otonecine and retronecine

Pyrrolizidine alkaloids (PAs) are natural toxins found in about 3%-5% of flowering plants. Dehydropyrrolizidine alkaloids contain a double bond in 1, 2-position of the necine bases, including retronecine type PAs (RET-PAs) and their N-oxides (RET N-oxide-PAs), and otonecine type PAs (OTO-PAs), and are known for their significant hepatotoxicity. Most dehydropyrrolizidine alkaloids are metabolically activated by cytochrome P450 (CYP450) enzymes to generate active pyrroles, which further bind to proteins to form pyrrole-protein adducts (PPAs). Methods for predicting PA-induced liver injury are generally performed on in vitro models with extremely low activities of CYP450 enzymes, which is different from the situation in vivo. In this regard, primary cultured mouse hepatocytes, which showed comparable CYP450 activity with the in vivo models, were applied to illustrate the structure-toxicity relationship of 13 dehydropyrrolizidine alkaloids, namely, eight RET-PAs, three RET N-oxide-PAs, and two OTO-PAs. PA-induced cytotoxicity and PA-generated PPAs were analyzed in primary mouse hepatocytes treated with different PAs. Results showed that PA-induced toxicity was correlated with the amount of PA-generated PPAs. RET-PAs and OTO-PAs were generally more toxic than RET N-oxide-PAs and generated higher amount of PPAs. PPAs were utilized to evaluate the efficiency of metabolic activation and predict the toxic potencies of dehydropyrrolizidine alkaloids. The proposed model could be a new approach for toxicity evaluation and risk control of exposure to PAs.

Topics: Activation, Metabolic; Alkaloids; Animals; Hepatocytes; Mice; Proteins; Pyrrolizidine Alkaloids

Pyrrolizidine alkaloids (PAs) are characteristic metabolites of some plant families and form a powerful defense mechanism against herbivores. More than 600 different PAs are known. PAs are ester alkaloids composed of a necine base and a necic acid, which can be used to divide PAs in different structural subcategories. The main target organs for PA metabolism and toxicity are liver and lungs. Additionally, PAs are potentially genotoxic, carcinogenic and exhibit developmental toxicity. Only for very few PAs, in vitro and in vivo investigations have characterized their toxic potential. However, these investigations suggest that structural differences have an influence on the toxicity of single PAs. To investigate this structural relationship for a large number of PAs, a quantitative structural-activity relationship (QSAR) analysis for hepatotoxicity of over 600 different PAs was performed, using Random Forest- and artificial Neural Networks-algorithms. These models were trained with a recently established dataset specific for acute hepatotoxicity in humans. Using this dataset, a set of molecular predictors was identified to predict the hepatotoxic potential of each compound in validated QSAR models. Based on these models, the hepatotoxic potential of the 602 PAs was predicted and the following hepatotoxic rank order in 3 main categories defined (1) for necine base: otonecine > retronecine > platynecine; (2) for necine base modification: dehydropyrrolizidine ≫ tertiary PA = N-oxide; and (3) for necic acid: macrocyclic diester ≥ open-ring diester > monoester. A further analysis with combined structural features revealed that necic acid has a higher influence on the acute hepatotoxicity than the necine base.

Topics: Algorithms; Chemical and Drug Induced Liver Injury; Cyclic N-Oxides; Databases, Factual; Dicarboxylic Acids; Heterocyclic Compounds, 2-Ring; Humans; Liver; Machine Learning; Molecular Structure; Neural Networks, Computer; Pyrrolizidine Alkaloids; Quantitative Structure-Activity Relationship; Reproducibility of Results; Risk Assessment

A rapid, but sensitive and selective method for simultaneous screening and quantification of toxic pyrrolizidine alkaloids (PAs) by ultra performance liquid-chromatography (UPLC) coupled with tandem mass spectrometry (MS/MS) on a tandem quadrupole mass spectrometer (TQ-MS) is described. This was accomplished by incorporating the precursor ion scan (PIS) acquisition and multiple reaction monitoring (MRM) acquisition in the same UPLC-MS/MS run. Notably, the developed PIS approach for detecting two pairs of characteristic product ions at m/z 120/138 or 168/150, allowed specific identification of toxic retronecine and otonecine types PAs. This PIS method is highly sensitive with over 10-fold sensitivity improvement upon previously published LC-MS method. Moreover, this new approach is suitable for high sample throughput and was applied to the screening and quantifying toxic PAs in 22 samples collected from seven Parasenecio species and four Senecio species. In addition, coupling the MRM with PIS approach generated quantitative results equivalent to those obtained by conventional MRM-only approach. This coupled MRM with PIS approach could provide both qualitative and quantitative results without the need of repetitive analyses.

Topics: Asteraceae; Chromatography, Liquid; Pyrrolizidine Alkaloids; Senecio; Tandem Mass Spectrometry

To detect the hepatotoxic pyrrolizidine alkaloids (HPA) in the genus Ligularia Cass... The alkaloid extracts of Ligularia plant materials were detected and analyzed by the method of combination of TLC, and LC/MSn.. Among 22 species of Ligularia Cass., HPA were detected in 18 species with LC/MSn, and no HPA was detected in the remaining 4 species.. HPA was first detected with LC/MSn in L. tongelensis and other 15 species of Ligularia Cass.; HPA from these plants should be isolated, separated and identified and it is necessary to study the activities and toxicities of the HPA. The types and kinds of HPA from different species and sources are different, they should be detected separately.

Topics: Asteraceae; Chromatography, Thin Layer; Molecular Structure; Plants, Medicinal; Pyrrolizidine Alkaloids; Species Specificity; Spectrometry, Mass, Electrospray Ionization

Clivorine (1) and ligularine (2), two hepatotoxic otonecine-type pyrrolizidine alkaloids isolated from Ligularia hodgsonii, an antitussive traditional Chinese medicine, were investigated in CDCl(3) and D(2)O by various NMR techniques to delineate why this type of alkaloid displays uncharacteristic solubility properties by dissolving in both nonpolar organic and aqueous solutions. The results demonstrated that both alkaloids exist in a non-ionized form in CDCl(3), but in an ionized form in D(2)O, suggesting that this unique dual solubility may play a role in the intoxication resultant from consumption of water extracts of herbs, including herbal teas, containing otonecine-type pyrrolizidine alkaloids.

Topics: Antitussive Agents; Asteraceae; Carcinogens; Magnetic Resonance Spectroscopy; Medicine, Chinese Traditional; Phytotherapy; Pyrrolizidine Alkaloids; Solubility; Tea