Toxicity testing of four heavy metals (Cd, Cu, Mn and As) using four species of tropical marine phytoplankton, Chaetoceros calcitrans, Isochrysis galbana, Tetraselmis tetrahele and Tetraselmis sp., was carried out in multiwell plates with test volumes of 2 mL and the results compared to those of standard, large volume, shake-flasks. IC50 values (concentrations of metals estimated to inhibit 50% growth relative to the control) were determined after 96 hours based on automated O.D. readings measured in Elisa microplates by a Multiskan spectrophotometer. Good agreement was achieved between O.D. readings and cell counts indicating that this new method is a simple, economical, practical and rapid technique for toxicity testing, and provides good reproducibility of IC50 values. Results of the toxicity tests indicate that Cu was the most toxic metal (average IC50 values ranging from 0.04 to 0.37 mg L(-1)), followed by Cd (0.06-5.7 mg L(-1)), Mn (7.2-21.4 mg L(-1)) and As (33.9-319.3 mg L(-1)). Test species had different degrees of sensitivity to the metals tested, with I. galbana and C. calcitrans the most sensitive to Cu, Cd and Mn. Based on these findings it is recommended that the existing Malaysian Interim Standards for Marine Water Quality for Cd and Cu be reviewed.
Studies on toxicities and tolerances of cadmium (Cd), copper (Cu), lead (Pb) and zinc (Zn) in the brown alga Isochrysis galbana and in the green-lipped mussel Perna viridis were conducted by short-term bioassays using endpoints growth production and mortality, respectively. The 5-day EC(50) and 24-h LC(50) of these heavy metals were determined in the brown alga and mussel, respectively. The EC(50) values calculated for the alga were 0.74 mg/l for Cd, 0.91 mg/l for Cu, 1.40 mg/l for Pb and 0.60 mg/l for Zn. The LC(50) values for the mussels were 1.53 mg/l for Cd, 0.25 mg/l for Cu, 4.12 mg/l for Pb and 3.20 mg/l for Zn. These LC(50) values were within the concentration ranges as reported by other authors who used P. viridis as the test organism. Based on these EC(50) and LC(50) values, the alga was most sensitive to Zn, followed by Cd, Cu and Pb while the mussel was most sensitive to Cu, followed by Cd, Zn and Pb. Differences in the trophic levels, metal handling strategies, biology and ecology of the primary producer (brown alga) and the primary consumer (mussel) are believed to be the plausible causes for the different toxicities and tolerances of the metals studied.
A study was conducted on the long term effects of nine heavy metals on the Chironomus plumosus and Culicoides furens larvae. This study tested the effect of the heavy metals on several generations of the larvae to observe the formation of increased hardiness against pollutants present within the aquatic habitat. From this study it was observed that susceptibility or sensitivity to heavy metals decreased with LC50 values becoming larger indicating a decreased toxicity level. Significant variations (p < 0.05) were observed between first generation and third generation culicoides for all metals and at all concentrations. Variations between third and fourth generation culicoides were also significantly different (p < 0.05) with the exception of chromium at 25 degrees C and nickel and lead at every temperature range group. The variation between all generations 4, 5 and 6 was found to be insignificant (p > 0.05). This would indicate that metal tolerance would have occurred in these generations and the effect of metals was less toxic to the culicoides. Generation 9 was found to have LC50 values (p > 0.05) the same as the LC50 values obtained in third generation culicoides. Thus it would appear that heavy metal resistance was developed when the organisms were exposed to prolonged exposure of the heavy metals but was lost when the organisms were bred in non-contaminated water.
The present study was undertaken to determine whether Ligularia fischeri leaf extract (LF) is efficacious against collagen-induced arthritis (CIA) in mice. DBA/1J mice were immunized with bovine type II collagen and treated with LF (100 and 200 mg/kg) for 49 days. Mice were assessed regularly for signs of arthritis and the levels of rheumatoid factor, anti-type II collagen antibody, cytokines, AST, ALT, and creatinine in serum were also examined after the animals were killed. The arthritis score and paw edema were markedly suppressed in the groups treated with LF. Moreover, levels of rheumatoid factor, anti-type II collagen antibody, tumor necrosis factor-alpha, interleukin (IL)-1, and IL-6 in sera were reduced by LF administration. These data suggest that L. fischeri might be effective for the treatment of inflammatory arthritis like human rheumatoid arthritis.
Malaysian locally processed raw food products are widely used as main ingredients in local cooking. Previous studies showed that these food products have a positive correlation with the incidence of cancer. The cytotoxicity effect was evaluated using MTT assay (3-(4,5-dimetil-2-thiazolil)-2,5-diphenyl-2H-tetrazolium bromide) against Chang liver cells at 2000 microg/ml following 72 h incubation. Findings showed all methanol extracts caused a tremendous drop in the percentage of cell viability at 2000 microg/ml (shrimp paste - 41.69+/-3.36%, salted fish - 37.2+/-1.06%, dried shrimp - 40.32+/-1.8%, p<0.05). To detect DNA damage in a single cell, alkaline Comet Assay was used. None of the extracts caused DNA damage to the Chang liver cells at 62.5 microg/ml following 24 h incubation, as compared to the positive control, hydrogen peroxide (tail moment - 9.50+/-1.50; tail intensity - 30.50+/-2.50). Proximate analysis which was used for the evaluation of macronutrients in food showed that shrimp paste did not comply with the protein requirement (<25%) as in Food Act 1983. Salt was found in every sample with the highest percentage being detected in shrimp paste which exceeded 20%. Following heavy metal analysis (arsenic, cadmium, lead and mercury), arsenic was found in every sample with dried shrimps showing the highest value as compared to the other samples (6.16 mg/kg). In conclusion, several food extracts showed cytotoxic effect but did not cause DNA damage against Chang liver cells. Salt was found as the main additive and arsenic was present in every sample, which could be the probable cause of the toxicity effects observed.
A study was conducted to determine the suitability of using selected aquatic dipterian larvae for biomonitoring bioassays. The organisms included a member of the biting midge family that was identified as Culicoides furens and a member of the non-biting midge family, identified as Chironomus plumosus. Median lethal toxicity tests were conducted to observe the variation between metal sensitivities between the two larval forms and how variations in temperature could affect the experimental setup. Nine heavy metals were used in the study. It was observed that the 96 h LC(50) (in mg/L) for the different metals was found to be Zn-16.21 (18.55 +/- 13.87); Cr-0.96 (1.08 +/- 0.84); Ag-4.22 (6.87 +/- 1.57); Ni-0.42 (0.59 +/- 0.25); Hg-0.42 (0.59 +/- 0.25); Pb-16.21 (18.31 +/- 14.11); Cu-42.24 (45.18 +/- 39.30); Mn-4.22 (7.19 +/- 1.25); Cd-0.42 (0.59 +/- 0.25) for the Chironomus plumosus and Zn-4.22 (6.56 +/- 1.88); Cr-0.42 (0.54 +/- 0.30); Ag-0.42 (0.54 +/- 0.30); Ni-0.42 (0.54 +/- 0.30); Hg-0.04 (0.07 +/- 0.01); Pb-0.42 (0.54 +/- 0.30); Cu-42.24 (45.18 +/- 39.30); Mn-4.22 (6.56 +/- 1.88); Cd-0.42 (0.54 +/- 0.30) in the case of the Culicoides furens. With temperature as a variable the LC(50) values were observed to increase from 2.51 mg/L at 10 degrees C to 4.22 ppm at 30 degrees C and to reduce slightly to 3.72 mg/L at 35 degrees C as seen in the case of Zn. It was also observed that at 40 degrees C thermal toxicity and chemical toxicity overlapped as 100% mortality was observed in the controls. This trend was observed in all metals for both C. plumosus and C. furens. Thus indicating temperature played an important role in determining LC(50) values of toxicants.
In this study, the kinetics of adsorption of Pb(II) from aqueous solution onto palm shell-based activated carbon (PSAC) were investigated by employing ion selective electrode (ISE) for real-time Pb(II) and pH monitoring. Usage of ISE was very appropriate for real-time adsorption kinetics data collection as it facilitated recording of adsorption data at very specific and short time intervals as well as provided consistent kinetics data. Parameters studied were initial Pb(II) concentration and agitation speed. It was found that increases in initial Pb(II) concentration and agitation speed resulted in higher initial rate of adsorption. Pseudo first-order, pseudo second-order, Elovich, intraparticle diffusion and liquid film diffusion models were used to fit the adsorption kinetics data. It was suggested that chemisorption was the rate-controlling step for adsorption of Pb(II) onto PSAC since the adsorption kinetics data fitted both the pseudo second-order and Elovich models well.
In this study on green turtles, Chelonia mydas, from Peninsular Malaysia, the effect of selected environmental toxicants was examined in vitro. Emphasis was placed on purported hormone-mimicking chemicals such as dichlorodiphenyltrichloroethane (DDT), dichlorodiphenyldichloroethylene, dieldrin, lead, zinc and copper. Five concentrations were used: high (1 mg/L), medium (10(-1) mg/L), low (10(-2) mg/L), very low (10(-6) mg/L) and control (diluted carrier solvent but no toxicants). The results suggest that environmental pesticides and heavy metals may significantly alter the binding of steroids [i.e. testosterone (T) and oestradiol] to the plasma proteins in vitro. Competition studies showed that only Cu competed for binding sites with testosterone in the plasma collected from nesting C. mydas. Dieldrin and all heavy metals competed with oestradiol for binding sites. Furthermore, testosterone binding affinity was affected at various DDT concentrations and was hypothesised that DDT in vivo may act to inhibit steroid-protein interactions in nesting C. mydas. Although the precise molecular mechanism is yet to be described, DDT could have an effect upon the protein conformation thus affecting T binding (e.g. the T binding site on the steroid hormone binding protein molecule).
The 1,053-bp promoter of the oil palm metallothionein gene (so-called MSP1) and its 5' deletions were fused to the GUS reporter gene, and analysed in transiently transformed oil palm tissues. The full length promoter showed sevenfold higher activity in the mesocarp than in leaves and 1.5-fold more activity than the CaMV35S promoter in the mesocarp. The 1,053-bp region containing the 5' untranslated region (UTR) gave the highest activity in the mesocarp, while the 148-bp region was required for minimal promoter activity. Two positive regulatory regions were identified at nucleotides (nt) -953 to -619 and -420 to -256 regions. Fine-tune deletion of the -619 to -420 nt region led to the identification of a 21-bp negative regulatory sequence in the -598 to -577 nt region, which is involved in mesocarp-specific expression. Gel mobility shift assay revealed a strong interaction of the leaf nuclear extract with the 21-bp region. An AGTTAGG core-sequence within this region was identified as a novel negative regulatory element controlling fruit-specificity of the MSP1 promoter. Abscisic acid (ABA) and copper (Cu(2+)) induced the activity of the promoter and its 5' deletions more effectively than methyl jasmonate (MeJa) and ethylene. In the mesocarp, the full length promoter showed stronger inducibility in response to ABA and Cu(2+) than its 5' deletions, while in leaves, the -420 nt fragment was the most inducible by ABA and Cu(2+). These results suggest that the MSP1 promoter and its regulatory regions are potentially useful for engineering fruit-specific and inducible gene expression in oil palm.
Adult Macrobrachium lanchesteri were exposed for a 4-day period in laboratory conditions to a range of copper (Cu), cadmium (Cd), zinc (Zn) and lead (Pb) concentrations. Mortality was assessed and median lethal times (LT₅₀) and concentrations (LC₅₀) were calculated. At the end of the 4-day period, live prawns were used to determine bioconcentration of the metals. LT₅₀ and LC₅₀ increased with the decrease in mean exposure concentrations and times, respectively, for all metals. LC₅₀s for 96 hours for Cu, Cd, Zn and Pb were 32.3, 7.0, 525.1 and 35.0 µg/L, respectively. Cu, Cd, Zn and Pb bioconcentration in M. lanchesteri increases with exposure to increasing concentrations and Cd was the most toxic to M. lanchesteri, followed by Pb, Cu and Zn. Comparison of LC₅₀ values for metals for this species with those for other freshwater crustacean organisms reveals that M. lanchesteri is equally or more sensitive to heavy metals than most other tested crustaceans.
The genotoxic effects of increasing concentrations (below lethal concentration [LC₅₀]) of cadmium ([Cd] 0.1, 1 and 10 mg/L), copper ([Cu] 0.2, 2 and 20 mg/L) and zinc ([Zn] 0.5, 5 and 50 mg/L) on Chironomus kiiensis were evaluated using alkaline comet assay after exposure for 24 h. Both the tail moment and the olive tail moment showed significant differences between the control and different concentrations of Cd, Cu and Zn (Kruskal-Wallis, p < 0.05). The highest concentration of Cd was associated with higher DNA damage to C. kiiensis larvae compared with Cu and Zn. The potential genotoxicity of these metals to C. kiiensis was Cd > Cu > Zn.
As well as for chemical and environmental reasons, acrylamide is widely used in many industrial applications. Due to its carcinogenicity and toxicity, its discharge into the environment causes adverse effects on humans and ecology alike. In this study, a novel acrylamide-degrading yeast has been isolated. The isolate was identified as Rhodotorula sp. strain MBH23 using ITS rRNA analysis. The results showed that the best carbon source for growth was glucose at 1.0% (w/v). The optimum acrylamide concentration, being a nitrogen source for cellular growth, was at 500 mg l(-1). The highest tolerable concentration of acrylamide was 1500 mg l(-1) whereas growth was completely inhibited at 2000 mg l(-1). At 500 mg l(-1), the strain MBH completely degraded acrylamide on day 5. Acrylic acid as a metabolite was detected in the media. Strain MBH23 grew well between pH 6.0 and 8.0 and between 27 and 30 °C. Amides such as 2-chloroacetamide, methacrylamide, nicotinamide, acrylamide, acetamide, and propionamide supported growth. Toxic heavy metals such as mercury, chromium, and cadmium inhibited growth on acrylamide.
Cupriavidus sp. strain BIS7 is a Malaysian tropical soil bacterium that exhibits broad heavy-metal resistance [Co(II), Zn(II), Ni(II), Se(IV), Cu(II), chromate, Co(III), Fe(II), and Fe(III)]. It is particularly resistant to Fe(II), Fe(III), and Zn(II). Here we present the assembly and annotation of its genome.
Freshwater quality criteria for copper (Cu), cadmium (Cd), aluminum (Al), and manganese (Mn) were developed with particular reference to aquatic biota in Malaysia, and based on USEPA's guidelines. Acute toxicity tests were performed on eight different freshwater domestic species in Malaysia, which were Macrobrachiumlanchesteri (prawn), two fish -Poeciliareticulata and Rasborasumatrana, Melanoidestuberculata (snail), Stenocyprismajor (ostracod), Chironomusjavanus (midge larvae), Naiselinguis (annelid), and Duttaphrynusmelanostictus (tadpole), to determine 96-h LC50 values for Cu, Cd, Al, and Mn. The final acute values (FAVs) for Cu, Cd, Al, and Mn were 2.5, 3.0, 977.8, and 78.3 μgL(-1), respectively. Using an estimated acute-to-chronic ratio (ACR) of 8.3, the value for final chronic value (FCV) was derived. Based on FAV and FCV, a Criterion Maximum Concentration (CMC) and a criterion Continuous Concentration (CCC) for Cu, Cd, Al, and Mn of 1.3, 1.5, 488.9, and 39.1 μgL(-1) and 0.3, 0.36, 117.8, and 9.4 μgL(-1), respectively, were derived. The results of this study provide useful data for deriving national or local water quality criteria for Cu, Cd, Al, and Mn based on aquatic biota in Malaysia. Based on LC50 values, this study indicated that R.sumatrana, M.lanchesteri, C.javanus, and N.elinguis were the most sensitive to Cu, Cd, Al, and Mn, respectively.
Pollutants put great stress on the environment, especially the aquatic ecosystem; therefore, the ease with which pollutants migrate in water is a subject of global concern. In this study, leachate from landfill that was analyzed with the objective of understanding the potential impact to the environment was tested on Pangasius sutchi. Heavy metals available at various concentrations in raw leachate samples of both closed and active landfills necessitated the determination of their degree of bioaccumulation in this fish species in order to enrich the risk data on toxicity of effluents. Zinc (3.2 µg g(-1)), iron (2.1 µg g(-1)) and chromium (0.24 µg g(-1)) detected in the fish within 96 h of acute exposure is of concern. A histopathology test on excised liver of P. sutchi indicated cellular disruption from normal stain. Heterogeneous effluents like leachate may affect not only groundwater but can endanger aquatic ecosystems, especially in some regions where improper waste disposal and treatment allow the flow of leachate into surface water courses. Though metals might be beneficial to organisms, the extent at which they can accumulate in leachate-exposed fish is a risk and can initiate metal toxicity in aquatic life.
Whole cell biosensors always face the challenge of low stability of biological components and short storage life. This paper reports the effects of poly(2-hydroxyethyl methacrylate) (pHEMA) immobilization on a whole cell fluorescence biosensor for the detection of heavy metals (Cu, Pb, Cd), and pesticides (dichlorophenoxyacetic acid (2,4-D), and chlorpyrifos). The biosensor was produced by entrapping the cyanobacterium Anabaena torulosa on a cellulose membrane, followed by applying a layer of pHEMA, and attaching it to a well. The well was then fixed to an optical probe which was connected to a fluorescence spectrophotometer and an electronic reader. The optimization of the biosensor using several factors such as amount of HEMA and drying temperature were undertaken. The detection limits of biosensor without pHEMA for Cu, Cd, Pb, 2,4-D and chlorpyrifos were 1.195, 0.027, 0.0100, 0.025 and 0.025 µg/L respectively. The presence of pHEMA increased the limits of detection to 1.410, 0.250, 0.500, 0.235 and 0.117 µg/L respectively. pHEMA is known to enhance the reproducibility of the biosensor with average relative standard deviation (RSD) of ±1.76% for all the pollutants tested, 48% better than the biosensor without pHEMA (RSD = ±3.73%). In storability test with Cu 5 µg/L, the biosensor with pHEMA performed 11.5% better than the test without pHEMA on day-10 and 5.2% better on day-25. pHEMA is therefore a good candidate to be used in whole cell biosensors as it increases reproducibility and enhances biosensor storability.
Perna viridis (P. viridis) has been identified as a good biological indicator in identifying environmental pollution, especially when there are various types of Heavy Metals Accumulations (HMA) inside its tissue. Based on the potential of P. viridis to accumulate heavy metals and the data on its physical properties, this study proffers to determine the relationships between both properties. The similarities of the physical properties are used to mathematical model their relationships, which included the size (length, width, height) and weight (wet and dry) of P. viridis, whilst the heavy metals are focused on concentrations of Pb, Cu, Cr, Cd and Zn. The concentrations of metal elements are detected by using Flame Atomic Adsorption Spectrometry. Results show that the mean concentration of Pb, Cu, Cr, Cd, Zn, length, width, height, wet weight and dry weight are: 1.12 +/- 1.00, 2.36 +/- 1.65, 2.12 +/- 2.74, 0.44 +/- 0.41 and 16.52 +/- 10.64 mg kg(-1) (dry weight), 105.08 +/- 14.35, 41.64 +/- 4.64, 28.75 +/- 3.92 mm, 14.56 +/- 3.30 and 2.37 +/- 0.86 g, respectively. It is also found out that the relationships between the Heavy Metals Concentrations (HMA) and the physical properties can be represented using Multiple Linear Regressions (MLR) models, relating that the HMA of Zinc has affected significantly the physical growth properties of P. viridis.
A fluorescence-based fiber optic toxicity biosensor based on genetically modified Escherichia coli (E. coli) with green fluorescent protein (GFP) was developed for the evaluation of the toxicity of several hazardous heavy metal ions. The toxic metals include Cu(II), Cd(II), Pb(II), Zn(II), Cr(VI), Co(II), Ni(II), Ag(I) and Fe(III). The optimum fluorescence excitation and emission wavelengths of the optical biosensor were 400 ± 2 nm and 485 ± 2 nm, respectively. Based on the toxicity observed under optimal conditions, the detection limits of Cu(II), Cd(II), Pb(II), Zn(II), Cr(VI), Co(II), Ni(II), Ag(I) and Fe(III) that can be detected using the toxicity biosensor were at 0.04, 0.32, 0.46, 2.80, 100, 250, 400, 720 and 2600 μg/L, respectively. The repeatability and reproducibility of the proposed biosensor were 3.5%-4.8% RSD (relative standard deviation) and 3.6%-5.1% RSD (n = 8), respectively. The biosensor response was stable for at least five weeks, and demonstrated higher sensitivity towards metal toxicity evaluation when compared to a conventional Microtox assay.
Two freshwater fish, Rasbora sumatrana (Cyprinidae) and Poecilia reticulata (guppy; Poeciliidae), were exposed to a range of eight heavy metals (copper (Cu), cadmium (Cd), zinc (Zn), lead (Pb), nickel (Ni), iron (Fe), aluminium (Al), and manganese (Mn)) at varied concentrations for 96 h in the laboratory. Mortality was assessed and median lethal concentrations (LC50) were calculated. It was observed that the LC50 values increased with a decrease in mean exposure times, for all metals and for both fish types. The 96-h LC50 values for Cu, Cd, Zn, Pb, Ni, Fe, Al, and Mn were 0.006, 0.10, 0.46, 0.63, 0.83, 1.71, 1.53, and 5.71 mg/L for R. sumatrana and 0.038, 0.17, 1.06, 1.99, 15.62, 1.46, 6.76, and 23.91 mg/L for P. reticulata, respectively. The metal toxicity trend for R. sumatrana and P. reticulata from most to least toxic was Cu > Cd > Zn > Pb > Ni > Al > Fe > Mn and Cu > Cd > Zn > Fe > Pb > Al > Ni > Mn, respectively. Results indicated that Cu was the most toxic metal on both fish, and R. sumatrana was more sensitive than P. reticulata to all the eight metals.
Biosorption process is a promising technology for the removal of heavy metals from industrial wastes and effluents using low-cost and effective biosorbents. In the present study, adsorption of Pb(2+), Cu(2+), Fe(2+), and Zn(2+) onto dried biomass of red seaweed Kappaphycus sp. was investigated as a function of pH, contact time, initial metal ion concentration, and temperature. The experimental data were evaluated by four isotherm models (Langmuir, Freundlich, Temkin, and Dubinin-Radushkevich) and four kinetic models (pseudo-first-order, pseudo-second-order, Elovich, and intraparticle diffusion models). The adsorption process was feasible, spontaneous, and endothermic in nature. Functional groups in the biomass involved in metal adsorption process were revealed as carboxylic and sulfonic acids and sulfonate by Fourier transform infrared analysis. A total of nine error functions were applied to validate the models. We strongly suggest the analysis of error functions for validating adsorption isotherm and kinetic models using linear methods. The present work shows that the red seaweed Kappaphycus sp. can be used as a potentially low-cost biosorbent for the removal of heavy metal ions from aqueous solutions. Further study is warranted to evaluate its feasibility for the removal of heavy metals from the real environment.