (dissolved organic matter), chloride, carbonates, sulfide] for binding with the biotic ligand, the site of toxic action on the organism. On the basis of fish gill research, the biotic ligands appear to be active ion uptake pathways (e.g., Na+ transporters for copper and silver, Ca2+ transporters for zinc, cadmium, lead, and cobalt), whose geochemical characteristics (affinity = log K, capacity = Bmax) can be quantified in short-term (3-24 h) in vivo gill binding tests. In general, the greater the toxicity of a particular metal, the higher the log K. The BLM quantitatively relates short-term binding to acute toxicity, with the LA50 (lethal accumulation) being predictive of the LC50 (generally 96 h for fish, 48 h for daphnids). We critically evaluate currently available BLMs for copper, silver, zinc, and nickel and gill binding approaches for cadmium, lead, and cobalt on which BLMs could be based. Most BLMs originate from tests with fish and have been recalibrated for more sensitive daphnids by adjustment of LA50 so as to fit the results of toxicity testing. Issues of concern include the arbitrary nature of LA50 adjustments; possible mechanistic differences between daphnids and fish that may alter log K values, particularly for hardness cations (Ca2+, Mg2+); assumption of fixed biotic ligand characteristics in the face of evidence that they may change in response to acclimation and diet; difficulties in dealing with DOM and incorporating its heterogeneity into the modeling framework; and the paucity of validation exercises on natural water data sets. Important needs include characterization of biotic ligand properties at the molecular level; development of in vitro BLMs, extension of the BLM approach to a wider range of organisms, to the estuarine and marine environment, and to deal with metal mixtures; and further development of BLM frameworks to predict chronic toxicity and thereby generate chronic AWQC.     

Development of a biotic ligand model and a regression model predicting acute copper toxicity to the earthworm Aporrectodea caliginosa

The purpose of this study was to develop a terrestrial biotic ligand model (BLM) for predicting acute copper toxicity to the earthworm Aporrectodea caliginosa. To overcome the basic problems hampering development of BLMs for terrestrial organisms, an artificial flow-through exposure system was developed consisting of an inert quartz sand matrix and a nutrient solution, of which the composition was univariately modified. A. caliginosa was exposed for 7 days under varying concentrations of copper and the major cations modifying toxicity: H+, Ca2+, Mg2+, and Na+. In addition copper speciation was modulated by means of EDTA or dissolved organic carbon (DOC). An increase in pH or pNa resulted in a linear decrease of 7-days median lethal concentrations. Increasing Ca2+ and Mg2+ activities had inconsistent effects. EDTA addition decreased toxicity when the total copper concentration in the pore water was kept the same. This is attributed to the strong complexation capacity of EDTA and shows that total copper is not the toxic species. DOC was more protective than could be explained by its metal complexing properties. The BLM developed incorporates the effects of H+ and Na+. This BLM was validated with the results of a set of bioassays with artificial pore water in quartz sand and by a set of bioassays in spiked field soils. Prediction error was within a factor of 2, but some predictions were not within the 95% confidence interval. Therefore a more widely applicable regression type model was developed that was able to explain >95% of the (lack of) toxicity observed. To our knowledge this is the first report of the successful development of a terrestrial BLM.     

Bioavailability and chronic toxicity of zinc to juvenile rainbow trout (Oncorhynchus mykiss): comparison with other fish species and development of a biotic ligand model

In this study, the effects of modifying Ca (0.2-4 mM), Mg (0.05-3 mM), Na (0.75-5 mM), and pH (5.5-7.5) on the chronic toxicity of zinc to juvenile rainbow trout (Oncorhynchus mykiss) were investigated using standard 30-d assays in which survival and growth were monitored. Survival was observed to be a more sensitive end point than growth, and mortality mainly occurred during the initial stages of the exposure. This suggested that the mode of action of zinc toxicity was mainly of an acute nature. A review and analysis of existing literature demonstrated similar results for most other fish species investigated. Overall, up to a 30-fold variation of zinc toxicity was observed, as indicated by no observed effect concentrations varying between 32.7 and 974 microg of Zn L(-1). Increased concentrations of Ca2+, Mg2+, Na+, and H+ (within the tested ranges) resulted in a reduction of chronic zinc toxicity by a factor of 12, 3, >2, and 2, respectively. This suggests the major importance of Ca competing with zinc and protecting against zinc toxicity, which seems to be a ubiquitous concept in fish species (and probably also invertebrate). On the basis of the toxicity data obtained, a chronic biotic ligand model (BLM) was developed that takes into account both chemical speciation of zinc and competition between zinc and the above-mentioned cations. The developed model was able to predict chronic effect concentrations with an error of less than a factor of 2 in most cases. Hence, it was concluded that the chronic Zn BLM can reduce toxicity variability due to bioavailability to a considerable extent and that the BLM can become an important tool in criteria setting and risk assessment practice of zinc and zinc substances.     

Copper toxicity to bioluminescent Nitrosomonas europaea in soil is explained by the free metal ion activity in pore water

The terrestrial biotic ligand model (BLM) for metal toxicity in soil postulates that metal toxicity depends on the free metal ion activity in solution and on ions competing for metal sorption to the biotic ligand. Unequivocal evidence for the BLM assumptions is most difficult to obtain for native soil microorganisms because the abiotic and biotic compartments cannot be experimentally separated. Here, we report copper (Cu) toxicity to a bioluminescent Nitrosomonas europaea reporter strain that was used in a solid phase-contact assay and in corresponding soil extracts and artificial soil solutions. The Cu(2+) ion activities that halve bioluminescence (EC50) in artificial solutions ranged 10(-5) to 10(-7) M and increased with increasing activities of H(+), Ca(2+) and Mg(2+) according to the BLM concept. The solution based Cu(2+) EC50 values of N. europaea in six contaminated soils ranged 2 × 10(-6) to 2 × 10(-9) M and these thresholds for both solid phase or soil extract based assays were well predicted by the ion competition model fitted to artificial solution data. In addition, solution based Cu(2+) EC50 of the solid phase-contact assay were never smaller than corresponding values in soil extracts suggesting no additional solid phase toxic route. By restricting the analysis to the same added species, we show that the Cu(2+) in solution represents the toxic species to this bacterium.     

Ni uptake by a green alga. 2. Validation of equilibrium models for competition effects

It is generally admitted that the presence of major cations and H+ can attenuate trace metal uptake. Recent models such as the biotic ligand model (BLM) aim to quantify and predict this effect by determining stability constants for each of the major competitors for any given interaction of a trace metal with a biological organism. In this study, short-term Ni internalization fluxes (J(int)) were used to quantitatively assess the binding of H+, Mg2+, Ca2+ (K(H-Rs), K(Mg-Rs), K(Ca-Rs)), and trace metals to transport sites (R(s)) leading to Ni biouptake by Chlamydomonas reinhardtii. H+ and Mg2+ are shown to compete directly for the entry of Ni with affinity constants that are of the same order of magnitude (K(Mg-Rs) = 10(5.1) M(-1); K(H-Rs) = 10(5.3) M(-1)) as that measured for Ni (K(Ni-Rs) = 10(5.1) M(-1)). The Ni internalization fluxes were also strongly linked to the Mg cell status. In contrast, the role of Ca2+ could not be explained by a simple competitive equilibrium with the Ni transport sites. Aluminum (K(Al-Rs) = 10(8) M(-1)), Zn (K(Zn-Rs) = 10(6.5) M(-1)), and Cu (K(Cu-Rs) = 10(6.6) M(-1)) were all shown to compete strongly with Ni for uptake. In addition to the determination of uptake constants, these studies provide insight into the transport mechanisms of Ni by the green alga, C. reinhardtii.     

A terrestrial biotic ligand model. 1. Development and application to Cu and Ni toxicities to barley root elongation in soils

A Terrestrial Biotic Ligand Model (TBLM) was developed using noncalcareous soils from Europe based on Cu and Ni speciation and barley (Hordeum vulgare cv. Regina) root elongation bioassays. Free metal ion (M2+) activity was computed by the WHAM VI model using inputs of soil metal, soil organic matter, and alkali and alkaline earth metals concentrations, and pH in soil solution. The TBLM assumes that metal in soil and in the solution are in equilibrium. Metal ions react with the biotic ligand, the receptor site, and inhibit root elongation. Other ions, principally H+, Ca2+ and Mg2+, compete with M2+ and, therefore, affect its toxicity. Toxicity is correlated only to the fraction of the total biotic ligand sites occupied by M2+. Compared to other models using either the soil metal concentration or M2+ activity as the toxic dose, the TBLM provides a more consistent method to normalize and compare Cu and Ni toxicities to root elongation among different soils. The TBLM was able to predictthe EC50 soil Cu and Ni concentrations generally within a factor of 2 of the observed values, a level of precision similar to that for the aquatic Biotic Ligand Model, indicating its potential utility in metals risk assessment in soils.     

不同金属离子对多酚氧化酶的影响

采用匀浆法从茄子中提取多酚氧化酶(PPO),以绿茶粉为底物,研究Cu2+、Mn2+、Mg2+、Ca2+、Na+、Al3+和Fe3+7种金属离子对PPO活性的影响。结果表明:Cu2+、Mn2+和Mg2+3种金属离子对PPO的活性有不同程度的促进作用,而Ca2+、Na+、Al3+和Fe3+对PPO的活性有抑制作用。其中,Cu2+浓度为3mmol/L时对PPO有激活作用,此时茶黄素总量达到7.45mg;Fe3+浓度为5mmol/L对PPO有显著抑制作用,此时茶黄素总量降低至0.26mg。     

Zinc toxicity to nitrification in soil and soilless culture can be predicted with the same biotic ligand model

The inhibitory effect of Zn on the nitrification process in ZnCl2 spiked soils (12 soils, pH range 4.8-7.5) was compared to toxic effects of Zn on the nitrification by Nitrosospira sp. in soilless solutions with varying pH (pH 6-8) and ionic composition. The nitrification was reduced by 20% at Zn solution concentrations (EC20) ranging between 7 and 1200 microM Zn in the soil pore water and between 5 and 150 microM Zn in the soilless solutions. Protective effects of H+, Ca2+, and Mg2+ against Zn2+ toxicity were observed in both systems. Zinc speciation was determined, and 60-90% of the Zn in the soils and 35-80% of the Zn in the soilless solutions was present as Zn2+. A biotic ligand model and a Freundlich-type model, incorporating the competition of Zn2+ ions with H+, Ca2+, and Mg2+ for binding on the biotic ligands, were used to model the results. The Zn2+ activities resulting in 20% reduction of the nitrification were well predicted using the same parameters for both (soil and soilless) systems, indicating that microorganisms in soil are exposed to zinc through the free zinc ion in soil pore water.     

Alleviation of Cu and Pb rhizotoxicities in cowpea (Vigna unguiculata) as related to ion activities at root-cell plasma membrane surface

Cations, such as Ca and Mg, are generally thought to alleviate toxicities of trace metals through site-specific competition (as incorporated in the biotic ligand model, BLM). Short-term experiments were conducted with cowpea (Vigna unguiculata L. Walp.) seedlings in simple nutrient solutions to examine the alleviation of Cu and Pb toxicities by Al, Ca, H, Mg, and Na. For Cu, the cations depolarized the plasma membrane (PM) and reduced the negativity of ψ(0)(o) (electrical potential at the outer surface of the PM) and thereby decreased {Cu(2+)}(0)(o) (activity of Cu(2+) at the outer surface of the PM). For Pb, root elongation was generally better correlated to the activity of Pb(2+) in the bulk solution than to {Pb(2+)}(0)(o). However, we propose that the addition of cations resulted in a decrease in {Pb(2+)}(0)(o) but a simultaneous increase in the rate of Pb uptake (due to an increase in the negativity of E(m,surf), the difference in potential between the inner and outer surfaces of the PM) thus offsetting the decrease in {Pb(2+)}(0)(o). In addition, Ca was found to alleviate Pb toxicity through a specific effect. Although our data do not preclude site-specific competition (as incorporated in the BLM), we suggest that electrostatic effects have an important role.     

茶绿色素稳定性的研究

对茶绿色素的稳定性进行研究.结果表明:荼绿色素的光谱特性在663 nm为特征吸收峰;茶绿色素对热较稳定;金属离子K+、Na+、Zn2+、Cu2+、Mg2+,Ca2+对该色素的稳定性影响较小,Al3+、Fe2+和Fe3+对其稳定性有明显的影响或破坏作用;pH对该色素的影响明显,在弱酸性条件下(pH=5～6)稳定性较好;光对该色素的稳定性影响较大;苯甲酸钠对该色素的稳定性影响较小;该色素抗氧化性较弱,抗还原性较强.     

鸭血糯色素稳定性的研究

对鸭血糯色素的稳定性进行研究。研究表明,鸭血糯色素在酸性条件下稳定性较好,而在碱性条件下稳定性极差,并且颜色发生改变;鸭血糯色素的热稳定性、光稳定性较好;糖类、食盐、柠檬酸、淀粉、山梨酸钾和苯甲酸钠对鸭血糯色素的影响很小;金属离子Mn2+、Na+、K+、Zn2+、Cu2+、Ca2+ 和Mg2+ 对鸭血糯色素的稳定性影响较小,Fe3+ 对鸭血糯色素的稳定性影响较大。     

ICP-AES分析植酸酶对制麦过程中游离态金属离子含量的变化

运用电感耦合等离子体原子发射光谱法(ICP-AES)考察了外源添加植酸酶对大麦发芽过程中游离态K+、Na+、Ca2+、Mg2+、Zn2+含量的变化。实验发现:添加外源植酸酶能提高发芽过程中游离态金属离子的含量,K+、Na+、Ca2+、Mg2+、Zn2+含量同比可提高10.7%、23.4%、16.3%、8.0%、75.5%。通过比较金属离子含量的变化可以为制定合理的制麦工艺提供参考。     

Toward a biotic ligand model for freshwater green algae: surface-bound and internal copper are better predictors of toxicity than free Cu2+-ion activity when pH is varied

The freshwater green microalgae Chlorella sp. and Pseudokirchneriella subcapitata (P. subcapitata) were chronically (48 and 72 h, respectively) exposed to copper at various pH levels, i.e., pH 6-7.5 and pH 5.9-8.5, respectively. Concentrations resulting in 50% inhibition of exponential growth rate (EC50) were determined as dissolved Cu, estimated chemical activity of the free Cu2+ ion (as pCu = - log{Cu2+ activity as molarity}), and as external (surface-bound) Cu and internal Cu in the algal cells. With increasing pH, EC50dissolved decreased from 30 to 1.1 microg of Cu L(-1) for Chlorella sp. and from 46 to 18 microg of Cu L(-1) for P. subcapitata. The pH effect on copper toxicity was even more obvious when expressed as Cu2+ activity. The EC50pCu increased on average 1.4 pCu unit per pH unit for Chlorella sp. and 1.1 pCu unit per pH unit for P. subcapitata, thus indicating a marked increase of Cu2+ toxicity at higher pH (more than 1 order of magnitude per pH unit). In contrast, it was found that EC50 values expressed as surface bound or external copper (EC50external) and as internal copper (EC50internal) did not vary substantially when pH was increased. External Cu was operationally defined as the Cu fraction removable from the algal cell by short-term contact with ethylenediaminetetraacetic acid; internal copper was defined as the nonremovable fraction. For Chlorella sp. the EC50external varied between 5 and 10 fg of Cu/ cell (factor of 2 difference) and the EC50internal between 25 and 40 fg of Cu/cell (factor of 1.6 difference). For P. subcapitata the EC50external varied between 10 and 28 fg of Cu/cell (factor of 2.8 difference) and the EC50internal between 42 and 71 fg of Cu/cell (factor of 1.7 difference). Because the observed variation in EC50external and EC50internal is much less than the variation in EC50Cu2+, it is concluded that both external and internal copper are better predictors of copper toxicity than Cu2+ when pH is varied. From the perspective of toxicity modeling, this observation is the first step toward considering the use of the cell surface as the algal biotic ligand for Cu in a similar way as fish gills fulfill this role in the biotic ligand model for predicting metal toxicity to fish species.     

自然密滤在天然山泉水生产中的应用

本文以天然山泉水为研究水体,以自然密滤技术对山泉水进行了加工处理,研究了自然密滤工艺对山泉水COD值、铁、锰的去除效果,以及钾、钠、钙、镁离子的保留情况.结果证明自然密滤工艺对COD值具有良好的去除效应,有效去除了水体中的天然有机物;该工艺对铁、锰的去除效果良好,保证了天然山泉水清甜可口的口感;同时,该工艺对人体所需的...     

Using the biotic ligand model for predicting the acute sensitivity of cladoceran dominated communities to copper in natural surface waters

In this study, the acute copper sensitivity of field-collected cladoceran species was determined using their natural surface waters and a standard reconstituted test water as test medium. A total of 43 species were collected on two occasions from six different sites, representing different water types and chemistries in Europe. The collected species belonged to four different families (Daphniidae, Bosminidae, Macrothricidae, Chydoridae) and 11 different genera (Daphnia, Ctenodaphnia, Ceriodaphnia, Simocephalus, Scapholeberis, Alona, Acroperus, Chydorus, Eurycercus, Disparalona, Pleuroxus). In acute experiments with immobilization as end point, the 48-h median effective concentrations (48-h EC50) for the cladoceran species ranged from 5.30 to 70.6 microg of Cu L(-1) in standard test water and from 9.60 to 853 microg of Cu L(-1) in natural waters. The mean site sensitivity (the geometric mean of 48-h EC50 values of species within a community) ranged from 10.1 to 27.4 microg of Cu L(-1) in standard water and from 16.4 to 281 microg of Cu L(-1) in natural water. This indicates that bioavailability is more importantthan inter-community (species composition) differences in determining the variability of copper toxicity across different aquatic systems. For the four surface waters that had a pH within the range for which the acute Daphnia magna biotic ligand model (BLM) has previously been successfully validated, the BLM predicted 48-h EC50 values for 27 of the 28 tested cladoceran species within factor of 2 of the observed values. For the same sites, all community sensitivities were predicted within a factor of 2.3. The BLM was clearly over-protective for the two acidic surface waters tested. Hence, the BLM can be considered a valuable tool for estimating the potentially harmful effects of copperto natural cladoceran communities, but more research will be needed for acidic surface waters.     

镰刀霉菌(Fusarium sp)JN158紫色素的提取及理化性质

研究了由镰刀霉菌(Fusarium sp)JN158发酵所产紫色素的提取条件和理化性质,结果表明:该紫色素的提取溶剂为体积分数95%乙醇溶液;紫色素在酸性条件下对温度,光照,柠檬酸和苯甲酸钠有较好的稳定性;最大吸收波长为310 nm和510 am;对H2O2和Na2SO3敏感;蔗糖与葡萄糖有增色效果;金属离子Zn2+、...     

猪苓色素的提取工艺及稳定性研究

通过正交试验确定猪苓色素提取的最佳工艺条件,研究色素的溶解性、热稳定性、光稳定性以及氧化剂、pH值与金属离子对色素的影响。结果表明,料液比1:60时,以1mol/LNaOH溶液为提取剂、提取温度95℃、时间90min是猪苓色素的最佳提取工艺;干燥的猪苓色素为深褐色粉末,能溶于水,易溶于碱性溶液,热稳定性好,光照会加速其分解,对H2O2不稳定,溶液pH值5～10范围时色素表现稳定;Na+、K+、Ca2+、Mg2+、Zn2+等离子不影响色素的稳定性,而Mn2+、Fe2+、Fe3+及Cu2+则有影响,其中Fe3+和Cu2+影响尤为明显。     

红曲色素TY的稳定性研究

红曲色素是一种天然的安全的食用色素,在食品行业应用广泛。本文主要研究了光照、温度、pH、金属离子(Cu2+、Ca2+、Mg2+、K+和Na+)以及食品添加剂(酸度调节剂、甜味剂、防腐剂和抗氧化剂)对红曲色素稳定性的影响。结果表明,红曲色素TY溶液在室外光照能使红曲色素溶液褪色,在室内光照和避光条件下对溶液色价影响不明显;在高温下具有良好的热稳定性;在pH<4时溶液产生沉淀,且色泽变为暗淡,pH>5时溶液无沉淀现象且色泽明亮;Fe2+和Cu2+的存在能使溶液产生沉淀,其他金属离子影响小;酸度调节剂(柠檬酸、乙酸、苹果酸、乳酸、酒石酸)能使溶液产生沉淀并且溶液色泽变的较暗淡。     

金属离子对蜡状芽孢杆菌合成多聚γ-谷氨酸的影响

研究金属离子对蜡状芽孢杆菌合成多聚γ-谷氨酸(γ-PGA)的影响。通过在培养基中加入定量的Mn2+、K+、Fe3+、Mg2+、Ca2+、Mo6+、Zn2+、Co2+、Cd2+、Cu2+,定时取发酵液用原子吸收分光光度法测定蜡状芽孢杆菌合成γ-PGA过程中金属离子的消耗情况、菌体生物量及γ-PGA产量;在发酵培养基中,分别加入不同质量浓度的Mn2+、Ca2+、Zn2+、Co2+、Mo6+,通过组合不同金属离子以及控制各组合中离子质量浓度测定菌体生物量和γ-PGA含量的变化,结合金属离子的消耗量探讨金属离子对蜡状芽孢杆菌合成γ-PGA的影响。结果表明：在蜡状芽孢杆菌合成γ-PGA的过程中均消耗一定量的Mn2+、K+、Fe3+、Mg2+、Ca2+、Mo6+、Zn2+、Co2+,其中Mn2+、K+、Fe3+、Mg2+、Ca2+在生长代谢过程中利用较多,而Cd2+、Cu2+的质量浓度不变化;Mn2+是γ-PGA合成的必需元素,在其质量浓度为0.08g/L时,γ-PGA合成达最高峰(1.25g/L)。当Mn2+质量浓度为0.08g/L时,适当增加Ca2+质量浓度就能较大幅度提高γ-PGA合成量;同时固定Mn2+、Ca2+质量浓度,适量加入不同质量浓度的Zn2+、使γ-PGA的合成量增加明显,最高可达2.94g/L,而Mo6+对γ-PGA的合成影响不明显,但对菌体生物量的影响较大。     

托柄菝葜果色素提取及其性质研究

以民族药托柄菝葜的成熟果实为材料,对其红色素的提取条件及理化性质进行研究.结果表明,托柄菝葜果色素提取的最佳条件为:40％vol乙醇水溶液、浸提温度60℃、浸提时间lh.托柄菝葜果色素在自然光照下稳定,但添加金属离子的色素溶液对光不稳定;色素溶液在酸性环境中呈现鲜艳的橙红色,在中性及碱性环境中变为褐绿色.VC对托柄菝葜色素具有增色作用;柠檬酸、EDTA对色素无明显影响;亚硫酸钠、高锰酸钾及苯甲酸钠对色素具有减色作用.金属离子Ca2+、Mn2+、Zn2+、K+、Na+、Fe2+、Mg2+、Cu2+、Co2+、Al3+、Pb2+等离子能减弱托柄菝葜果色素对光的稳定性;Sn2+、Fe3+影响色素稳定性.