Pyrrolizidine alkaloid

Pyrrolizidine alkaloids (PAs), sometimes referred to as necine bases, are a group of naturally occurring alkaloids based on the structure of pyrrolizidine. Their use dates back centuries and is intertwined with the discovery, understanding, and eventual recognition of their toxicity on humans and other animals.

History PAs were first discovered in plants in the 19th century, but their toxic effects were not immediately recognized. Instead, many PA-containing plants were traditionally used for medicinal purposes in various cultures around the world. For example, herbs containing PAs were used in traditional Chinese medicine and by Native American tribes for their purported therapeutic properties. It has been estimated that 3% of the world's flowering plants contain pyrrolizidine alkaloids. Honey can contain pyrrolizidine alkaloids, as can grains, milk, offal and eggs. To date (2011), there is no international regulation of PAs in food, unlike those for herbs and medicines. In the early to mid-20th century, researchers began to observe and document cases of livestock poisoning linked to the consumption of PA-containing plants. These observations led to the recognition of PAs as potent hepatotoxic and genotoxic compounds. In response to growing concerns about PA exposure, regulatory agencies around the world began to establish guidelines and regulations to limit PA levels in food, herbal products, and animal feed. These regulations aim to protect human and animal health by minimizing PA exposure and mitigating the risk of toxicity. Despite regulatory efforts, the issue of PA exposure remains relevant today. Ongoing research continues to explore various aspects of PA toxicity, including the identification of new PA-containing plants, the development of sensitive analytical methods, and the assessment of human health risks associated with PA exposure. Additionally, efforts to raise awareness among healthcare professionals, herbal product manufacturers, and the general public about the risks of PA exposure are ongoing.

Natural occurrence

PAs are a group of naturally occurring compounds found in a wide range of plant species. These alkaloids are secondary metabolites synthesized by plants primarily as a defense mechanism against herbivores, insects, and pathogens. The biosynthesis of PAs was discovered to occur through the first pathway-specific enzyme homospermidine synthase.

The polyamines putrescine and spermidine are derived from the basic amino acid arginine. Subsequently, homospermidine synthase exchanges the 1,3-diaminopropane by putrescine and forms symmetric homospermidine. Oxidation of homospermidine by copper-dependent diamine oxidases initiates cyclization to pyrrolizidine-1-carbaldehyde, which is reduced, to 1-hydroxymethylpyrrolizidine. Desaturation and hydroxylation ultimately form retronecine, which is acylated with an activated necic acid, for instance with senecyl-CoA2 as in the example shown below. PAs are preferably found in the plant families Asteraceae (tribes Eupatorieae and Senecioneae), Boraginaceae (many genera), Fabaceae (mainly the genus Crotalaria), and Orchidaceae (nine genera). More than 95% of the PA-containing species investigated thus far belong to these four families.

Structure and reactivity

PAs are compounds made up of a necine base, a double five-membered ring with a nitrogen atom in the middle, and one or two carboxylic esters called necic acids. Four major necine bases are described, with retronecine and its enantiomer heliotridine being the largest group, and highly toxic. Another group is the platynecine, the difference between these groups is its saturated base, which makes it less toxic. Most bases have a 1,2-unsaturated base. Another difference in the groups is with otonecine, which cannot form N-oxides, due to the methylation of the nitrogen atom.

The alcohol groups on the necine bases can make esters in a wide variety of forms. Among the possibilities are mono-esters, like Floridine and Heliotrine, and di-esters either with an open or closed ring structure, like Usaramine and Lasiocarpine. In total more than 660 PAs and PA N-oxides have been identified in over 6000 plants.

Synthesis There are multiple ways to synthesize PAs and their derivatives. A flexible strategy would be to start with a Boc (tert-butoxycarbonyl) protected pyrrole molecule and use specific reaction for synthesis into the desired compound.

Mechanisms of actions and metabolism PAs are commonly introduced into the body via oral ingestion through contaminated food or traditional medicine, notably borage leaf, comfrey and coltsfoot. It can readily form salts with nitrates, chlorides and sulphates, which facilitate the uptake in the gastrointestinal tract. After which they travel to the liver via the portal vein.

Metabolites form mostly in the liver. Here esterases can hydrolyze the PAs to reduce the compound into its necine acids and bases, both forms are non-toxic for humans and do not damage the body. However, cytochrome P450 (CYP450) also metabolizes PAs, this enzyme can form pyrrolic esters (EPy), these are hepatotoxic due to their high reactivity. The EPy can also be hydrolyzed into alcoholic pyrroles, which are mutagenic and carcinogenic. Since this mostly happens in the liver, this is the most affected organ. Other affected organs are the lungs and kidneys. The EPy can escape the liver, and travel through the Disse space into the bloodstream. The electrophilic nature of pyrroles makes them an easy target for nucleophilic attack from nucleic acids and protein. If bound by glutathione they can become a non-toxic conjugate and be excreted via the kidneys. A second detoxifying pathway is the formation of the N-oxide In the liver and lungs of certain mammal species enzymes called monooxygenase can prevent aromatization of the double 5-ring and in turn prevent the formation of the pyrrole-protein adduct.

Toxicological effects The toxicity consequences resulting from the metabolism of PAs in humans primarily revolve around hepatotoxicity and genotoxicity.

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