Microcystin Toxins at Potentially Hazardous Levels in Algal Dietary Supplements Revealed by a Combination of Bioassay, Immunoassay, and Mass Spectrometric Methods.

Microcystins (MCs) are hepatotoxic heptapeptides produced by cyanobacteria and are potent inhibitors of protein phosphatases in eukaryotic cells. Algae for dietary supplements are harvested from outdoor environments and can be contaminated with MCs. Monitoring of MCs in these products is necessary but is complicated by their structural diversity (>250 congeners). We used a combination of protein phosphatase inhibition assay (PPIA), ELISA, LC-MS/MS, and nontargeted LC-high-resolution MS (LC-HRMS) with thiol derivatization to characterize the total MCs in 18 algal dietary supplements. LC-MS/MS revealed that some products contained >40 times the maximum acceptable concentration (MAC) of 1 µg/g MCs, but ELISA and PPIA showed up to 50-60 times the MAC. LC-HRMS identified all congeners targeted by LC-MS/MS plus MC-(H4)YR contributing up to 18% of total MCs, along with numerous minor MCs. Recommended dosages of the products greater than the MAC would result in 2.6-75 times the tolerable daily intake, presenting a risk to consumers. This study confirms the need for monitoring these products and presents strategies to fully describe the total MC pool in environmental samples and algal products.

A new benzophenanthridine alkaloid and other bioactive constituents from the stem bark of Zanthoxylum heitzii.

Heitziquinone (7), a new benzophenanthridine alkaloid, together with five known compounds; isoarnottianamide (5), rhoifoline B (6), isobauerenol (8), 6-hydroxypellitorine (9) and sylvamide (10), were isolated as minor compounds from the hexane extract of stem bark from Zanthoxylum heitzii. Four previously reported compounds (1-4) were found, as well. Compounds 5 and 7 were both found to exist as 4:1 mixtures of two atropisomers. The structures were elucidated by 1D and 2D NMR spectroscopy and by mass spectrometry. Compounds 5-10 were identified for the first time in this species, and they are all rare natural compounds. Pellitorine (4), one of the main compounds from the hexane bark extract, was found to be responsible for the brine shrimp larvae toxicity (LC50 37 µM, 8 µg/ml) of the crude extract (LC50 24 µg/ml). Low cytotoxicity against a macrophage cell line was observed.

Indole-diterpenes and ergot alkaloids in Cynodon dactylon (Bermuda grass) infected with Claviceps cynodontis from an outbreak of tremors in cattle.

Tremorgenic syndromes in mammals are commonly associated with indole-diterpenoid alkaloids of fungal origin. Cattle are sometimes affected by tremors (also called "staggers") when they graze on toxic grass pastures, and Bermuda grass ( Cynodon dactylon , kweek) has been known to be associated with tremors for several decades. This study reports the identification of paspalitrems and paspaline-like indole-diterpenes in the seedheads of Claviceps cynodontis -infected Bermuda grass collected from a pasture that had caused a staggers syndrome in cattle in South Africa and thereby links the condition to specific mycotoxins. The highest concentration (about 150 mg/kg) was found for paspalitrem B. Ergonovine and ergine (lysergic acid amide), together with their C-8 epimers, were found to co-occur with the indole-diterpenes at concentrations of about 10 microg/kg. The indole-diterpene profile of the extract from the ergotized Bermuda grass was similar to that of Claviceps paspali sclerotia. However, the C. paspali sclerotia contained in addition agroclavine and elymoclavine. This is the first study linking tremors associated with grazing of Bermuda grass to specific tremorgenic indole-diterpenoid mycotoxins.