Investigation of hepatotoxic and genotoxic potential as well as metabolism of food relvant pyrrolizidine alkaloids (PA2)

The findings of high pyrrolizidine alkaloid (PA) amounts in tea and herbal infusions and other plant derived food have to be considered as a relevant topic in food safety, especially, as some of these toxins are classified as genotoxic carcinogens. The PAs detected in different types of tea and herbal infusions belonged to different structural types. More precisely, PA amounts determined in food are the sum of a large number of individual PAs that occur in parallel. As no substantial data on the relative toxicities of PAs are available usually the sum-amounts of individual PAs were considered for risk assessment. It is questionable if this procedure is adequate and demonstrates the need for a better knowledge concerning relative PA toxicities. With an increase in the extensive production of plants for tea and herbal infusions and the intended reduction in herbicide use, natural toxins such as PAs are likely to become an increasing problem.In the past, several toxicity studies with laboratory animals were conducted for a limited number of PAs. Monitored endpoints were the acute toxic effects such as lethality or morphological changes in organs. Although hints for a structure dependent toxicity exist, no conclusions on the relative toxicities of individual PAs can be drawn due to the limited number of tested PAs and due to study designs that limited the comparability of data and the calculation of relative potencies. It is the goal of this project to apply in vitro test systems with different toxicological endpoints that enable the quantitative analysis of toxic effects of PAs. The data will be used for assessing relative PA toxicities. As PAs are pro-toxins and toxicity is caused after their metabolic activation only, there has to be a strong relationship between metabolism and toxicity. The correlation of in vitro tests and metabolism data will be used as tool for a better understanding of PA toxicity, a key to risk assessment of relative PA toxicities.

Pyrrolizidine alkaloids (PAs) are secondary plant metabolites formed by certain flowering plants in high structural diversity. They are metabolized in the liver to highly reactive metabolites, which can lead to adverse health effects in humans and animals. Due to the co-harvesting of PA containing weeds, food of plant origin, such as herbal teas and spices, may show contamination with PAs. It is likely that natural toxins such as PAs will become a challenge to crop food production due to climate change and the increase in extensive agriculture while reducing herbicide use. Combined with the desire and need to balance biodiversity and agricultural land use, it is likely that reliable data on the toxicity of individual PAs will be required for risk assessment.The aim of this project was to identify hepatically formed metabolites (in vitro incubation with liver microsomes) from humans and rats so that this process can be described as quantitatively as possible. In this project, for the first time, comprehensive analyses were performed to determine structural influences on the metabolism of PAs using 25 congeners with certified concentrations under standardized measurement conditions. From the set of 25 PAs, metabolite screening was performed for six representative PAs. The identified metabolites were classified into reactive or other metabolites based on their structures and toxic mechanisms of action reported in the literature. In this work, numerous such reactive, so-called pyrrolic metabolites, could be identified and structurally described in more detail. It was shown that the formation potential for reactive metabolites varies between the individual PAs and correlates with the observed toxic effect. The structural features that correlated with a high formation potential in the studies in this project can be summarized as follows: Open-chain and cyclic diesters of the heliotridine and retronecinol type substituted at the C7 position with angelic acid (including epoxide) exhibited high formation potential. For example, PAs such as lasiocarpine, echimedine, senecionine, retrorsine, riddelliine, jacobine belong to this group.Accordingly, low formation potential showed PAs that do not have this structure: PAs that are not 1,2 unsaturated, such as platyphylline.monoesters, such as lycopsamine, intermedin open-chain diesters substituted at C7 with short-chain necine acids, such as acetic acid, e.g. acetyllycopsaminecyclic diesters that lack the angelic acid motif, such as jacolin. The data generated in this research project suggest that PAs with a lower formation potential also have a lower toxic effect potential. Investigating this hint in more detail would be relevant, for example, in light of the following:In nature, plants occur that almost exclusively form monoesters or occur in association with open-chain diesters substituted at C7 with short-chain necic acids, such as acetic acid. Here, native plants such as Eupatorium or Borago may be mentioned as representatives. Against the above background, the toxic potential of such PA plants would have to be reconsidered, both from the point of view of consumer health protection and the protection of biodiversity.