Botanical-derived dietary supplements have widespread use in the general population. The complex and variable nature of botanical ingredients and reports of adverse responses have led to concern for negative human health impacts following consumption of these products. Toxicity testing of the vast number of available products, formulations, and combinations is not feasible due to the time and resource intensive nature of comprehensive testing. Methods are needed to assess the safety of a large number of products via more efficient frameworks. Identification of toxicologically-active constituents is one approach being used, with many advantages toward product regulation. Bioassay-guided fractionation (BGF) is the leading approach used to identify biologically-active constituents. Most BGF studies with botanicals focus on identifying pharmacologically-active constituents for drug discovery or botanical efficacy research. Here, we explore BGF in a toxicological context, drawing from both efficacy and poisonous plant research. Limitations of BGF, including loss of mixture activity and bias toward abundant constituents, and recent advancements in the field (e.g., biochemometrics) are discussed from a toxicological perspective. Identification of active constituents will allow better monitoring of market products for known toxicologically-active constituents, as well as surveying human exposure, two important steps to ensuring the safety of botanical dietary supplements.

ABSTRACT Primaquine (PQ) remains the sole available drug to prevent relapse of Plasmodium vivax malaria more than 60 years after licensure. While this drug was administered as a racemic mixture, prior studies suggested a pharmacodynamic advantage based on differential antirelapse activity and/or toxicities of its enantiomers. Oral primaquine enantiomers prepared using a novel, easily scalable method were given for 7 days to healthy rhesus macaques in a dose-rising fashion to evaluate their effects on the blood, liver, and kidneys. The enantiomers were then administered to Plasmodium cynomolgi -infected rhesus macaques at doses of 1.3 and 0.6 mg/kg of body weight/day in combination with chloroquine. The (−)-PQ enantiomer had higher clearance and apparent volume of distribution than did (+)-PQ and was more extensively converted to the carboxy metabolite. There is evidence for differential oxidative stress with a concentration-dependent rise in methemoglobin (MetHgb) with increasing doses of (+)-PQ greater than that seen for (−)-PQ. There was a marked, reversible hepatotoxicity in 2 of 3 animals dosed with (−)-PQ at 4.5 mg/kg. (−)-PQ in combination with chloroquine was successful in preventing P. cynomolgi disease relapse at doses of 0.6 and 1.3 mg/kg/day, while 1 of 2 animals receiving (+)-PQ at 0.6 mg/kg/day relapsed. While (−)-PQ was also associated with hepatotoxicity at higher doses as seen previously, this has not been identified as a clinical concern in humans during >60 years of use. Limited evidence for increased MetHgb generation with the (+) form in the rhesus macaque model suggests that it may be possible to improve the therapeutic window for hematologic toxicity in the clinic by separating primaquine into its enantiomers.