Xingqiang Wu

An effective pathway for the removal of microcystin LR via anoxic biodegradation in lake sediments

Aerobic biodegradation has been considered to be the main attenuation mechanism for microcystins, but the role of anoxic biodegradation remains unclear. We investigated the potential for anoxic biodegradation of microcystin and the effects of environmental factors on the process through a series of well-controlled microcosm experiments using lake sediments as inocula. Microcystin LR could be degraded anoxically from 5mgL(-1) to below the detection limit at 25 degrees C within 2 days after a lag phase of 2 days. The rate was highly dependent on temperature, with a favorable temperature range of 20-30 degrees C. The addition of glucose or low levels of NH(4)-N had no effect on the anoxic biodegradation of microcystin, whereas the addition of NO(3)-N significantly inhibited the biodegradation at all experimental concentrations, and the inhibition increased with increasing amount of NO(3)-N-amended. Adda (3-amino-9-methoxy-2,6,8-trimethyl-10-phenyl-deca-4,6-dienoic acid), a previously reported nontoxic product of aerobic degradation of microcystin, was identified as the anoxic biodegradation product. This is the first report of Adda as a degradation product of microcystin under anoxic conditions. No other product containing Adda residue was detected during the anoxic degradation of microcystin. These results strongly indicated that anoxic biodegradation is an effective removal pathway of microcystin in lake sediments, and represents a significant bioremediation potential.

Mechanisms and Factors Affecting Sorption of Microcystins onto Natural Sediments

The sorption of microcystins (MCs) to fifteen lake sediments and four clay minerals was studied as a function of sediment/clay properties, temperature, and pH through well-controlled batch sorption experiments. All sorption data for both sediments and clays are well described by a nonlinear Freundlich model (n(f) varies between 0.49 and 1.03). The sorption process for MCs exhibited different adsorptive mechanisms in different lake sediments mainly dependent on the sediment organic matter (OM). For sediments with lower OM (i.e., less than 8%), the sorption of MCs decreases with increasing OM and is dominated by the competition for adsorption sites between MCs and OM. In contrast, MC sorption to organic-rich (i.e., more than 8%) sediments increases with increasing OM and is dominated by the interaction between MCs and adsorbed OM. The sorption thermodynamics of MCs onto sediments showed that MC sorption is a spontaneous physisorption process with two different mechanisms. One mechanism is an exothermic process for sediment with lower OM, and the other is an endothermic process for sediment with higher OM. Furthermore, the sorption of MCs onto sediments is pH dependent (sorption decreased with increasing pH). These results provide valuable informations for a better understanding of the natural abiotic attenuation mechanisms for MCs in aquatic ecosystems.

Effects of microcystin-LR, linear alkylbenzene sulfonate and their mixture on lettuce (Lactuca sativa L.) seeds and seedlings

Microcystin-LR (MCLR) and linear alkylbenzene sulfonate (LAS) are present widely in aquatic and terrestrial ecosystems, but their combined ecotoxicological risk is unknown. This study investigated the toxic effects of MCLR, LAS and their mixture on lettuce (Lactuca sativa L.) and evaluated MCLR accumulation level in lettuce with or without LAS. The changes in seed germination and shoot/root growth, responses of the antioxidative defense system, and the accumulation of MCLR in lettuce were tested to evaluate the single and combined toxic effect of MCLR and LAS in well-controlled conditions. The results showed that seedling growth (except for root elongation and leaf weight) was more sensitive to toxicant exposure than seed germination. For seedling leaves, lipid peroxidation was not observed when the antioxidative defense system (including superoxide dismutase, catalase and glutathione) was activated to relieve the adverse effects of oxidative stress via different pathways. Our results also confirmed that the interaction between MCLR and LAS was synergistic. Both toxicants in combination not only significantly inhibited seedling growth, but also increased the activities of superoxide dismutase and catalase, as well as the contents of glutathione. Furthermore, LAS dramatically enhanced the accumulation of MCLR in the plant, thus leading to a reduction in quality and yield and posing greater potential risk to humans via consumption of these edible plants.

Kinetic study of the 2-methyl-3-methoxy-4-phenylbutanoic acid produced by oxidation of microcystin in aqueous solutions†

Microcystins (MCs) are a family of related cyclic hepatotoxic heptapeptides, of which more than 70 types have been identified. The chemically unique nature of the C20 beta-amino acid, (2S,3S,8S,9S)-3-amino-9-methoxy-2,6,8-trimethyl-10-phenyldeca-4,6-dienoic acid (Adda), portion of the MCs has been exploited to develop a strategy to analyze the entirety. Oxidation of MCs causes the cleavage of MC Adda to form 2-methyl-3-methoxy-4-phenylbutanoic acid (MMPB). In the present study, we investigated the kinetics of MMPB produced by oxidation of the most-often-studied MC variant, MC-LR (L = leucine, R = arginine), with permanganate-periodate. This investigation allowed insight regarding the influence of the reaction conditions (concentration of the reactants, temperature, and pH) on the conversion rate. The results indicated that the reaction was second order overall and first order with respect to both permanganate and MC-LR. The second-order rate constant ranged from 0.66 to 1.35 M/s at temperatures from 10 to 30 degrees C, and the activation energy was 24.44 kJ/mol. The rates of MMPB production can be accelerated through increasing reaction temperature and oxidant concentration, and sufficient periodate is necessary for the formation of MMPB. The initial reaction rate under alkaline and neutral conditions is higher than that under acidic conditions, but the former decreases faster than the latter except under weakly acidic conditions. These results provided new insight concerning selection of the permanganate-periodate concentration, pH, and temperature needed for the oxidation of MCs with a high and stable yield of MMPB.

Effects of arsenate on microcystin content and leakage of Microcystis strain PCC7806 under various phosphate regimes.

Both arsenic pollution and eutrophication are prominent environmental issues when considering the problem of global water pollution. It is important to reveal the effects of arsenic species on cyanobacterial growth and toxin yields to assess ecological risk of arsenic pollution or at least understand naturally occurring blooms. The sensitivity of cyanobacteria to arsenate has often been linked to the structural similarities of arsenate and phosphate. Thus, we approached the effect of arsenate with concentrations from 10−8 to 10−4 M on Microcystis strain PCC7806 under various phosphate regimes. The present study showed that Microcystis strain PCC7806 was arsenate tolerant up to 10−4 M. And such tolerance was without reference to both content of intra‐ and extra‐cellular phosphate. It seems that arsenate involved the regulation of microcystin synthesis and cellular polyphosphate contributed to microcystin production of Microcystis responding to arsenate, since there was a positive linear correlation of the cellular microcystin quota with the exposure concentration of arsenate when the cells were not preconditioned to phosphate starvation. It is presumed that arsenate could help to actively export microcystins from living Microcystis cells when preconditioned to phosphate starvation and incubated with the medium containing 1 μM phosphate. This study firstly provided evidence that microcystin content and/or release of Microcystis might be impacted by arsenate if it exists in harmful algal blooms.

Rapid quantification of total microcystins in cyanobacterial samples by periodate-permanganate oxidation and reversed-phase liquid chromatography

Microcystins (MCs) comprise a family of more than 80 related cyclic hepatotoxic heptapeptides. Oxidation of MCs causes cleavage of the chemically unique C20 beta-amino acid (2S, 3S, 8S, 9S)-3-amino-9-methoxy-2,6,8-trimethyl-10-phenyldeca-4,6-dienoic acid (Adda) amino to form 2-methyl-3-methoxy-4-phenylbutanoic acid (MMPB), which has been exploited to enable analysis of the entire family. In the present study, the reaction conditions (e.g. concentration of the reactants, temperature and pH) used in the production of MMPB by oxidation of cyanobacterial samples with permanganate-periodate were optimized through a series of well-controlled batch experiments. The oxidation product (MMPB) was then directly analyzed by high-performance liquid chromatography with diode array detection. The results of this study provided insight into the influence of reaction conditions on the yield of MMPB. Specifically, the optimal conditions, including a high dose of permanganate (> or = 50 mM) in saturated periodate solution at ambient temperature under alkaline conditions (pH approximately 9) over 1-4 h were proposed, as indicated by a MMPB yield of greater than 85%. The technique developed here was applied to determine the total concentration of MCs in cyanobacterial bloom samples, and indicated that the MMPB technique was a highly sensitive and accurate method of quantifying total MCs. Additionally, these results will aid in development of a highly effective analytical method for detection of MMPB as an oxidation product for evaluation of total MCs in a wide range of environmental sample matrices, including natural waters, soils (sediments) and animal tissues.

Optimal strategies for determination of free/extractable and total microcystins in lake sediment

The optimization of analytical procedures for the quantification of free and total microcystins (MCs) in natural sediments was systematically examined based on solvent extraction and Lemieux oxidation. In this optimized analytical procedure, a sequential solvent extraction using 50% (v/v) methanol and EDTA-sodium pyrophosphate was selected as the optimal extraction solvent for free MCs analysis, after which the purified extracts and sediment residuals were applied to the optimized Lemieux oxidation for determination of total MCs in lake sediments. The optimized procedures were shown to be efficient and reliable for the routine analysis of both free and total MCs in lake sediment samples, as indicated by the minimal adverse impact of sediment organic matter on the recovery of free MCs and yield of MMPB (2-methyl-3-methoxy-4-phenylbutyric acid). Finally, the developed procedures were applied to field sediment samples collected from Lake Dianchi during a bloom season and seven of thirty samples showed positive results.

A sensitive method for the determination of total microcystins in water and sediment samples by liquid chromatography with fluorescence detection

Microcystins (MCs) are potent hepatotoxins that comprise a family of more than 90 different variants. MCs can bind to matrices, after which they cannot be completely extracted by the solvent. Currently, the MMPB (2-methyl-3-methoxy-4-phenylbutyric acid) method is the only effective way of determining total MCs without solvent extraction. In this study, a sensitive method to determine the total MCs in water and sediment was developed based on the MMPB method. Specifically, MCs were oxidized to MMPB with an improved oxidation reagent (20 mg mL−1 NaIO4, 4000 mg L−1 KMnO4, pH ∼ 9) and a stable MMPB yield of about 35% was obtained for both water and sediment samples. The minimum volume of the oxidation reagent could be determined by the organic content in the matrix. After being concentrated, the MMPB was derivatized with 1,2-benzo-3,4-dihydrocarbazole-9-ethyl-p-toluenesulfonate (BDETS) and detected by liquid chromatography using a fluorescence detector. Quantification limits of MCs in water and sediment samples were 125 ng L−1 and 100 ng g−1 respectively. The method was successfully applied to Lake Dianchi and the soluble MC concentrations determined by this method were correlated well with those obtained by the enzyme-linked immunosorbent assay (ELISA).

Evaluation of the potential of anoxic biodegradation of intracellular and dissolved microcystins in lake sediments

The kinetics of the anoxic biodegradation of intracellular and dissolved microcystin (MCs) and the effects of soluble organic materials on the process were investigated via a series of well-controlled microcosm experiments. The potential for the removal of intracellular and dissolved MCs from lake sediment differed. The dissolved MCs could be degraded to below the detection limit at 20 °C within one to 3 days after a lag phase of 2-6 days under anoxic conditions. The levels of intracellular MCs were also significantly reduced in the sun-dried cyanobacterial samples but not rapidly reduced in fresh cyanobacterial samples until the cells were ruptured. The addition of soluble organic matter enhanced the anoxic biodegradation of MCs. These results indicate that the application of anoxic biodegradation via lake sediments is an effective method to remove dissolved and intracellular MCs and that this process exhibits significant bioremediation potential for the further treatment of cyanobacteria.

Effect of long-term cultivation on soil organic carbon fractions and metal distribution in humic and fulvic acid in black soil, Northeast China

Cultivation affects soil organic matter and its fractions. Fulvic acid (FA) and humic acid (HA) make up an important part of soil organic matter, and their binding capacity influences heavy metal behaviour in soil. This research studied changes in soil organic components and the distribution of copper (Cu), lead (Pb), and zinc (Zn) in HA and FA affected by long-term cultivation in black soils. Uncultivated sites and their adjacent cultivated sites (18, 50, and >200 years) were selected. Alkaline sodium hydroxide/pyrophosphate extraction of humic substances and precipitation of HA by acidification were used to separate the HA and FA fractions. Concentrations of Cu, Pb, and Zn in HA and FA were determined. The content of soil organic carbon (C) had decreased by 30% after 200 years of cultivation. Cultivation led to a moderate decrease (38%) in HA and a minor decrease (7%) in FA. The CHA/CFA ratio, which is a humification parameter, decreased from 2.05 in the uncultivated soil to 1.38 in the soil cultivated for 200 years, indicating a lower degree of humification of organic matter in cultivated soils. Of the Na4P2O7 + NaOH-extracted Pb and Zn, 47–60% and 63–76%, respectively, was associated with FA, showing that FA has a high affinity for Pb and Zn. Of the Na4P2O7 + NaOH-extracted Cu, 55% was in the HA fraction. The share of Cu and Zn in the HA fraction decreased with cultivation time, but cultivation did not bring about a systematic change in Pb distribution in humic substances. The results show that cultivation can decrease the humified C content and metals bound to the HA fraction, and suggest that cultivation may potentially increase the mobility of heavy metals.