The integrated use of in silico methods for the hepatotoxicity potential of Piper methysticum.

Herbal products as supplements and therapeutic intervention have been used for centuries. However, their toxicities are not completely evaluated and the mechanisms are not clearly understood. Dried rhizome of the plant kava (Piper methysticum) is used for its anxiolytic, and sedative effects. The drug is also known for its hepatotoxicity potential. Major constituents of the plant were identified as kavalactones, alkaloids and chalcones in previous studies. Kava hepatotoxicity mechanism and the constituent that causes the toxicity have been debated for decades. In this paper, we illustrated the use of computational tools for the hepatotoxicity of kava constituents. The proposed mechanisms and major constituents that are most probably responsible for the toxicity have been scrutinized. According to the experimental and prediction results, the kava constituents play a substantial role in hepatotoxicity by some means or other via glutathione depletion, CYP inhibition, reactive metabolite formation, mitochondrial toxicity and cyclooxygenase activity. Some of the constituents, which have not been tested yet, were predicted to involve mitochondrial membrane potential, caspase-3 stimulation, and AhR activity. Since Nrf2 activation could be favorable for prevention of hepatotoxicity, we also suggest that these compounds should undergo testing given that they were predicted not to be activating Nrf2. Among the major constituents, alkaloids appear to be the least studied and the least toxic group in general. The outcomes of the study could help to appreciate the mechanisms and to prioritize the kava constituents for further testing.

Liver transplantation and the use of KAVA: Case report.

BACKGROUND: Self-medication and the belief that herbal products are free of health risks are common in Brazil. The kava (Piper methysticum), known for its anxiolytic action, has a widespread popular use. Hepatotoxicity of kava is reported, including cases of liver transplantation and death. The kava had its use prohibited or restricted in countries like Germany, France, among others. Toxicity may be related to overdosage; however, factors such as botanical characteristics of the plant, the harvesting, storage, and production process may be associated with the development of hepatotoxic substances, such as triggering idiosyncratic reactions. HYPOTHESIS: In this case, there is a suspicion that the toxicide is intrinsic to the drug; however, the possibility of adulterants and contaminants must be ruled out. STUDY DESIGN: This study reports the case of a patient who, after using the herbal kava for 52 days, evolved into acute liver failure and liver transplantation. METHODS: The data were collected directly with the patient and compared with their clinical records. Causality was determined through the RUCAM algorithm. In addition, a phytochemical analysis of the drug used was performed. RESULTS: According to the patient's report, there is no evidence of overdosage. Results from RUCAM algorithm infer causality between liver damage and the use of kava. The analysis chemical constituents did not find any possible contaminants and major changes in the active compounds. Seven months after transplantation, the patient is well and continues to be followed up by a medical team. CONCLUSION: Our investigation indicates that there was kava-induced hepatotoxicity at standard dosages. In Brazil, self-medication by herbal medicines is frequent and many patients and health professionals do not know the risks associated with their use. Diagnosing and notifying cases in which plants and herbal medicine induce liver damage is of paramount importance to increase the knowledge about DILI and to prevent or treat similar cases quickly.

Chemical Exacerbation of Light-induced Retinal Degeneration in F344/N Rats in National Toxicology Program Rodent Bioassays.

Retinal degeneration due to chronic ambient light exposure is a common spontaneous age-related finding in albino rats, but it can also be related to exposures associated with environmental chemicals and drugs. Typically, light-induced retinal degeneration has a central/hemispherical localization whereas chemical-induced retinal degeneration has a diffuse localization. This study was conducted to identify and characterize treatment-related retinal degeneration in National Toxicology Program rodent bioassays. A total of 3 chronic bioassays in F344/N rats (but not in B6C3F1/N mice) were identified that had treatment-related increases in retinal degeneration (kava kava extract, acrylamide, and leucomalachite green). A retrospective light microscopic evaluation of the retinas from rats in these 3 studies showed a dose-related increase in the frequencies of retinal degeneration, beginning with the loss of photoreceptor cells, followed by the inner nuclear layer cells. These dose-related increased frequencies of degenerative retinal lesions localized within the central/hemispherical region are suggestive of exacerbation of light-induced retinal degeneration.

Contemporary Pacific and Western perspectives on `awa (Piper methysticum) toxicology.

In 2010, a National Science Foundation project in Hawai`i assembled a collaboration of Pacific indigenous scientists, Hawaiian cultural practitioners and scientists trained in Western pharmacology. The objective of the collaborative project was to study Kava, a culturally significant Pacific beverage, and to address and ultimately transcend, long-standing barriers to communication and collaboration between these groups. Kava is a product of the `awa plant (Piper methysticum) that has been used ceremonially and medicinally throughout the history of Pacific Island cultures, and is now in widespread recreational and nutraceutical use in the US. This project, culminating in 2015, has enriched the participants, led to published work that integrates cultural and Western pharmacologic perspectives and established a paradigm for collaboration. This review paper integrates cultural and Western perspectives on efficacy, toxicity and the future cultural and commercial significance of `awa in the Pacific. Here we present a detailed review of traditional and non-traditional kava usage, medicinal efficacy and potential toxicological concerns. Recent mechanistic data on physiological action and potential pathological reactions are evaluated and interpreted.

Contaminant hepatotoxins as culprits for kava hepatotoxicity--fact or fiction?

The culprit of kava hepatotoxicity will continue to remain a mystery in humans, if the underlying reaction is of idiosyncratic, unpredictable, and dose-independent nature due potentially to some metabolic aberration in a few individuals emerging from kava use. In addition, kava hepatotoxicity is presently not reproducible experimentally in preclinical models, as demonstrated by studies showing whole kava extracts are not hepatotoxic. This led us to propose our 'working hypothesis' that contaminant hepatotoxins including moulds might have caused rare kava hepatotoxicity in humans. Further studies are now warranted to proof or disproof our working hypothesis, because kava hepatotoxicity possibly based on contaminant hepatotoxins could be a preventable disease. In the meantime, however, for minimizing toxicity risk in kava users, a pragmatic approach should focus on the medicinal use of an aqueous extract derived from peeled rhizomes and roots of a non-mouldy noble kava cultivar, limited to maximum 250-mg kavalactones daily for acute or intermittent use.

Phototoxicity of kava - formation of reactive oxygen species leading to lipid peroxidation and DNA damage.

Kava is one of the most widely sold herbal dietary supplements in the United States. It has been reported that, besides exhibiting hepatotoxicity, kava also possesses photosensitivity and induces dermopathy in humans. In this study, we determined that UVA irradiation of kava in the presence of a lipid, methyl linoleate, generated lipid peroxidation which was mediated by singlet oxygen generated during photoirradiation. The six major kavalactones(yangonin, 7,8-dihydrokawa in, kawain, 7,8-dihydromethysticin, methysticin, and 5,6-dehydrokawain) were also studied in parallel; only 5,6-dehydrokawain and yangonin-induced a low level of lipid peroxidation. UVA irradiation of kava in human HaCaT skin keratinocytes induced cytotoxicity which was mediated by oxidative stress, led to DNA strand cleavage, and produced 8-hydroxy-2'-deoxyguanosine (8-OHdG) adduct. Study by the electron spin resonance (ESR) method revealed that UVA irradiation of kava produced singlet oxygen and carbon-centered radicals. The overall results suggest that kava is photocytotoxic and photogenotoxic, both mediated by free radicals generated during photoirradiation.

Toxicology and carcinogenesis studies of kava kava extract (CAS No. 9000-38-8) in F344/N rats and B6C3F1 mice (Gavage Studies).

UNLABELLED: Kava beverages, made from dried roots of the shrub Piper methysticum, have been used ceremonially and socially in the South Pacific and in Europe since the 1700s. The drink is reported to have pleasant mild psychoactive effects, similar to alcoholic beverages. In the United States, kava kava is an herbal product used extensively as an alternative to anti-anxiety drugs such as Xanax and Valium. It has also been reported as being used to help children with hyperactivity and as a skin-conditioning agent in cosmetics. Kava kava was nominated by the National Cancer Institute for study because of its increasing use as a dietary supplement in the mainstream United States market and reports of liver toxicity among humans. Male and female F344/N rats and B6C3F1 mice received kava kava extract in corn oil by gavage for 2 weeks, 3 months, or 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium, Escherichia coli, and mouse peripheral blood erythrocytes. 2-WEEK STUDY IN RATS: Groups of five male and five female rats were administered kava kava extract in corn oil by gavage at doses of 0, 0.125, 0.25, 0.5, 1.0, or 2.0 g/kg body weight, 5 days per week for 16 days. One female rat administered 2.0 g/kg kava kava extract died on day 3 of the study. Mean body weights of all dosed groups of rats were similar to those of the vehicle controls. Clinical findings included abnormal breathing, ataxia, and lethargy in the 2.0 g/kg groups of males and females and ataxia and lethargy in the 1.0 g/kg group of females. Liver weights were significantly increased in 1.0 and 2.0 g/kg males and in 0.5 g/kg or greater females compared to the vehicle controls. Minimal hepatocellular hypertrophy occurred in all 2.0 g/kg males and in all females administered 0.25 g/kg or greater. 2-WEEK STUDY IN MICE: Groups of five male and five female mice were administered kava kava extract in corn oil by gavage at doses of 0, 0.125, 0.25, 0.5, 1.0, or 2.0 g/kg body weight, 5 days per week for 17 days. In the 2.0 g/kg group of males, one died on day 2 and one died on day 3. Mean body weights of all dosed groups of mice were similar to those of the vehicle controls. Clinical findings included abnormal breathing, ataxia, and lethargy in males and females in the 1.0 and 2.0 g/kg groups. Liver weights of 2.0 g/kg males and females were significantly increased. The incidence of hepatocellular hypertrophy in 2.0 g/kg female mice was significantly greater than that in the vehicle control group. 3-MONTH STUDY IN RATS: Groups of 10 male and 10 female rats were administered kava kava extract in corn oil by gavage at doses of 0, 0.125, 0.25, 0.5, 1.0, or 2.0 g/kg, 5 days per week for 14 weeks. Deaths attributed to kava kava extract administration included three males and four females in the 2.0 g/kg groups and one female in the 1.0 g/kg group. One 0.25 g/kg male and one vehicle control female also died before the end of the study. The mean body weights of males in the 1.0 and 2.0 g/kg groups and females in the 2.0 g/kg group were significantly less than those of the vehicle controls. Ataxia and lethargy were observed in males and females in the 1.0 g/kg groups during week 1 and in the 2.0 g/kg groups throughout the study. Increased -glutamyltransferase activity in 1.0 g/kg females and 2.0 g/kg males and females may represent enzyme induction. However, the hepatocellular hypertrophy observed in the 2.0 g/kg females may have contributed to the increased -glutamyltransferase activity. The liver weights of 0.25 g/kg or greater males and 0.5 g/kg or greater females were significantly increased compared to the vehicle controls. The kidney weights of 0.5 g/kg or greater males and females were significantly increased compared to the vehicle controls. The incidence of hepatocellular hypertrophy in 2.0 g/kg females was significantly greater than that in the vehicle controls. 3-MONTH STUDY IN MICE: Groups of 10 male and 10 female mice were administered kava kava extract in corn oil by gavage at doses of 0, 0.125, 0.25, 0.5, 1.0, or 2.0 g/kg, 5 days per week for 14 weeks. Four male and three female 2.0 g/kg mice died during week 1; these deaths were attributed to kava kava extract administration. One additional 2.0 g/kg female died during week 6 due to a gavage accident. The mean body weights of dosed males and females were similar to those of the vehicle controls. Ataxia and lethargy occurred in males and females in the 1.0 and 2.0 g/kg groups during week 1. The liver weights of 2.0 g/kg males and 1.0 and 2.0 g/kg females were significantly increased compared to those of the vehicle control groups. The incidences of centrilobular hypertrophy in the liver of 0.5 g/kg or greater males and 1.0 and 2.0 g/kg females were significantly greater than those in the vehicle controls. 2-YEAR STUDY IN RATS: Groups of 49 or 50 male and 50 female rats were administered kava kava extract in corn oil by gavage at doses of 0, 0.1, 0.3, or 1.0 g/kg, 5 days per week for 104 (males) or 105 (females) weeks. Survival of dosed groups of males and females was similar to that of the vehicle controls. Mean body weights of males administered 1.0 g/kg were less than those of the vehicle controls after week 65, and those of the 1.0 g/kg females were less than those of the vehicle controls after week 41. Clinical findings included ataxia and lethargy that occurred in 21 males and 14 females in the 1.0 g/kg groups during the first 4 weeks of the study. After week 5, ataxia and lethargy were noted in 10 males and eight females in the 1.0 g/kg groups and these findings were observed randomly and intermittently throughout the study. At approximately 1 year into the study, twitching and seizures were observed in males and females in all dosed groups but mainly in the 1.0 g/kg groups. There was a dose-related increase in the incidences of interstitial cell adenoma in the testis with increased incidences of bilateral neoplasms. The incidences of hepatocellular hypertrophy in 1.0 g/kg males and females were significantly greater than those in the vehicle controls. Increased -glutamyltransferase activity and/or bile salt concentrations in males and females may represent a cholestatic event related to the hepatocellular hypertrophy observed in rats. Enzyme induction may have played a role in the increased -glutamyltransferase activity. Significantly increased incidences of centrilobular fatty change occurred in 0.1 and 1.0 g/kg males. The incidences of inflammation, ulcer, and epithelial hyperplasia in the forestomach were significantly increased in 1.0 g/kg males and females. The severity of nephropathy was increased in 1.0 g/kg male rats, and the incidence of nephropathy was significantly increased in 1.0 g/kg females. Incidences of transitional epithelial hyperplasia of the pelvis of the kidney were significantly increased in 1.0 g/kg males and 0.3 and 1.0 g/kg females. The incidences of retinal degeneration in the eye were significantly increased in 1.0 g/kg males and females. The incidences of metaplasia of pancreatic acinar cells to a hepatocytic morphology increased in 1.0 g/kg males and females, and the increase in males was significant. Significantly decreased incidences of pars distalis adenoma in the pituitary gland occurred in 1.0 g/kg males and in 0.1 and 1.0 g/kg females. The incidence of fibroadenoma of the mammary gland in 1.0 g/kg females was significantly less than that in the vehicle control group. 2-YEAR STUDY IN MICE: Groups of 50 male and 50 female mice received kava kava extract in corn oil by gavage at doses of 0, 0.25, 0.5, or 1.0 g/kg, 5 days per week for 105 weeks. Survival of dosed groups of males and females was similar to that of the vehicle controls. Mean body weights of males administered 1.0 g/kg were generally similar to those of the vehicle controls until the end of the study; however, those of 1.0 g/kg females were less than those of the vehicle controls after week 21. Clinical findings included ataxia and lethargy that occurred in 13 males and 31 females in the 1.0 g/kg groups during the first week of the study. Decreasing numbers of animals exhibited ataxia or lethargy during the remainder of the study, but these findings were observed in 1.0 g/kg females as late as week 101. The incidences of hepatoblastoma in 0.5 and 1.0 g/kg males were significantly increased compared to the vehicle controls. The incidences of hepatocellular carcinoma or hepatoblastoma (combined) were significantly increased in 0.5 g/kg males. Incidences of hepatocellular carcinoma were increased in all dosed groups of females, and the increase was significant in the 0.25 g/kg group. The incidences of hepatocellular adenoma or carcinoma (combined) were significantly increased in 0.25 and 0.5 g/kg females. In the liver, the incidences of centrilobular hypertrophy in all dosed groups of males and females were significantly greater than those in the vehicle control groups. Significantly increased incidences of eosinophilic foci occurred in 0.5 g/kg males and in 1.0 g/kg males and females, and the incidence of angiectasis was significantly increased in the 1.0 g/kg males. The incidences of hepatocellular necrosis were significantly increased in 0.25 and 1.0 g/kg males. In the forestomach, the incidences of chronic inflammation, epithelial hyperplasia, and erosion were significantly increased in 0.5 and 1.0 g/kg females, and the incidence of ulceration was significantly increased in 1.0 g/kg females. GENETIC TOXICOLOGY: Kava kava extract was tested for bacterial mutagenicity over a broad range of concentrations in two independent assays using several strains of bacteria (S. typhimurium tester strains TA97, TA98, TA100, and TA1535 and E. coli strain WP2 uvrA/pKM101), with and without exogenous metabolic activation. No increase in mutant colonies was seen in any of the tester strains, under any activation condition. (ABSTRACT TRUNCATED)

Are mould hepatotoxins responsible for kava hepatotoxicity?

Previous studies with kava components such as kavalactones, pipermethystine and flavokavain B have demonstrated hepatotoxicity from these constituents. Regardless, there has recently been speculation that adulterants or impurities such as the mould hepatotoxin aflatoxin are a more likely cause of kava hepatotoxicity, despite a paucity of supporting evidence. Although there is limited similarity between acute kava hepatotoxicity and acute aflatoxicosis, and background levels of aflatoxin have been detected in kava samples, unless epidemiological investigations can uncover direct evidence implicating mould hepatotoxins, it remains more likely that chemical constituents of kava are the cause of the hepatotoxicity from kava.

Kava hepatotoxicity in traditional and modern use: the presumed Pacific kava paradox hypothesis revisited.

Kava, a Pacific herb consumed worldwide for medicinal, recreational and cultural purposes, has been associated with rare hepatotoxicity, and there is currently a critical need to determine this causation. The previously proposed Pacific kava paradox was based on the theory that kava hepatotoxicity was not observed following use of traditional aqueous extracts in the Pacific region, but was restricted to use of Western acetonic and ethanolic extracts. Subsequent cases analyzed by the World Health Organization and published case reports revealed that traditional aqueous extracts used in New Caledonia, Australia, the USA and Germany may also be hepatotoxic; thus, there is no longer a basis to sustain the previously proposed Pacific kava paradox. It appears that the primary cause of toxicity may reside in the time before the preparation of the various kava extracts, possibly attributed to poor quality of the raw material caused by mould hepatotoxins. Rigorous testing of kava raw material is urgently advised, in addition to Pan-Pacific kava manufacturing quality standards.

Liver toxicity and carcinogenicity in F344/N rats and B6C3F1 mice exposed to Kava Kava.

Kava Kava is an herbal supplement used as an alternative to antianxiety drugs. Although some reports suggest an association of Kava Kava with hepatotoxicity , it continues to be used in the United States due to lack of toxicity characterization. In these studies F344/N rats and B6C3F1 mice were administered Kava Kava extract orally by gavage in corn oil for two weeks, thirteen weeks or two years. Results from prechronic studies administered Kava Kava at 0.125 to 2g/kg body weight revealed dose-related increases in liver weights and incidences of hepatocellular hypertrophy. In the chronic studies, there were dose-related increases in the incidences of hepatocellular hypertrophy in rats and mice administered Kava Kava for up to 1g/kg body weight. This was accompanied by significant increases in incidences of centrilobular fatty change. There was no treatment- related increase in carcinogenic activity in the livers of male or female rats in the chronic studies. Male mice showed a significant dose-related increase in the incidence of hepatoblastomas. In female mice, there was a significant increase in the combined incidence of hepatocellular adenoma and carcinoma in the low and mid dose groups but not in the high dose group. These findings were accompanied by several nonneoplastic hepatic lesions.

Kava and kava hepatotoxicity: requirements for novel experimental, ethnobotanical and clinical studies based on a review of the evidence.

Kava hepatotoxicity is a well described disease entity, yet there is uncertainty as to the culprit(s). In particular, there is so far no clear evidence for a causative role of kavalactones and non-kavalactone constituents, such as pipermethystine and flavokavain B, identified from kava. Therefore, novel enzymatic, analytical, toxicological, ethnobotanical and clinical studies are now required. Studies should focus on the identification of further potential hepatotoxic constituents, considering in particular possible adulterants and impurities with special reference to ochratoxin A and aflatoxins (AFs) producing Aspergillus varieties, which should be urgently assessed and published. At present, Aspergillus and other fungus species producing hepatotoxic mycotoxins have not yet been examined thoroughly as possible contaminants of some kava raw materials. Its occurence may be facilitated by high humidity, poor methods for drying procedures and insufficient storage facilities during the time after harvest. Various experimental studies are recommended using aqueous, acetonic and ethanolic kava extracts derived from different plant parts, such as peeled rhizomes and peeled roots including their peelings, and considering both noble and non-noble kava cultivars. In addition, ethnobotanical studies associated with local expertise and surveillance are required to achieve a good quality of kava as the raw material. In clinical trials of patients with anxiety disorders seeking herbal anxiolytic treatment with kava extracts, long-term safety and efficacy should be tested using traditional aqueous extracts obtained from peeled rhizomes and peeled roots of a noble kava cultivar, such as Borogu, to evaluate the risk: benefit ratio. Concomitantly, more research should be conducted on the bioavailability of kavalactones and non-kavalactones derived from aqueous kava extracts. To be on the side of caution and to ensure lack of liver injury, kava consuming inhabitants of the kava producing or importing South Pacific islands should undergo assessment of their liver function values and serum aflatoxin levels. The primary aim is to achieve a good quality of kava raw material, without the risk of adulterants and impurities including ochratoxin A and AFs, which represent the sum of aflatoxin B1, B2, G1 and G2. Although it is known that kava may naturally be contaminated with AFs, there is at present no evidence that kava hepatotoxicity might be due to aflatoxicosis. However, appropriate studies have yet to be done and should be extended to other mould hepatotoxins, with the aim of publishing the obtained results. It is hoped that with the proposed qualifying measures, the safety of individuals consuming kava will substantially be improved.

Does inflammation play a role in kava hepatotoxicity?

The pathophysiology of kava hepatotoxicity remains inconclusive. There is circumstantial evidence for the roles of toxic metabolites, inhibition of cyclooxygenase (COX) enzymes and depletion of liver glutathione. Pharmacogenomic effects are likely, particularly for Cytochrome P450 genes. Experimental and clinical cases of hepatotoxicity show evidence of hepatitis. The question remains whether this inflammation is caused by components of kava directly, or indirectly due to the downstream effects.

Gene expression profiling in male B6C3F1 mouse livers exposed to kava identifies--changes in drug metabolizing genes and potential mechanisms linked to kava toxicity.

The association of kava products with liver-related health risks has prompted regulatory action in many countries. We used a genome-wide gene expression approach to generate global gene expression profiles from the livers of male B6C3F1 mice administered kava extract by gavage for 14 weeks, and identified the differentially expressed drug metabolizing genes in response to kava treatments. Analyses of gene functions and pathways reveal that the levels of significant numbers of genes involving drug metabolism were changed and that the pathways involving xenobiotics metabolism, Nrf2-mediated oxidative stress response, mitochondrial functions and others, were altered. Our results indicate that kava extract can significantly modulate drug metabolizing enzymes, potentially leading to herb-drug interactions and hepatotoxicity.

Kava hepatotoxicity: regulatory data selection and causality assessment.

BACKGROUND: Kava hepatotoxicity in 20 patients from Germany has been debated worldwide following a regulatory ad hoc causality assessment and ban of kava, an anxiolytic herbal remedy obtained from the rhizome of Piper methysticum Forster. AIMS: We assessed causality with a quantitative structured causality analysis in all 20 cases of patients with liver disease, presented by the German regulatory agency that assumed a causal relationship with the use of kava extracts. METHODS: The quantitative scale of CIOMS (Council for International Organizations of Medical Sciences) in its updated form was employed for causality assessment and quality evaluation of the regulatory data presentation. RESULTS: The regulatory information is scattered and selective, and items essential for causality assessment, such as exclusion of kava independent causes, were not, or only marginally, considered by the regulator. Quantitative causality assessment for kava was possible (n=2), unlikely (n=12), or excluded (n=6), showing no concordance with the regulatory ad hoc causality evaluation. CONCLUSION: The regulatory data regarding kava hepatotoxicity is selective and of low quality, not supportive of the regulatory proposed causality; but instead, is an explanation of the overall causality discussions of kava hepatotoxicity. We are proposing that the regulatory agency reports data in full length and reevaluates causality.

Analysis of gene expression changes of drug metabolizing enzymes in the livers of F344 rats following oral treatment with kava extract.

The association of kava product use with liver-related risks has prompted regulatory action in many countries. We studied the changes in gene expression of drug metabolizing enzymes in the livers of Fischer 344 male rats administered kava extract by gavage for 14 weeks. Analysis of 22,226 genes revealed that there were 14, 41, 110, 386, and 916 genes significantly changed in the 0.125, 0.25, 0.5, 1.0, and 2.0 g/kg treatment groups, respectively. There were 16 drug metabolizing genes altered in all three high-dose treatment groups, among which seven genes belong to cytochrome P450 isozymes. While gene expression of Cyp1a1, 1a2, 2c6, 3a1, and 3a3 increased; Cyp 2c23 and 2c40 decreased, all in a dose-dependent manner. Real-time PCR analyses of several genes verified these results. Our results indicate that kava extract can significantly modulate drug metabolizing enzymes, particularly the CYP isozymes, which could cause herb-drug interactions and may potentially lead to hepatotoxicity.

High dose of commercial products of kava (Piper methysticum) markedly enhanced hepatic cytochrome P450 1A1 mRNA expression with liver enlargement in rats.

Commercial products containing the kava plant (Piper methysticum), known to have the anxiolytic activity, are banned in several European countries and Canada because of the suspicion of a potential liver toxicity. In some reports, kava and kavalactones (major constituents of kava) inhibited activities of cytochrome P450 (CYP) isoforms including CYP1A2. On the other hand, a few studies showed that administration of kava to rats moderately increased CYP1A2 proteins in the liver. CYP1A isoforms are likely responsible for the metabolic activation of potent carcinogenic environmental toxins such as aflatoxins, benzo[a]pyrene, and others. On these bases, we have investigated the effects of administration of commercial kava products on gene expression of hepatic CYP1A isoforms in rats. A high dose (equivalent to approximately 380mg kavalactones/kg/day; 100 times of the suggested dosage for human use) of two different types of kava products for 8 days significantly increased liver weights. CYP1A2 mRNA expression was moderately increased (2.8-7.3 fold). More importantly, the high dose of kava markedly enhanced CYP1A1 mRNA expression (75-220 fold) as well as ethoxyresorufin O-deethylase activities and CYP1A1 immunoreactivities. Thus, no observed adverse effect levels of kavalactones would be lower than 380mg/kg/day. When the safety factor of kavalactones is assumed to be 100, a value most often used upon the risk analysis of chemicals and designed to account for interspecies and intraspecies variations, a number of kava product users likely ingest more kavalactones than acceptable daily intakes. Based on overall evidence, we should pay considerable attention to the possibility that kava products induce hepatic CYP1A1 expression in human especially in sensitive individuals.

Evaluation of commercial kava extracts and kavalactone standards for mutagenicity and toxicity using the mammalian cell gene mutation assay in L5178Y mouse lymphoma cells.

Kava (Piper methysticum) is a member of the pepper family and has been cultivated by South Pacific islanders for centuries and used as a social and ceremonial drink. Traditionally, kava extracts are prepared by grinding or chewing the rhizome and mixing with water and coconut milk. The active constituents of kava are a group of approximately 18 compounds collectively referred to as kavalactones or kava pyrones. Kawain, dihydrokawain, methysticin, dihydromethysticin, yangonin, and desmethoxyyangonin are the six major kavalactones. Kava beverages and other preparations are known to be anxiolytic and are used for anxiety disorders. Dietary supplements containing the root of the kava shrub have been implicated in several cases of liver toxicity in humans, including several who required liver transplants after using kava supplements. In order to study the toxicity and mutagenicity, two commercial samples of kava, Kaviar and KavaPure, and the six pure kavalactones including both D-kawain and DL-kawain, were evaluated in L5178Y mouse lymphoma cells. Neither the kava samples nor the kavalactones induced a mutagenic response in the L5178Y mouse lymphoma mutation assay with the addition of human liver S9 activation.

Immunohistochemical analysis of expressions of hepatic cytochrome P450 in F344 rats following oral treatment with kava extract.

Kava (Piper methysticum), used for relaxation and pain relief, has been one of the leading dietary supplements and several reports linking hepatic functional disturbances and liver failure to kava have resulted in a ban on sales in Europe and Canada and the issuance of warnings by the US FDA. The National Toxicology Program conducted 14-week rat studies to characterize the toxicology of kava exposure in Fischer 344 rats [National Toxicity Program. 90 day gavage toxicity studies of KAVA KAVA EXTRACT in Fischer rats and B6C3F1 mice. Research Triangle Park, NC; 2005a; National Toxicity Program. Testing status of agents at NTP (KAVA KAVA EXTRACT M990058). Research Triangle Park, NC; 2005b. (http://ntp.niehs.nih.gov/index.cfm?objectid=071516E-C6E1-7AAA-C90C751E23D14C1B)]. Groups of 10 male and 10 female rats were administered kava extract by gavage at 0, 0.125, 0.25, 0.5, 1.0, and 2.0 g/kg/day. Increased gamma-glutamyl-transpeptidase (GGT) activities were observed in the 2.0 g/kg males and 1.0 and 2.0 g/kg females, as well as increased serum cholesterol levels in males and females at 0.5 g/kg and higher. Increases in incidence and severity of hepatocellular hypertrophy (HP) were noted in males at 1.0 g/kg and females at 0.5 g/kg and higher, as well as increased liver weights. Immunohistochemical analyses of the expression of cytochrome-P450 (CYP) enzymes in liver of the control and 1.0- and 2.0-g/kg-treated groups indicated decreased expression of CYP2D1 (human CYP2D6 homolog) in 2.0 g/kg females and increased expression of CYP1A2, 2B1, and 3A1 in 1.0 and 2.0 g/kg groups of both sexes. The no observed adverse effect levels were decided as 0.25 g/kg in both genders, based on neurotoxic effects, increases in GGT, cholesterol, liver weight, and HP and decreases in body weight. Kava-induced hepatic functional changes in the F344 rat might be relevant to human clinical cases of hepatotoxicity following exposure.

Safety of ethanolic kava extract: Results of a study of chronic toxicity in rats.

BACKGROUND: Recently, potential liver toxicity was discussed with the intake of kava extract preparations (Piper methysticum) as anxiolytic drugs. The aim of this study was to test chronic toxicity in rats by oral application of an ethanolic kava full extract. METHODS: Wistar rats of both sexes were fed 7.3 or 73 mg/kg body weight of ethanolic kava extract for 3 and 6 months. The animals were examined for changes in body weight, hematological and liver parameters, and macroscopical and microscopical histological changes in the major organs. RESULTS: No signs of toxicity could be found. CONCLUSIONS: The results are in accordance with the medical experience regarding the use of kava preparations and the long tradition of kava drinking in the South Pacific island states. Specifically, the results do not back the suspicion of potential liver toxicity.

In vitro cytotoxicity of nonpolar constituents from different parts of kava plant (Piper methysticum).

Kava (Piper methysticum), a perennial shrub native to the South Pacific islands, has been used to relieve anxiety. Recently, several cases of severe hepatotoxicity have been reported from the consumption of dietary supplements containing kava. It is unclear whether the kava constituents, kavalactones, are responsible for the associated hepatotoxicity. To investigate the key components responsible for the liver toxicity, bioassay-guided fractionation was carried out in this study. Kava roots, leaves, and stem peelings were extracted with methanol, and the resulting residues were subjected to partition with a different polarity of solvents (hexane, ethyl acetate, n-butanol, and water) for evaluation of their cytotoxicity on HepG2 cells based on the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and lactate dehydrogenase and aspartate aminotransferase enzyme leakage assays. Organic solvent fractions displayed a much stronger cytotoxicity than water fractions for all parts of kava. The hexane fraction of the root exhibited stronger cytotoxic effects than fractions of root extracted with other solvents or extracts from the other parts of kava. Further investigations using bioassay-directed isolation and analysis of the hexane fraction indicated that the compound responsible for the cytotoxicity was flavokavain B. The identity of the compound was confirmed by (1)H and (13) C NMR and MS techniques.