Detection of the tremorgenic mycotoxin paxilline and its desoxy analog in ergot of rye and barley: a new class of mycotoxins added to an old problem

Abstract

A newly developed enzyme immunoassay (EIA) for the detection of the tremorgenic indole-diterpene alkaloid paxilline (PAX) and closely related analogs was used to analyze ergot sclerotia collected from rye and barley fields. The mean EIA standard curve detection limit was 0.47 ± 0.14 ng/mL; relative cross-reactivity of toxin standard solutions was found for 11-hydroxy-paspaline (terpendole E, 1.1%) but not for lolitrem B or ergot alkaloids. Sclerotia from all fields were positive in the PAX-EIA at concentration levels of 620 ± 200 and 160 ± 37 μg/kg in ergot of rye and 130 ± 47 μg/kg in ergot of barley. Confirmatory analyses of sclerotia by liquid chromatography-tandem mass spectrometric detection identified PAX and its analog 13-desoxypaxilline. To the best of our knowledge, this is the first report on the natural occurrence of tremorgenic indole-diterpene alkaloid mycotoxins in ergot sclerotia from rye and barley. Along with details on the analytical methodology developed in this study, particularly PAX-antibody production, the relevance and implications of these findings for food and feed safety are discussed. Presence or absence of elevated levels of tremorgenic mycotoxins, along with the ergot alkaloids, would help in explaining the difference between the two distinct manifestations of historic ergotism, the convulsive and the gangrenous form. Further method development for paxilline and other tremorgenic mycotoxins in cereals used for food and feed is a prerequisite for a comprehensive risk assessment, which seems to be necessary in light of the findings reported here. Paxilline in ergot of rye.

Alkaloid Concentrations of Lolium perenne Infected with Epichloë festucae var. lolii with Different Detection Methods—A Re-Evaluation of Intoxication Risk in Germany?

“…It can be assumed that other methods of alkaloid detection differ in their absolute amounts of alkaloid concentrations and can only be compared relatively. Bauer et al (2018), for example, showed that paxilline concentrations detected with HPLC-MS/MS were less than 3% of the paxilline level detected with enzyme immunoassay [35], but this might be explained due to the often unselective detection of immunoassays [60]. Hence, only validated methods should be used for the detection of alkaloids to compare alkaloid concentrations with other studies and toxicity thresholds from literature.…”

“…However, methodologies differ substantially between studies. Most studies used high-performance liquid chromatography (HPLC) with different detection methods like mass spectrometry [30], tandem mass spectrometry [24,31,32], or fluorescence detection [16,20,21,33,34] for the quantitation of lolitrem B, paxilline, ergovaline, and peramine, while one study used enzyme immunoassays [35]. Analytical reference substances are used for the validation of analytical method or as external standard for quantitation of alkaloids (e.g., lolitrem B in [20,21]) but are rarely commercially available.…”

Tremorgenic and neurotoxic paspaline-derived indole-diterpenes: biosynthetic diversity, threats and applications

“…Tellingly, the paspalitrem producer C. paspali was isolated from the unwholesome grain (Aaronson 1988). Even today, another IDT producer, C. purpurea is often isolated from rye and barley, and paxilline can be detected in such specimen at the maximum concentration of 0.6 mg/kg (Bauer et al, 2017). C. purpurea sclerotia contain at least seven paspalenes, including paspaline (Uhlig et al 2014).…”

“…This raises the interesting possibility that these toxins may have played a role in the development of the feared symptoms of ergotism in the Middle Ages. Some authors, including Bauer and coworkers hypothesized that IDTs might have contributed to outbreaks of convulsive ergotism during history (Bauer et al 2017). …”

Ergochromes: Heretofore Neglected Side of Ergot Toxicity

“…He noted the complexity of this area of research as EA toxicity is affected by changes in alkaloid concentrations, proportions, and availability as well as individual’s genetic predispositions, prior exposures, and ambient environments. This led to the overall conclusion that the impacts of EAs on livestock, especially convulsive (neurological and abortogenic) symptoms are not caused by the sole action of a single toxin, but rather the combined impact and synergistic action of multiple EAs derived from Claviceps and Epichloë species [9,10]. While most of the available data of EAs effects on animals both address the symptoms and define the problem, there is still a lack of research on other fungal metabolites and their potential harmful effects on livestock and humans.…”