Search Results (13)

Cyanobacteria can produce a wide range of toxins which have acute and chronic adverse health effects. Affecting a variety of mammalian systems, they are generally characterized according to their mode of action and the organs affected. Cyanobacterial neurotoxins are one cyanotoxin class that can negatively affect human health, and representatives of other cyanotoxins classes are increasingly showing neurotoxic effects. Of the various human exposure routes to cyanobacterial toxins, the significance of the airborne and inhalation route requires much greater clarity and understanding. People may be exposed to mixtures of cyanobacterial neurotoxins through the inhalation of sprays and dust, along with the potential to directly enter the central nervous system when crossing the blood-brain barrier. This review aims to summarize the current state of knowledge concerning airborne cyanobacterial neurotoxins, research gaps, health effects, and the need for management practices to protect human and animal health. Full article

Lake Winnipeg in Manitoba, Canada is heavily impacted by harmful algal blooms that contain non-protein amino acids (NPAAs) produced by cyanobacteria: N-(2-aminoethyl)glycine (AEG), β-aminomethyl-L-alanine (BAMA), β-N-methylamino-L-alanine (BMAA), and 2,4-diaminobutyric acid (DAB). Our objective was to investigate the impact of microbial diversity on NPAA production by cyanobacteria using semi-purified crude cyanobacterial cultures established from field samples collected by the Lake Winnipeg Research Consortium between 2016 and 2021. NPAAs were detected and quantified by ultra-performance liquid chromatography–tandem mass spectrometry (UPLC-MS/MS) using validated analytical methods, while Shannon and Simpson alpha diversity scores were determined from 16S rRNA metagenomic sequences. Alpha diversity in isolate cultures was significantly decreased compared to crude cyanobacterial cultures (p < 0.001), indicating successful semi-purification. BMAA and AEG concentrations were higher in crude compared to isolate cultures (p < 0.0001), and AEG concentrations were correlated to the alpha diversity in cultures (r = 0.554; p < 0.0001). BAMA concentrations were increased in isolate cultures (p < 0.05), while DAB concentrations were similar in crude and isolate cultures. These results demonstrate that microbial community complexity impacts NPAA production by cyanobacteria and related organisms. Full article

Of the wide variety of toxic compounds produced by cyanobacteria, the neurotoxic amino acid β-N-methylamino-l-alanine (BMAA) has attracted attention as a result of its association with chronic human neurodegenerative diseases such as ALS and Alzheimer’s. Consequently, specific detection methods are required to assess the presence of BMAA and its isomers in environmental and clinical materials, including cyanobacteria and mollusks. Although the separation of isomers such as β-amino-N-methylalanine (BAMA), N-(2-aminoethyl)glycine (AEG) and 2,4-diaminobutyric acid (DAB) from BMAA has been demonstrated during routine analysis, a further compounding factor is the potential presence of enantiomers for some of these isomers. Current analytical methods for BMAA mostly do not discriminate between enantiomers, and the chiral configuration of BMAA in cyanobacteria is still largely unexplored. To understand the potential for the occurrence of D-BMAA in cyanobacteria, a chiral UPLC-MS/MS method was developed to separate BMAA enantiomers and isomers and to determine the enantiomeric configuration of endogenous free BMAA in a marine Lyngbya mat and two mussel reference materials. After extraction, purification and derivatization with N-(4-nitrophenoxycarbonyl)-l-phenylalanine 2-methoxyethyl ester ((S)-NIFE), both L- and D-BMAA were identified as free amino acids in cyanobacterial materials, whereas only L-BMAA was identified in mussel tissues. The finding of D-BMAA in biological environmental materials raises questions concerning the source and role of BMAA enantiomers in neurological disease. Full article

β-N-methylamino-L-alanine (BMAA) and its isomers, 2,4-diaminobutyric acid (2,4-DAB) and N-(2-aminoethyl)-glycine (AEG), along with microcystins (MCs)-RR, -LR, and -YR (the major MC congeners), are cyanotoxins that can cause detrimental health and environmental impacts during toxic blooms. Currently, there are no reverse-phase (RP) LC-MS/MS methods for the simultaneous detection and quantification of BMAA, its isomers, and the major MCs in a single analysis; therefore, multiple analyses are required to assess the toxic load of a sample. Here, we present a newly developed and validated method for the detection and quantification of BMAA, 2,4-DAB, AEG, MC-LR, MC-RR, and MC-YR using RP LC-MS/MS. Method validation was performed, assessing linearity (r² > 0.996), accuracy (>90% recovery for spiked samples), precision (7% relative standard deviation), and limits of detection (LODs) and quantification (LOQs) (ranging from 0.13 to 1.38 ng mL⁻¹). The application of this combined cyanotoxin analysis on a culture of Microcystis aeruginosa resulted in the simultaneous detection of 2,4-DAB (0.249 ng mg⁻¹ dry weight (DW)) and MC-YR (4828 ng mg⁻¹ DW). This study provides a unified method for the quantitative analysis of BMAA, its isomers, and three MC congeners in natural environmental samples. Full article

In recent years, the consumption of blue-green algae (BGA) dietary supplements is increasing because of their health benefits. However, cyanobacteria can produce cyanotoxins, which present serious health risks. In this work we propose hydrophilic interaction liquid chromatography coupled with tandem mass spectrometry (HILIC-MS/MS) to determine cyanotoxins in BGA dietary supplements. Target toxins, including microcystin-leucine-arginine (MC-LR) and microcystin-arginine-arginine (MC-RR), nodularin, anatoxin-a and three non-protein amino acids, β-N-methylamino-L-alanine (BMAA), 2,4-diaminobutyric acid (DAB) and N-(2-aminoethyl)glycine (AEG), were separated using a SeQuant ZIC-HILIC column. Cyanotoxin extraction was based on solid–liquid extraction (SLE) followed by a tandem-solid phase extraction (SPE) procedure using Strata-X and mixed-mode cation-exchange (MCX) cartridges. The method was validated for BGA dietary supplements obtaining quantification limits from 60 to 300 µg·kg⁻¹. Nine different commercial supplements were analyzed, and DAB, AEG, and MCs were found in some samples, highlighting the relevance of monitoring these substances as precaution measures for the safe consumption of these products. Full article

Polymyxins are considered a last-line treatment against infections caused by multidrug-resistant (MDR) Gram-negative bacteria. In addition to their use as a potent antibiotic, polymyxins have also been utilized as outer membrane (OM) permeabilizers, capable of augmenting the activity of a partner antibiotic. Several polymyxin derivatives have been developed accordingly, with the objective of mitigating associated nephrotoxicity. The conversion of polymyxins to guanidinylated derivatives, whereby the L-γ-diaminobutyric acid (Dab) amines are substituted with guanidines, are described herein. The resulting guanidinylated colistin and polymyxin B (PMB) exhibited reduced antibacterial activity but preserved OM permeabilizing properties that allowed potentiation of several antibiotic classes. Rifampicin, erythromycin, ceftazidime and aztreonam were particularly potentiated against clinically relevant MDR Gram-negative bacteria. The potentiating effects of guanidinylated polymyxins with ceftazidime or aztreonam were further enhanced by adding the β-lactamase inhibitor avibactam. Full article

Cyanobacteria are a diverse group of oxygenic photosynthetic prokaryotes, which are believed to be one of the oldest life forms on Earth. They live in a wide range of ecosystems and withstand extreme environmental conditions. An important proportion of cyanobacteria is known to be producers of harmful cyanotoxins, which are toxic secondary metabolites that can impact the ecosystem and human health. The oral route is one of the main ways whereby humans can be exposed to cyanotoxins. Therefore, the consumption of contaminated algae-based food supplements is becoming more relevant due to its upsurge, which underlines the importance of controlling these toxins in this kind of products. This work describes the simultaneous determination of seven cyanotoxins belonging to three different classes: the cyclic peptides microcystin-LR (MC-LR), microcystin-RR (MC-RR) and nodularin (NOD); the alkaloid anatoxin-a (ANA) and three non-protein amino acids isomers β-methylamine-L-alanine (BMAA), 2,4-diaminobutyric acid (DAB) and N-(2-aminoethyl)glycine (AEG). These have been determined in spirulina-derived food supplements using a novel solid–liquid extraction coupled with a solid phase extraction procedure for clean up and preconcentration (SLE-tandem-SPE) and analysis by hydrophilic interaction liquid chromatography with tandem mass spectrometry detection (HILIC-MS/MS). A SeQuant® Zwitterionic Hydrophilic Interaction Liquid Chromatography (ZIC-HILIC) column (EMD Millipore, Billerica, MA, USA) was employed to achieve the chromatographic separation in less than 12 min using water and acetonitrile, both acidified with 0.3% of formic acid, as mobile phase. Previously, an SLE was developed, using 4 mL of aqueous 5% formic acid to extract the most polar compounds, followed by 4 mL of 80% MeOH. Both extracts were combined and submitted to a tandem-SPE using mixed-mode cation exchange (MCX) and Strata-X cartridges. Elution from both cartridges was performed using 10% NH₃·H₂O in MeOH. Method validation was carried out in terms of linearity, limit of detection (LOD) and quantification (LOQ), recoveries, matrix effect and repeatability and intermediate precision. LOQs in the range of 50–300 µg·kg⁻¹ and recoveries ranging between 64.2% and 102.9% with an associated relative standard deviation < 19.2% were achieved. Satisfactory precision was obtained with RSD values lower than 19.6% in all cases, with the exception of BMAA, which reported the highest RSD values, reaching 25.1%. The method was satisfactorily applied to determine the occurrence of cyanotoxins in blue green algae (BGA) dietary supplements. DAB was the most frequently detected cyanotoxin, at concentrations up to 2408 µg·kg⁻¹, and AEG was found in few samples at concentrations up to 194 µg·kg⁻¹. However, MC-LR and MC-RR were found in one sample at concentration levels higher than 5 mg·kg⁻¹, which underlines the need to control these substances in these matrices. Full article

The neurotoxic alkaloid β-N-methyl-amino-l-alanine (BMAA) and related isomers, including N-(2-aminoethyl glycine) (AEG), β-amino-N-methyl alanine (BAMA), and 2,4-diaminobutyric acid (DAB), have been reported previously in cyanobacterial samples. However, there are conflicting reports regarding their occurrence in surface waters. In this study, we evaluated the impact of amending lake water samples with trichloroacetic acid (0.1 M TCA) on the detection of BMAA isomers, compared with pre-existing protocols. A sensitive instrumental method was enlisted for the survey, with limits of detection in the range of 5–10 ng L⁻¹. Higher detection rates and significantly greater levels (paired Wilcoxon’s signed-rank tests, p < 0.001) of BMAA isomers were observed in TCA-amended samples (method B) compared to samples without TCA (method A). The overall range of B/A ratios was 0.67–8.25 for AEG (up to +725%) and 0.69–15.5 for DAB (up to +1450%), with absolute concentration increases in TCA-amended samples of up to +15,000 ng L⁻¹ for AEG and +650 ng L⁻¹ for DAB. We also documented the trends in the occurrence of BMAA isomers for a large breadth of field-collected lakes from Brazil, Canada, France, Mexico, and the United Kingdom. Data gathered during this overarching campaign (overall, n = 390 within 45 lake sampling sites) indicated frequent detections of AEG and DAB isomers, with detection rates of 30% and 43% and maximum levels of 19,000 ng L⁻¹ and 1100 ng L⁻¹, respectively. In contrast, BAMA was found in less than 8% of the water samples, and BMAA was not found in any sample. These results support the analyses of free-living cyanobacteria, wherein BMAA was often reported at concentrations of 2–4 orders of magnitude lower than AEG and DAB. Seasonal measurements conducted at two bloom-impacted lakes indicated limited correlations of BMAA isomers with total microcystins or chlorophyll-a, which deserves further investigation. Full article

The determination of small highly polar metabolites at low concentrations is challenging when reverse-phase (RP) chromatography is used for multiclass analysis. A mixed cationic-RP column coupled to high-resolution tandem MS (HR-MS/MS) was tested for highly polar compounds in biological fluids, i.e., trimethylamine N-oxide (TMAO) and the isobaric molecules beta-methylamino-L-alanine (BMAA) and 2,4-diaminobutyric acid (DAB). The efficient retention and separation of the above compounds were obtained with common and MS-friendly RP conditions, reaching high selectivity and sensitivity. The method was firstly assessed in plasma and urine, showing good linearity in the range 50–1000 µg/L and 500–10,000 µg/L for TMAO and both BMAA and DAB, respectively. Excellent precision (RDS < 3%) and good accuracies (71–85%) were observed except for BMAA in plasma, whose experimental conditions should be specifically optimized. Preliminary tests performed on compounds with biological relevance and a wider range of polarities proved the effectiveness of this chromatographic solution, allowing the simultaneous analysis of a larger panel of metabolites, from very small and polar compounds, like trimethylamine, to quite lipophilic molecules, such as corticosterone. The proposed LC-HRMS protocol is an excellent alternative to hydrophilic interaction liquid chromatography and ion-pairing RP chromatography, thus providing another friendly analytical tool for metabolomics. Full article

β-methylamino-L-alanine (BMAA) is a non-protein amino acid that has been implicated as a risk factor for motor neurone disease (MND). BMAA is produced by a wide range of cyanobacteria globally and by a small number of marine diatoms. BMAA is commonly found with two of its constitutional isomers: 2,4-diaminobutyric acid (2,4-DAB), and N-(2-aminoethyl)glycine (AEG). The isomer 2,4-DAB, like BMAA, has neurotoxic properties. While many studies have shown BMAA production by cyanobacteria, few studies have looked at other algal groups. Several studies have shown BMAA production by marine diatoms; however, there are no studies examining freshwater diatoms. This study aimed to determine if some freshwater diatoms produced BMAA, and which diatom taxa are capable of BMAA, 2,4-DAB and AEG production. Five axenic diatom cultures were established from river and lake sites across eastern Australia. Cultures were harvested during the stationary growth phase and intracellular amino acids were extracted. Using liquid chromatography triple quadrupole mass spectrometry (LC-MS/MS), diatom extracts were analysed for the presence of both free and protein-associated BMAA, 2,4-DAB and AEG. Of the five diatom cultures analysed, four were found to have detectable BMAA and AEG, while 2,4-DAB was found in all cultures. These results show that BMAA production by diatoms is not confined to marine genera and that the prevalence of these non-protein amino acids in Australian freshwater environments cannot be solely attributed to cyanobacteria. Full article

The neurotoxin β-N-methylamino-l-alanine (BMAA) has been identified as an environmental factor triggering neurodegenerative diseases such as Amyotrophic Lateral Sclerosis (ALS) and Alzheimer’s disease (AD). We investigated the possible vectors of BMAA and its isomers 2,4-diaminobutyric acid (DAB) and N-2(aminoethyl)glycine (AEG) in marine mollusks collected from the Chinese coast. Sixty-eight samples of marine mollusks were collected along the Chinese coast in 2016, and were analyzed by an HILIC-MS/MS (hydrophilic interaction liquid chromatography with tandem quadrupole mass spectrometer) method without derivatization. BMAA was detected in a total of five samples from three species: Neverita didyma, Solen strictus, and Mytilus coruscus. The top three concentrations of free-form BMAA (0.99~3.97 μg·g⁻¹ wet weight) were detected in N. didyma. DAB was universally detected in most of the mollusk samples (53/68) with no species-specific or regional differences (0.051~2.65 μg·g⁻¹ wet weight). No AEG was detected in any mollusk samples tested here. The results indicate that the gastropod N. didyma might be an important vector of the neurotoxin BMAA in the Chinese marine ecosystem. The neurotoxin DAB was universally present in marine bivalve and gastropod mollusks. Since N. didyma is consumed by humans, we suggest that the origin and risk of BMAA and DAB toxins in the marine ecosystem should be further investigated in the future. Full article

Exposure to β-N-methylamino-l-alanine (BMAA) might be linked to the incidence of amyotrophic lateral sclerosis, Alzheimer’s disease and Parkinson’s disease. Analytical chemistry plays a crucial role in determining human BMAA exposure and the associated health risk, but the performance of various analytical methods currently employed is rarely compared. A CYANOCOST initiated workshop was organized aimed at training scientists in BMAA analysis, creating mutual understanding and paving the way towards interlaboratory comparison exercises. During this workshop, we tested different methods (extraction followed by derivatization and liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) analysis, or directly followed by LC-MS/MS analysis) for trueness and intermediate precision. We adapted three workup methods for the underivatized analysis of animal, brain and cyanobacterial samples. Based on recovery of the internal standard D₃BMAA, the underivatized methods were accurate (mean recovery 80%) and precise (mean relative standard deviation 10%), except for the cyanobacterium Leptolyngbya. However, total BMAA concentrations in the positive controls (cycad seeds) showed higher variation (relative standard deviation 21%–32%), implying that D₃BMAA was not a good indicator for the release of BMAA from bound forms. Significant losses occurred during workup for the derivatized method, resulting in low recovery (<10%). Most BMAA was found in a trichloroacetic acid soluble, bound form and we recommend including this fraction during analysis. Full article

A cluster of amyotrophic lateral sclerosis (ALS) has been previously described to border Lake Mascoma in Enfield, NH, with an incidence of ALS approximating 25 times expected. We hypothesize a possible association with cyanobacterial blooms that can produce β-N-methylamino-L-alanine (BMAA), a neurotoxic amino acid implicated as a possible cause of ALS/PDC in Guam. Muscle, liver, and brain tissue samples from a Lake Mascoma carp, as well as filtered aerosol samples, were analyzed for microcystins (MC), free and protein-bound BMAA, and the BMAA isomers 2,4-diaminobutyric acid (DAB) and N-(2-aminoethyl)glycine (AEG). In carp brain, BMAA and DAB concentrations were 0.043 μg/g ± 0.02 SD and 0.01 μg/g ± 0.002 SD respectively. In carp liver and muscle, the BMAA concentrations were 1.28 μg/g and 1.27 μg/g respectively, and DAB was not detected. BMAA was detected in the air filters, as were the isomers DAB and AEG. These results demonstrate that a putative cause for ALS, BMAA, exists in an environment that has a documented cluster of ALS. Although cause and effect have not been demonstrated, our observations and measurements strengthen the association. Full article