Structure-dependent cytotoxicity of different pyrrolizidine alkaloids in primary rat hepatocytes

Background:

Pyrrolizidine alkaloids (PAs) are secondary metabolites occurring in a wide range of plant species. Some 1,2-unsaturated PAs exert toxic effects through metabolic activation which form the corresponding dehydropyrrolizidine derivatives, primarily in the liver, catalyzed by cytochrome P450 monooxygenases. Due to their hepatotoxicity, genotoxicity and carcinogenicity, the accidental presence of PAs in food, feed and herbal medicinal products can be, depending on the dose, a cause for safety concerns.

Objectives:

In order to assess potential risks and confirm the connection between structure and in vitro toxicity [1], we generate data firstly concerning cytotoxicity of some food-relevant PAs. In particular, we wanted to expand the database for the derivation of congener-specific relative potency factors to take into account the widely differing toxic potencies among PAs.

Methods:

After 24h and 48h exposure, cytotoxicity of the selected PAs was determined at concentrations ranging from 1 to 300µM by the Alamar blue assay in primary rat hepatocytes. A kinetic assay analyzing 7-benzyloxyresorufin-O-dealkylation (BROD) was used for measuring the activity of CYP enzymes.

Results:

It was found that cyclic and open di-esters were much more toxic than mono-esters. Levels of BROD activity, decreasing over time in culture, played an important role for activation of PA. With a highly toxic PA (lasiocarpine), inhibition of CYP with ketoconazole markedly reduced toxicity while this was not obvious with the less toxic congener lycopsamine. Overall, our data [2] mostly confirm previously published interim Relative Potency (iREP) factors although for echimidine and monocrotaline substantial deviations were found, possibly due to specific toxicokinetic properties of these congeners in vivo.

Acknowledgement:

We would like to thank the DFG and the Kooperation Phytopharmaka, Bonn for their financial support of this study.

References:

[1] Merz KH, Schrenk D. Toxicol Lett 2016; 263: 44 – 57

[2] Gao L et al. Food Chem Toxicol 2019, in press