Microcystins Research Articles

Simultaneous electrochemical removal of three microcystin congeners and sulfamethoxazole in natural water

Microcystins (MCs), frequently detected in freshwater ecosystems, have raised significant human health and ecological concerns. New approaches are being developed to control and remove MCs. In this study, we examined factors influencing the efficacy of electrochemical oxidation as a means of control. Anode material (Pt/Ti, Ta2O5-IrO2/Ti, SnO2-SbO2/Ti, boron-doped diamond (BDD)/Si), anode surface area ratios and solution volumes, initial pollutant concentrations, and the co-existing antibiotic sulfamethoxazole (SMX) were investigated. MCs and SMX were dissolved in filtered Taihu Lake water to simulate the natural aquatic environment. The results showed that non-active anodes, lower initial concentration of MC, larger surface area ratio of anode to cathode, and smaller ratio of reaction solution volume to anode surface area could promote the degradation target pollutants. Under optimal conditions in this study, the degradation rates of MC-LR, MC-YR, MC-RR, and SMX each reached more than 90% within 6 hours, and the removal efficiency of MC-YR was the highest among three congeners. The effect of SMX on the degradation of MC congeners depended mainly on their concentration differences, such that when the initial concentration of SMX was one to two orders of magnitude lower than microcystin, the presence of SMX would promote the degradation of MCs. In contrast, when the initial concentration of SMX was higher than that of microcystin by approximately an order of magnitude, sulfamethoxazole would inhibit the degradation of MCs by between 4.6% and 24.5%. Ultra-high-performance liquid chromatography tandem mass spectrometry analysis revealed that the three MC congeners were electrochemically degraded through aromatic ring oxidation, alkene oxidation, and bond cleavage on the ADDA (3-amino-9-methoxy-2,6,8-trimethyl-10-phenyldeca-4,6-dienoic acid) side chain. Notably, the removal of MCs was accompanied by a decline in the hardness of the reaction water. This study provided insights into electrochemical degradation of microcystins and antibiotics in natural water, offering suggestions for its practical application.

Effects of Ciprofloxacin on the Production and Composition of Cellular Microcystins in Microcystis aeruginosa.

Antibiotics can affect the photosynthetic system of Microcystis, potentially altering the balance of carbon and nitrogen, which may influence the synthesis of different microcystin (MC) congeners. However, the regulatory mechanisms by which antibiotics affect the synthesis of various MC congeners in Microcystis remain unknown. In this study, the effects of ciprofloxacin (CIP) on the growth, carbon and nitrogen balance, amino acid composition, mcyB gene expression, and production of different MC congeners were investigated in two toxin-producing strains of Microcystis aeruginosa. The results show that CIP exposure significantly inhibited the growth of both strains, achieving an inhibition rate of 71.75% in FACHB-315 and 41.13% in FACHB-915 at 8 μg/L CIP by the end of the cultivation. The intracellular C:N ratio in FACHB-315 increased by 51.47%, while no significant change was observed in FACHB-915. The levels of leucine, tyrosine, and arginine, as identified and quantified by UPLC-MS/MS, were significantly altered at higher CIP concentrations, leading to a reduction in leucine percentage and a notable increase in tyrosine in both strains, which contributed to a reduction in MC-LR proportion and an increase in MC-RR and MC-YR proportion. Additionally, the expression of the mcyB gene was upregulated by as much as 5.57 times, indicating that antibiotic stress could enhance MC synthesis at the genetic level, contributing to the increased toxicity of cyanobacteria. These findings emphasize the significant role of CIP in the biochemical processes of M. aeruginosa, particularly in MC synthesis and composition, providing valuable insights into the ecological risks posed by antibiotics and harmful cyanobacteria.

Using dredged sediments from Lake Taihu as a plant-growing substrate: Focusing on the impact of microcystins

Using dredged sediment as plant growth substrates is a promising way to deal with large amounts of excavated sediments. However, it is a big challenge to deal with various pollutants in sediments, among which microcystins (MCs) gained limited attention. In this study, sediments collected from Lake Taihu were mixed with agricultural soil at a 1:1 ratio to create various growing substrates for lettuce (Lactuca sativa L. var. ramosa Hort.). Results indicated that fresh weight and leaf area of lettuce increased in some sediment-amended treatments due to additional nutrients, but food quality was negatively affected by sediment amendment as suggested by the soluble sugar and Vitamin C levels. MCs were detected in all lettuce grown in sediment-amended substrates, particularly in treatments with sediments collected during the bloom. The highest MC contents were found in treatment amended with sediments collected from Meiliang Bay in August (88.6 μg kg−1 for MC-LR and 65.6 μg kg−1 for MC-RR). MC accumulation in lettuce and the associated human health risks were significant, especially in treatments with sediments from the bloom period. Ecological risk assessments revealed high RQ values, indicating potential harm to the soil ecosystem. This study underscores the importance of considering MC content in sediments when evaluating their use as growing substrates. The findings contribute to understanding the environmental and health implications of sediment reuse, offering insights for safer agricultural practices and sediment management.

Laboratory-Simulated Inhibitory Effects of the Floating-Bed Plants on Microcystis aeruginosa and Their Microbial Communities' Responses to Microcystins.

Three common floating bed plants, Eichhornia crassipes, Pistia stratiotes, and Ipomoea aquatica, were selected in the present study to investigate their inhibitory effects on toxic Microcystis aeruginosa. The results showed that all three types of floating-bed plants could considerably inhibit the growth of M. aeruginosa and effectively remove the microcystins (MCs) from water systems, among which, E. crassipes and P. stratiotes were more effective in resisting M. aeruginosa, and the removal rate of the intracellular MCs could be up to 100%. In addition, the roots and leaves of the three plants were enriched with a large number of MCs and demonstrated significant antioxidant responses, as evidenced by the increase in the content of catalase (CAT), glutathione peroxidase (GSH-PX), superoxide dismutase (SOD), and malondialdehyde (MDA) in the roots, stems, and leaves of the plants. Furthermore, this study also showed that Proteobacteria, Bacteroidota, Myxococcota, Verrucomicrobiota, and Actinobacteriota dominated the root microorganisms of the three plants. Moreover, a variety of MC-degrading bacteria, including Sphingomonas, Acinetobacter, Novosphingobium, and Pseudomonas, were found at the genus level, which further provides important basic data for the regulation of eutrophic water bodies and the removal of MCs.

Surface amphiphilic hybrid porous polymers based on cage-like organosiloxanes for pipette tip solid-phase extraction of microcystins in water

The occurrence of microcystins (MCs) during harmful algal blooms (HABs) represents a major threat to freshwater environments. In this work, a novel surface amphiphilic hybrid porous polymers based on cage-like organosiloxanes (PCSs) was prepared for the enrichment of MCs. The copolymerization of bifunctional amphiphilic monomers, 2-methacryloyloxyethyl phosphorylcholine (MPC) and N-benzylquininium chloride (BQN), with the cross-linker methacryl substituted polyhedral oligomeric silsesquioxane (POSS) was achieved in an ionic liquid-based porogenic medium. The hierarchical porous structure, a variety of surface functional groups and weak hydrophilicity were well characterized on the prepared materials using scanning electron microscopy, nitrogen adsorption/desorption analysis, Fourier transform infrared spectroscopy, X-ray photoelectron spectroscopy, zeta potential analysis and water contact angle testing, respectively. The as-prepared surface amphiphilic PCSs was used as an adsorbent for pipette tip solid-phase extraction (PT-SPE) to enrich microcystins (MCs) from surface waters before their analysis by capillary electrochromatography (CEC) and liquid chromatography-mass spectrometry (LC-MS). Under the optimal conditions, the established PT-SPE-LC-MS method exhibited a wide linear range (10–10,000 ng L−1), low limits of detection (4.0–8.0 ng L−1) and satisfactory recoveries (89.5–102.8 %) for MCs. An adsorption mechanism involving electrostatic interactions, hydrogen bonding, hydrophilic interactions, and π-π stacking has been proposed. The findings suggest that the use of surface amphiphilic PCSs materials as adsorbents in the PT-SPE platform facilitates efficient enrichment of MCs for subsequent chromatographic analysis. These investigations offer a new perspective on the simple and uncomplicated pretreatment of complex environmental samples.

Contaminant Exposure and Liver and Kidney Lesions in North American River Otters in the Indian River Lagoon, Florida.

The harmful algal bloom (HAB) liver toxin microcystin (MC) and trace element biomagnification were previously detected in organisms in the Indian River Lagoon (IRL), Florida. Since there are no routine screening programs for these contaminants, liver tissue from North American river otters (Lontra canadensis), an important sentinel species in the IRL, was screened for MC via enzyme-linked immunoassay (ELISA), followed by confirmatory analyses via liquid-chromatography/mass spectrometry methods (LC-MS/MS). Liver and kidney samples were evaluated for trace element (As, Cd, Co, Cu, Fe, Hg, Mn, Mo, Pb, Se, Tl, and Zn) bioaccumulation via inductively coupled plasma mass spectrometry (ICP-MS). Histopathologic evaluation of the liver and kidney was conducted to assess possible correlation with toxic insults. Forty-three river otters were evaluated (2016-2022). Microcystin was not detected in any river otter sample (n = 37). Of those tested for trace element bioaccumulation (n = 22), no sample measured above provided reference ranges or estimated toxic thresholds for this species. There were no statistically significant patterns observed based on season, year, or age class, but sex had a small influence on trace element levels in the kidney. One individual had a kidney Cu level (52 μg/g dry weight) higher than any previously reported for this species. Trace elements were detected at presumed background levels providing baselines for future monitoring. For otters with available histopathologic evaluation (n = 28), anomalies indicative of contaminant exposure (non-specific inflammation, necrosis, and/or lipidosis) were present in the liver and kidney of 18% and 4% of individuals, respectively. However, since these lesions were not linked to abnormal trace element bioaccumulation or MC exposure, other causes (e.g., infectious disease) should be considered.

Evaluating physiological responses of microalgae towards environmentally coexisting microplastics: A meta-analysis

Microplastics (MPs) are abundantly present in aquatic environments, where the phytoplankton—microalgae, are now inevitably bound to a long-term coexistence with them. While numerous studies have focused on the toxicological effects of high-concentration MPs exposure, there remains controversy over whether and how MPs affect microalgae at environmentally relevant concentrations. This study aims to draw conclusions that narrow the gap from 52 studies with varying results. Overall, MPs can inhibit growth and photosynthesis, induce oxidative damage, from which microalgae can recover after an appropriate period. Cyanobacteria exhibit greater vulnerability than chlorophyta. The relative size of MPs to algal cells potentially governs their coexistence behavior, thereby altering the mechanisms of impact. Pristine MPs may increase the production of extracellular polymeric substances (EPS) and microcystins (MCs), while aged MPs have the opposite effect. Additionally, relevant factors are systematically discussed, offering insights for future research.

Microcystins are present in water treatment plant residuals and are impacted by extraction and quantification methodology

ABSTRACT Microcystins (MCs), a toxin produced by some species of the photosynthetic autotrophic cyanobacteria, are the most studied and monitored cyanotoxin in water. Water treatment plant (WTP) residuals are the byproduct of water treatment consisting of solids removed from WTP processes and have been shown to contain cyanobacterial cells. However, the presence of MCs in WTP residuals has not been systematically demonstrated. Samples from four different WTPs across the United States were used to quantify MCs in residuals while assessing extraction and quantification methods adapted from water samples for solid matrices. MCs were present in 100% of samples. MC-LA was the most prevalent variant in these samples (70.05% of MCs quantified by UPLC-PDA). Natural degradation observed in a WTP storage lagoon was also investigated to determine the impact of physical, chemical, and biological processes on MC concentrations in high-biomass residuals. This study demonstrates that residuals of various characteristics across the United States contain MCs, and no one method was found to maximize results consistently across all samples. Cyanotoxins accumulating in WTP residuals are a growing concern. Implications of this work can help regulations and future studies of potential reuse applications and understanding of potential ecological significance of MCs accumulating in WTP residuals.

New tools for effective production and long-term stabilization of microcystinase (MlrA) - A biotechnological perspective towards hepatotoxic microcystins remediation

Microcystinase (MlrA) is the most specific catalyst and the most efficient enzyme of known microcystins (MCs) detoxification pathways. Very recently, direct MlrA application has effectively degraded MCs within industrial processes, demonstrating MlrA application for sustainable in situ MCs remediation. Heterologous MlrA expression in cyanobacteria offers a unique opportunity, linking harmful MCs remediation with emerging cyanobacterial biotechnologies. Here, we first generate a novel Synechocystis sp. PCC 6803 (6803) using non-native trc promoter for MlrA expression. Whole-cell MlrA activity was comparable to previously described expression via native 6803 PcpcB560 promoter, while cellular extracts of the new strain showed significantly higher MlrA yields (2–15 times, depending on the age of the cultures). Furthermore, efficiency of MlrA production under multiple photoautotrophic cultivation conditions varied, but was not improved by Na2CO3 supplementation nor under increased light, indicating the need to explore new photoautotrophic chassis for higher MlrA productivity.Methods for MlrA stabilization are critical for industrial development, thus lyophilization of MlrA-enriched cellular extracts was explored. Recovered MlrA activity was not statistically different from initial MlrA activity following storage of lyophilized extracts at −20 °C for 20 weeks. In contrast, storage at 20 °C and storage of aqueous lysates at 4 °C resulted in progressive MlrA activity loss. Such stabilized lyophilizate may offer a checkpoint for further optimization of upstream processes (production), while expanding potential for downstream investigations (application), contributing simultaneously to novel MlrA-based MCs remediation approaches and to cyanobacterial biotechnology.

Accumulation of microcystin toxin in irrigation water and alfalfa (Medicago sativa) forage plant, and assessing the potential risk to animal health

Microcystin (MC) toxin produced by cyanobacteria has become a significant concern for societies worldwide. The risk of MC in drinking water has been assessed to human health. Nonetheless, its risk to animal health has not been thoroughly evaluated. This study investigated MCs in irrigation water and alfalfa plant from nearby farmlands. Both irrigation water and alfalfa shoots contained greater MC concentrations (1.8–17.4 μg L−1 and 0.053–0.128 μg g−1) during summer than winter (2.4 μg L−1 and 0.017 μg g−1). These MC concentrations showed a correlation with the predominance of cyanobacteria in the sites, triggering the potential risk of these microorganisms in irrigation waters. Accordingly, there would be a high risk (risk quotient, RQ > 1) during summer and a moderate risk (0.1<RQ < 1) during winter for cattle and sheep that drink polluted irrigation water or eat contaminated alfalfa plants. Therefore, the study suggests that cyanotoxins in forage plants and irrigation water sources should be regularly monitored to protect animals from consuming contaminated food and water.

Microcystin-degrading bacteria reduce bioaccumulation in Fragaria vulgaris and enhance fruit yield and quality

In Morocco, red fruit production has thrived, primarily utilizing hydroponic methods to control crops, increase fruit yield and quality, and avoid soil-related problems. However, the irrigation of these expansive hydroponic farms relies heavily on water sourced from dams, many of which are contaminated with Microcystins (MCs). To address this contamination issue, ongoing research is focused on discovering effective and cost-efficient biological solutions for eliminating MCs. In this study, we isolate and identify bacterial strains capable of degrading MCs, evaluate the rate of degradation, and investigate how soil inoculated with these bacteria affects the accumulation of MCs in plant tissue. The partial 16S rRNA analyses of three bacterial sequences were conducted, identifying them through NCBI as follows: Ensifer sp. (B1) isolated from soil, Shinella sp. (B2) from a cyanobacterial bloom, and Stutzerimonas sp. (B3) from water. These bacteria exhibited the ability to degrade MCs, with approximately 34.75%, 73.75%, and 30.1% of the initial concentration (20 µg/L) being removed after a 6-day period for B1, B2, and B3, respectively. Moreover, strawberry plants were cultivated hydroponically in a greenhouse for a duration of 90 days. These plants were subjected to extracts of cyanobacteria containing 10 and 20 µg/L of Microcystins (MC), as well as water from an artificial lake contaminated with MC, both with and without the presence of isolated bacterial strains. Among these strains, Shinella sp. exhibited the highest efficacy in mitigating MC accumulation. Specifically, it resulted in a reduction of approximately 1.159 µg of MC per kilogram of root dry weight, leading to complete elimination in the leaves and fruits. The findings also indicated that the inoculation of perlite with the three MC-degrading bacterial strains significantly enhanced growth, photosynthetic pigments, yield, biochemical constituents, and quality attributes of strawberries (p ≤ 0.05). These promising outcomes suggest the potential of this approach for addressing the adverse impacts of crops irrigated with MC-contaminated water in future agricultural practices.

Co-Occurrence of Cyanotoxins and Phycotoxins in One of the Largest Southeast Asian Brackish Waterbodies: A Preliminary Study at the Tam Giang—Cau Hai Lagoon (Vietnam)

The Tam Giang-Cau Hai lagoon (TGCH) in Thua Thien Hue province (Vietnam) is a marsh/lagoon system and ranks among the largest waterbodies in Southeast Asia. It plays a significant role in terms of both socio-economic and environmental resources. However, anthropogenic stress, as well as the discharge of untreated domestic and industrial sewage with agricultural runoff from its three major tributaries, dramatically damages the water quality of the lagoon. Especially after heavy rain and flash floods, the continuous degradation of its water quality, followed by harmful algal and cyanobacterial bloom patterns (HABs), is more perceptible. In this study, several physicochemical factors, cyanotoxins (anatoxins (ATXs), saxitoxins (STXs), microcystins (MCs)), phycotoxins (STXs, okadaic acid (OA), and dinophysistoxins (DTXs)) were analyzed in water and shellfish samples from 13 stations in June 2023 from 13 stations, using enzyme-linked immunosorbent assay (ELISA) kits for the ATXs and STXs, and the serine/threonine phosphatase type 2A (PP2A) inhibition assay kit for the MCs, OA, and DTXs. The results showed for the first time the co-occurrence of freshwater cyanotoxins and marine phycotoxins in water and shellfish samples in this lagoon. Traces of ATXs and STXs were detected in the shellfish and the orders of magnitude were below the seafood safety action levels. However, toxins inhibiting the PP2A enzyme, such as MCs and nodularin (NODs), as well as OA and DTXs, were detected at higher concentrations (maximum: 130.4 μg equiv. MC-LR/kg shellfish meat wet weight), approaching the actionable level proposed for this class of toxin in shellfish (160 μg of OA equivalent per kg of edible bivalve mollusk meat). It is very important to note that due to the possible false positives produced by the ELISA test in complex matrices such as a crude shellfish extract, this preliminary and pilot research will be repeated with a more sophisticated method, such as liquid chromatography coupled with mass spectroscopy (LC-MS), in the upcoming research plan.

Interspecific competition enhances microcystin production by Microcystis aeruginosa under the interactive influences of temperature and nutrients

Global warming and eutrophication contribute to frequent occurrences of toxic algal blooms in freshwater systems globally, while there is a limited understanding of their combined impacts on toxin-producing algal species under interspecific competitions. This study investigated the influences of elevated temperatures, lights, nutrient enrichments and interspecific interactions on growth and microcystin (MC) productions of Microcystis aeruginosa in laboratory condition. Our results indicated that elevated temperatures and higher nutrient levels significantly boosted biomass and specific growth rates of Microcystis aeruginosa, which maintained a competitive edge over Chlorella sp. Specifically, with phosphorus levels between 0.10 and 0.70 mg P L−1, the growth rate of Microcystis aeruginosa in mixed cultures increased by 23 %–52 % compared to mono-cultures, while the growth rate of Chlorella sp. shifted from positive in mono-cultures to negative in mixed cultures. Redundancy and variance partition analyses suggested that Chlorella sp. stimulate MC production in Microcystis aeruginosa and nutrient levels outshine temperature for toxin productions during competition. Lotka‒Volterra model revealed a positive correlation between the intensities of competitions and MC concentration. Our findings indicate that future algal bloom mitigation strategies should consider combined influence of temperature, nutrients, and interspecific competition due to their synergistic effects on MC productions.

Widespread, low concentration microcystin detection in a subtropical Louisiana estuary

Spatiotemporal patterns and drivers of hepatotoxic microcystins (MC) were investigated in the Atchafalaya-Vermilion Bay System (AVBS), a subtropical, river-dominated estuary in Louisiana. Along with environmental data, monthly particulate MC (pMC) samples were examined over a two-year period (2016–2018), and biweekly pMC and dissolved MC (dMC) samples were examined over a five-month period in 2020. Solid phase adsorption toxin tracking (SPATT) samplers used to quantify time-integrated dMC concentrations were also deployed in 2020. Low, but detectable concentrations of pMC (≤0.033 μg L−1) and dMC (≤0.190 μg L−1) were found throughout the AVBS in 37.8 and 21.2 % of samples, respectively. Time integrative SPATT samplers detected dMC in nearly 100 % of the deployments, compared to dMC detections in 30.8 % of the discrete samples. This study documents widespread MC presence throughout the AVBS and while concentrations were low, knowledge gaps remain regarding the potential long-term impacts of sublethal MC exposure to estuarine organisms.

Exposure to microcystin-LR promotes the progression of colitis-associated colorectal cancer by inducing barrier disruption and gut microbiota dysbiosis

Microcystins (MCs) are secondary metabolites generated by cyanobacterial blooms, among which microcystin-LR (MC-LR) stands out as the most widely distributed variant in aquatic environments. However, the effects of MC-LR on the colorectum and its role in promoting colorectal tumor progression remain unclear. Therefore, this study aims to scrutinize the impact of MC-LR on a mice model of colitis-associated colorectal cancer and elucidate the potential underlying molecular mechanisms. In this study, we used AOM/DSS mice and orally administered MC-LR at doses of 40 µg/kg or 200 µg/kg. Exposure to MC-LR increased tumor burden, promoted tumor growth, shortened colon size, and decreased goblet cell numbers and tight junction protein levels in intestinal tissues. Additionally, exposure to MC-LR induced alterations in the structure of gut microbiota in the mouse colon, characterized by an increase in the relative abundance of Escherichia_coli and Shigella_sonnei, and a decline in the relative abundance of Akkermansia_muciniphila. Transcriptomic analysis revealed that MC-LR exposure activated the IL-17 signaling pathway in mouse colorectal tissues and participated in inflammation regulation and immune response. Immunofluorescence results demonstrated an increase in T-helper 17 (Th17) cell levels in mouse colorectal tumors following MC-LR exposure. The results from RT-qPCR revealed that MC-LR induced the upregulation of IL-6, IL-1β, IL-10, IL-17A, TNF-α, CXCL1, CXCL2, CXCL5 and CCL20. The novelty of this study lies in its comprehensive approach to understanding the mechanisms by which MC-LR may contribute to CRC progression, offering new perspectives and valuable reference points for establishing guidance standards regarding MC-LR in drinking water. Our findings suggest that even at guideline value, MC-LR can have profound effects on susceptible mice, emphasizing the need for a reevaluation of guideline value and a deeper understanding of the role of environmental toxins in cancer progression.

Inhibition of JNK transcription via the Nrf2/Keap1a pathway to resist microcystin-induced oxidative stress and apoptosis in freshwater mussels Cristaria plicata

With global warming and increasing eutrophication of water bodies, a variety of algal toxins, including microcystin (MC), released into water by cyanobacterial blooms pose a serious threat to the survival of aquatic organisms. To investigate the mechanism of the Nrf2/Keap1a pathway on resisting MC-induced oxidative stress and apoptosis in Cristata plicata, we cloned the full-length cDNA of CpBcl-2. The cDNA full-length of CpBcl-2 was 760 bp, encoded a 177 amino acid peptide, and contained a highly conserved Bcl-2-like superfamily domain. MC stimulation increased the expression and activity levels of related antioxidant enzymes. After CpNrf2 knockdown, the transcription levels of NAD(P)H quinone redox Enzyme-1 (NQO1) and related antioxidant enzymes activity in the gills and kidney of C. plicata were significantly down-regulated upon MC stress, but that was significantly upregulated after knockdown of CpKeap1a. Additionally, Upon MC stress, the mRNA levels of CpBcl-2 were increased in the gills and kidney after knockdown of CpNrf2 at 24 h, and that of CpBcl-2 were decreased at 72 and 96 h in the CpKeap1a-siRNA+MC group. Moreover, MC stimulation significantly inhibited CpJNK expression in the gills and kidney, but which regulated the Nrf2/Keap1a pathway in C. plicata. However, the JNK inhibitor SP600125 promoted the expression of CpNrf2 and related enzymes with antioxidant response element (ARE-driven enzyme) in the gills and kidney. Then, we speculated that CpKeap1a was a negative regulator of CpNrf2, and C. plicata resisted MC-induced oxidative damage and apoptosis by inhibiting JNK transcription via the Nrf2/Keap1a pathway.

Self-assembled flower-like carbon nanosheets for magnetic solid-phase extraction of microcystins from aquatic organism

Adsorbents with good dispersibility and high efficiency are crucial for magnetic solid-phase extraction (MSPE). In this study, flower-like magnetic nanomaterials (F-Ni@NiO@ZnO2-C) were successfully prepared by calcination of metal-organic framework (MOF) precursors that was stacked by two-dimensional (2D) nanosheet. The synthesized F-Ni@NiO@ZnO2-C has a flower-like layered structure with a large amount of pore space, promoting the rapid diffusion of targets. In addition, Zn2+ doped in MOF precursors was still retained that further produced strong metal chelation with targets. The unique structure of F-Ni@NiO@ZnO2-C was used as MSPE adsorbent, and combined with high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) for extraction of three microcystins (MCs) detection, including microcystin-LR (MC-LR), microcystin-RR (MC-RR), microcystin-YR (MC-YR). The resulting method has a detection limit of 0.2–1.0 pg mL−1, a linear dynamic range of 0.6–500.0 pg mL−1 and has good linearity (R ≥ 0.9996). Finally, the established method was applied to the highly selective enrichment of MCs in biological samples, successfully detecting trace amounts of MCs (8.4–15.0 pg mL−1) with satisfactory recovery rates (83.7–103.1 %). The results indicated that flower-like magnetic F-Ni@NiO@ZnO2-C was a promising adsorbent, providing great potential for the determination of trace amounts of MCs in biological samples.

Analysis of Total Microcystins using LC-MS/MS: Assessment of Pretreatment Technique for MMPB(2-methyl-3-methoxy-4-phenylbutyric acid)

Algal toxin analysis, particularly for microcystin (MC) variants, is commonly performed using LC-MS/MS as the standard method. In 2016, the environmental protection agency (EPA) published method 546 for the determination of total MCs by ELISA. However, the limited availability of commercial algal toxin standards limits the efficacy of the LC-MS/MS method. This study proposes a novel approach using the 2-methyl-3-methoxy-4-phenylbutyric acid (MMPB) structure generated by oxidizing MCs with potassium permanganate and sodium periodate. MMPB is formed through the oxidation of the common (all-S,all-E)-3-amino-9-methoxy-2,6,8-trimethyl-10-phenyldeca-4,6-dienoic acid (ADDA) structure, eliminating the need for specific standard materials for each MC variant. This provides a method for measuring the total MCs. This study focused on optimizing sample pretreatment before oxidation, salt removal as a post-oxidation step, and the quantitative method to establish a comprehensive total MC detection approach applicable to surface water samples. The whole process included pretreatment using a freeze-drying method, purification using liquid-liquid extraction (LLE) for salt removal, and a comparison of quantitative methods using internal standards. Additionally, an evaluation of internal standards was conducted to assess availability during the oxidation process. In this study, an MMPB recovery rate of up to 50% was achieved, with an analysis precision of 14%. According to the results of QAQC, the satisfactory linearity of the calibration curve was a precision and the performance of the method for spiked samples was verified. The MMPB analysis method developed in this study showed relatively lower quantification limits and recovery rates than individual MC analysis methods. We believe these results stem from the inherent sensitivity differences between MCs and MMPB, as well as the complexity of the pre-treatment process centered around the oxidation process.

Microcystins in the benthic food-web of the Sacramento-San Joaquin Delta, California

Harmful cyanobacteria blooms are a growing threat in estuarine waters as upstream blooms are exported into coastal environments. Cyanobacteria can produce potent toxins, one of which—hepatotoxic microcystins (MCs)—can persist and accumulate within the food web. Filter-feeding invertebrates may biomagnify toxins up to 100× ambient concentrations. As such, bivalves can be used as an environmentally relevant and highly sensitive sentinel for MC monitoring. To date there has been little research on cyanotoxin bioaccumulation in estuaries. The Sacramento-San Joaquin Delta (Delta) aquatic food web has undergone a profound change in response to widespread colonization of aquatic invasive species such as Asian clams (Corbicula fluminea) in the freshwater portion of the Delta. These clams are prolific—blanketing areas of the Delta at densities up to 1000 clams/m2 and are directly implicated in the pelagic organism decline of threatened and endangered fishes. We hypothesized that Asian clams accumulate MCs which may act as an additional stressor to the food web and MCs would seasonally be in exceedance of public health advisory levels. MCs accumulation in Delta Asian clams and signal crayfish (Pacifastacus leniusculus) were studied over a two-year period. ELISA and LC-MS analytical methods were used to measure free and protein-bound MCs in clam and crayfish tissues. We describe an improved MC extraction method for use when analyzing these taxa by LC-MS. MCs were found to accumulate in Asian clams across all months and at all study sites, with seasonal maxima occurring during the summer. Although MC concentrations rarely exceeded public health advisory levels, the persistence of MCs year-round still poses a chronic risk to consumers. Crayfish at times also accumulated high concentrations of MCs. Our results highlight the utility of shellfish as sentinel organisms for monitoring in estuarine areas.

Sustained and enhanced inhibitory effects of allelochemicals on Microcystis Aeruginosa during its recruitment stage

Microcystis aeruginosa is the main toxic strain in cyanobacterial blooms, and the recruitment stage in its temperature-dependent seasonal succession is considered as the key to its subsequent growth. In this study, a protocol with specific temperature settings was developed as the simulated recruitment stage in order to investigate and confirm the superior inhibitory effects of allelochemicals on M. aeruginosa at that stage of recruitment. One of the most common allelochemicals, gallic acid (GA) (10 mg/L, 20 mg/L) was employed to treat M. aeruginosa under initially low temperature condition (15 °C), then intermediate (20 °C) and last normal (26 °C), which corresponds to the critical temperatures for cyanobacterial recruitment and growth. Growth, metabolism, photosynthetic activity, extracellular polysaccharides (EPS) and microcystins (MCs) release were analyzed and discussed in this study, and a more sustained and better inhibitory effect over a 20-day period was achieved. Notably, GA (10 mg/L) markedly delayed the recruitment of M. aeruginosa from low temperature, with an inhibition efficiency of 85.71 %, and suppressing Fv/Fm and photosynthetic pigments production. It is also observed that M. aeruginosa at recruitment stage exhibited higher sensitivity and poorer resistance to allelochemical treatment, with variable responses suggesting that optimal dosages may alter. The antioxidant enzyme activities remained high under prolonged stress, and the secretion of EPS was stimulated, indicating that cyanobacteria were more inclined to form colonies. While the laboratory-based inhibitory mechanism appeared to increase the release of microcystins in individual cells, the actual concentration of microcystins in natural aquatic environments requires further investigation.