Mc wr

Manufactured by Enzo Life Sciences

Sourced in United Kingdom

The MC-WR is a laboratory centrifuge designed for general-purpose applications. It features a robust construction and can accommodate different rotor types to accommodate a variety of sample sizes and volumes. The centrifuge provides reliable performance and consistent results.

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7 protocols using mc wr

Microcystin Toxin Analysis in Water Samples

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In triplicate, 200 mL of water sample was filtered (CFC, Whatman, Maidstone, UK), filter papers were wrapped individually in aluminium foil and preserved at −80 °C. On analysis, filter papers were subjected to three cycles of freeze-thawing before submersion in 10 mL of 80% aqueous methanol. Samples were left in the dark at 4–6 °C for 24 h, before ~0.5 mL was aliquoted into a LCMS certified vial. Toxin analysis was carried by ultra-high-performance liquid chromatography (UHPLC) (Acquity, Waters, Manchester, UK) coupled to a tandem quadruple mass spectrometer (Xevo TQ, Waters, Manchester, UK). All instrument solvents and chemicals were of LC-MS-grade (Fisher Optima, Thermo Fisher, Manchester, UK). Reference toxins used for the detection method included the microcystin analogues MC-RR, MC-LA, MC-LY, MC-LF, MC-LW, MC-YR, MC-WR, MC-HilR, MC-HtyR, MC-LR & Asp³-MC-LR (Enzo Life Sciences, Exeter, UK) and [Dha⁷]-MC-LR and matrix reference material of blue-green algae (RM-BGA, Lot 201301) containing a range of microcystins (Institute of Biotoxin Metrology, National Research Council Canada). Analysis of microcystins was conducted following the method by Turner et al. [50 (link)].

Hartnell D.M., Chapman I.J., Taylor N.G., Esteban G.F., Turner A.D, & Franklin D.J. (2020). Cyanobacterial Abundance and Microcystin Profiles in Two Southern British Lakes: The Importance of Abiotic and Biotic Interactions. Toxins, 12(8), 503.

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Quantitative Analysis of Cyanotoxin Standards

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Mobile phases were prepared from LC-MS-grade acetonitrile (Fisher Optima, ThermoFisher, Greater London, UK) and water used for LC-MS was obtained in-house. Sample preparation reagents were HPLC grade. Toxin standards used for preparation of calibration solutions (MC-RR, MC-LA, MC-LY, MC-LF, MC-LW, MC-YR, MC-WR, MC-HilR, MC-HtyR, MC-LR, [Asp3] MC-LR and Nodularin) were all obtained from Enzo Life Sciences, Exeter, UK. A certified standard of [Dha7] MC-LR was obtained from the Institute of Biotoxin Metrology, National Research Council Canada (NRCC, Halifax, NS, Canada). Reference standards received as solid films were dissolved in 50% aqueous methanol, to form stock solutions. A mixed stock solution was subsequently prepared by combining aliquots of each stock, followed by further dilutions to create seven-level suite of working calibration standards between 0.33 ng/mL to 327 ng/mL per toxin.

Turner A.D., Dhanji-Rapkova M., O’Neill A., Coates L., Lewis A, & Lewis K. (2018). Analysis of Microcystins in Cyanobacterial Blooms from Freshwater Bodies in England. Toxins, 10(1), 39.

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Extraction and Preparation of Cyanotoxin Standards

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The solvents for the extraction and for the basis of the mobile phase were UHPLC-MS grade. All toxin standards for MCs (MC-LR, MC-RR, MC-LA, MC-LY, MC-LF, MC-LW, MC-YR, MC-WR) and NOD came from Enzo Life Sciences^® (Antwerp, Belgium) and were received under the form of a solid powder. After dilution in 100% methanol (MeOH), mixed stock solutions were prepared in 50% methanol (MeOH) (50% Milli-Q water with 1% acetic acid (v/v)). The stock and the intermediate solutions were stored at −20 °C.

Van Hassel W.H., Ahn A.C., Huybrechts B., Masquelier J., Wilmotte A, & Andjelkovic M. (2022). LC-MS/MS Validation and Quantification of Cyanotoxins in Algal Food Supplements from the Belgium Market and Their Molecular Origins. Toxins, 14(8), 513.

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Comprehensive Microcystin Quantification

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LC-MS grade acetonitrile, water and formic acid and HPLC-grade methanol and water were purchased from Fisher (ThermoFisher, UK). Reference toxin standards of microcystins (MC-LR, MC-RR, MC-YR, MC-WR, MC-LW, MC-LA, MC-LY, MC-LF, MC-HtyR, MC-HiLR, [Asp3]-MC-LR/[Dha7]-MC-LR) and NOD ≥95% were as per Enzo Life Sciences (Exeter, UK) (Fig. S1). A certified standard of [Dha7]-MC-LR and a pre-certified freeze-dried matrix reference material of blue-green algae (RM-BGA, Lot 201301) containing a range of MCs were purchased from the Institute of Biotoxin Metrology, National Research Council Canada (Ontario, Canada).
A mixed stock solution was prepared by combining aliquots of each toxin to give a final concentration of 327 μg/L. For external calibration, a seven point calibration curve was prepared by serial dilution with methanol/water (1:1, v/v) in the range of 0.33–327 μg/L for each toxin and stored at – 18 °C. A quality control reference material (RM-BGA, National Research Council, Halifax, Canada) was prepared, with toxins extracted in the supernatant after 28 mL of methanol/water (1:1, v/v) + 0.1% acetic acid were added to RM-BGA (280 mg) and subsequent centrifugation (4500

\times

g; 10 min).
Shellfish diet 1800 (approximately 7.4 × 10¹¹ cells/mL) was purchased from ReedMariculture Inc., (US) and dilutions were made in water/seawater (10:0.86, v/v).

Camacho-Muñoz D., Waack J., Turner A.D., Lewis A.M., Lawton L.A, & Edwards C. (2021). Rapid uptake and slow depuration: Health risks following cyanotoxin accumulation in mussels?. Environmental Pollution (Barking, Essex : 1987), 271, 116400.

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Comprehensive Cyanotoxin Analysis in Water

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We measured toxins in 246 whole-water samples spanning 6 y (2013 to 2018). These samples were taken from the same water grabs used to characterize the microbial community. We quantified cyanotoxins using methods described in detail by Miller et al. (16 (link)). Briefly, lyophilized samples were resuspended in formic acid, subjected to three freeze-thaw cycles, and extracted in methanol and formic acid in a sonicating water bath. The extract supernatant was analyzed by targeted high-performance liquid chromatography coupled with tandem mass spectrometry (HPLC-MS/MS).
Eleven microcystin (MC) congeners and three anabaenopeptin (Apt) congeners were analyzed. Certified reference standards of MC-LR, [Dha⁷]MC-LR and nodularin were purchased from the National Research Council of Canada Biotoxins program (Halifax, Nova Scotia). MC-LA, MC-RR, MC-LF, MC-YR, MC-WR, MC-LY, MC-LW, MC-HtyR, and MC-HilR (all >95%) were purchased from Enzo Life Sciences (Farmingdale, NY). Anabaenopeptin A (Apt-A) (>95%), Apt-B (>95%), and Apt-F (>95%) were purchased from MARBIONC (Wilmington, NC). To calculate total concentrations, we summed nanomolar measurements of the congeners. 99 ± 4% of total microcystin was comprised of the congeners MC-LA and MC-LR. Toxin totals are available in a tabular format as Dataset S1.

Rohwer R.R., Hale R.J., Vander Zanden M.J., Miller T.R, & McMahon K.D. (2023). Species invasions shift microbial phenology in a two-decade freshwater time series. Proceedings of the National Academy of Sciences of the United States of America, 120(11), e2211796120.

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Accurate Quantification of Cyanotoxins

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Whenever possible, certified reference standards were used. Nodularin, MCLR and dmMCLR were certified reference materials from the National Research Council of Canada Biotoxins program (Halifax, Nova Scotia). Microcystin standards – MCLA (> 95%), MCRR (> 90%), and MCYR (> 90%) were purchased from Sigma-Aldrich (Milwaukee, WI) and MCLF (> 95%), MCLY (> 95%), MCWR (> 95%), MCLW (>95%), MCHtyR (> 95%), (> 95%), and MCHilR (> 95%) were purchased from Enzo Life Sciences (Farmington, NY, USA). AptA (> 95%), B (> 95%) and F (> 95%), Cpt1007 (> 95%), 1020 (> 95%), and 1041 (> 95%), and Mgn690 (> 95%) were purchased from MARBIONC (Wilmington, NC, USA).

Bartlett S.L., Brunner S.L., Klump J.V., Houghton E.M, & Miller T.R. (2018). Spatial analysis of toxic or otherwise bioactive cyanobacterial peptides in Green Bay, Lake Michigan. Journal of Great Lakes research, 44(5), 924-933.

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Cyanotoxin Quantification Protocol

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Instrument solvents used for preparation of mobile phases were of LC-MS-grade (Fisher Optima, ThermoFisher, UK) and all chemicals were LC-MS reagent grade where possible. Sample preparation reagents were HPLC grade. Reference toxin standards (MC-RR, MC-LA, MC-LY, MC-LF, MC-LW, MC-YR, MC-WR, MC-HilR, MC-HtyR, MC-LR, [Asp3] MC-LR and Nod) were all obtained from Enzo Life Sciences, Exeter, UK. A certified standard of [Dha 7 ]-MC-LR and a pre-certified freeze-dried matrix reference material of blue-green algae (RM-BGA, Lot 201301) containing a range of MC was obtained from the Institute of Biotoxin Metrology, National Research Council Canada (NRCC). Reference standards received as solid powders were dissolved in suitable volumes of 50% aqueous methanol, to form stock solutions. A mixed stock solution was subsequently prepared by combining aliquots of each stock, followed by a seven-level suite of working calibration standards resulting in a calibration range between 0.33 ng/mL to 327 ng/m per toxin. RM-BGA (280 mg) was extracted with 28.0 mL 50% aqueous MeOH + 0.1% acetic acid, prior to centrifugation (4,500 g; 10 min) and the supernatant collected prior to analysis. The seven-point calibration standards were used for external calibration of cyanotoxins in all sample matrices, adjusting dilution factors depending on the extraction applied.

Turner A.D., Waack J., Lewis A., Edwards C, & Lawton L. (2018). Development and single-laboratory validation of a UHPLC-MS/MS method for quantitation of microcystins and nodularin in natural water, cyanobacteria, shellfish and algal supplement tablet powders. Journal of chromatography. B, Analytical technologies in the biomedical and life sciences, 1074-1075.

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