Intestinal epithelial damage due to herbal compounds - an in vitro study.

Intestinal epithelial injury from herbal products has rarely been reported, despite the gut being the first point of contact for oral preparations. These products often consist of multiple herbs, thereby potentially exposing consumers to higher levels of reactive phytochemicals than predicted due to pharmacokinetic interactions. The phytochemical coumarin, found in many herbal products, may be taken in combination with herbal medicines containing astragalosides and atractylenolides, purported cytochrome P450 (CYP) modulators. As herbal use increases, the need to predict interactions in multiple at-risk organ systems is becoming critical. Hence, to determine whether certain herbal preparations containing coumarin may cause damage to the intestinal epithelium, Caco2 cells were exposed to common phytochemicals. Coumarin, astragaloside IV (AST-IV) or atractylenolide I (ATR-I) solutions were exposed to Caco2 cultures in increasing concentrations, individually or combined. Coumarin produced a significant concentration-dependant fall in cell viability that was potentiated when CYP enzymes were induced with rifampicin and incubated with CYP3A4 inhibitor econazole, suggesting a role for other CYP enzymes generating toxic metabolites. ATR-I alone produced no toxicity in uninduced cells but showed significant toxicity in rifampicin-induced cells. ATR-I had no effect on coumarin-induced toxicity. AST-IV was nontoxic alone but produced significant toxicity when combined with nontoxic concentrations of coumarin. The combination of coumarin, ATR-I and AST-IV was significantly toxic, but no synergistic interaction was seen. This investigation was conducted to determine the likelihood for intestinal-based interactions, with the results demonstrating coumarin is potentially toxic to intestinal epithelium, and combinations with other phytochemicals can potentiate this toxicity.

Could herbal soup be a potentially unrecognized cause of hepatotoxicity at autopsy?

Unexpected hepatic failure with liver necrosis is sometimes encountered during a forensic autopsy. Determining the etiology may sometimes be difficult, although increasingly herbal medicines are being implicated. To determine whether such effects might also be caused by foodstuffs, the following in vitro study was undertaken. Four formulations of traditional herbal soup advertised as bak kut teh were prepared and added to cultures of liver carcinoma cells (HepG2). Cell viability was assessed using an MTT colorimetric assay at 48 h demonstrating that all formulations had significant toxicity prior to dilution (p < 0.05). Formulation #1 showed 21% cell death (p = 0.023), Formulation #2 30% (p = 0.009), and Formulation #3 41% (p < 0.0001). Formulations #1-3 showed no significant toxicity once diluted (p > 0.05). Formulation 4 showed approximately 83% cell death before dilution (p < 0.0001) and persistent toxicity even with dilutions at 1:10 (15% ± 3.7, p = 0.023) and 1:1000 (14% ± 3.8, p = 0.024). This study has shown that herbal foodstuffs such as bak kut teh may be responsible for variable degrees of in vitro hepatotoxicity, thus extending the range of herbal products that may be potentially injurious to the liver. If unexpected liver damage is encountered at autopsy, information on possible recent ingestion of herbal food preparations should be sought, as routine toxicology screening will not identify the active components. Liver damage may therefore be caused not only by herbal medicines but possibly by herbal products contained in food.

Implications for herbal polypharmacy: coumarin-induced hepatotoxicity increased through common herbal phytochemicals astragaloside IV and atractylenolide I.

Hepatotoxicity is a well-known adverse effect of many substances, with toxicity often resulting from interactions of drugs with other drug-like substances. With the increased availability of complementary and alternative medicines, including herbal medicines, the likelihood of adverse interactions between drugs and drug-like substances in herbs increases. However, the impact of potential herb-herb interactions is little understood. To assess the potential of two cytochrome P450 enzyme modulating phytochemicals common to many herbal medicines, atractylenolide I (ATR-I) and astragaloside IV (AST-IV), to interact with coumarin, another phytochemical common in many foods, a hepatocyte function model with a liver carcinoma cell line, HepG2, was exposed to these agents. To determine the effects of cytochrome P450 modulation by these phytochemicals certain cells were induced with rifampicin to induce cytochrome P450. Increasing concentrations of ATR-I combined with a fixed, nontoxic concentration of coumarin (200 µM), demonstrated significant additive interactions. 300 µM ATR-I produced a 31% reduction in cell viability (p < 0.01) with coumarin in rifampicin uninduced cells. In rifampicin-induced cells, ATR-I (100-300 µM) produced a significant reduction in cell viability (p < 0.01) with coumarin (200 µM). AST-IV with fixed coumarin (200 µM) showed 27% toxicity at 300 µM AST-IV in rifampicin uninduced cells (p < 0.05) and 30% toxicity in rifampicin induced cells (p < 0.05). However, when fixed coumarin and AST-IV were combined with increasing concentrations of ATR-I no further significant increase in toxicity was observed (p > 0.05). These results demonstrate the potential toxic interactive capabilities of common traditional Chinese herbal medicine phytochemicals and underline the potential importance of coumarin-mediated toxicity.

Flavonoid-statin interactions causing myopathy and the possible significance of OATP transport, CYP450 metabolism and mevalonate synthesis.

3-hydroxy-3-methyl-glutaryl-coenzyme A (HMG-CoA) reductase inhibitors, statins, are a primary treatment for hyperlipidemic cardiovascular diseases which are a leading global cause of death. Statin therapy is life saving and discontinuation due to adverse events such as myotoxicity may lead to unfavourable outcomes. There is no known mechanism for statin-induced myotoxicity although it is theorized that it is due to inhibition of downstream products of the HMG-CoA pathway. It is known that drug-drug interactions with conventional medicines exacerbate the risk of statin-induced myotoxicity, though little attention has been paid to herb-drug interactions with complementary medicines. Flavonoids are a class of phytochemicals which can be purchased as high dose supplements. There is evidence that flavonoids can raise statin plasma levels, increasing the risk of statin-induced myopathy. This could be due to pharmacokinetic interactions involving hepatic cytochrome 450 (CYP450) metabolism and organic anion transporter (OATP) absorption. There is also the potential for flavonoids to directly and indirectly inhibit HMG-CoA reductase which could contraindicate statin-therapy. This review aims to discuss what is currently known about the potential for high dose flavonoids to interact with the hepatic CYP450 metabolism, OATP uptake of statins or their ability to interact with HMG-CoA reductase. Flavonoids of particular interest will be covered and the difficulties of examining herbal products will be discussed throughout.

The potential side effects of herbal preparations in domestic animals.

A recent series of deaths in previously healthy dogs in Victoria, Australia associated with the ingestion of raw meat contaminated by indospicine derived from native Australian plants of the Indigofera species draws attention to the potential that exists for herbal toxicity in domestic animals. Although the efficacy of herbal remedies generally remains unproven in domestic animals, herbal preparations are being increasingly used as supplements and treatments. Issues with incorrect ingredients, inadequate processing, faulty, incomplete or inaccurate product labelling, contamination with toxins, adulteration with undeclared pharmaceutical agents and herb-herb interactions are well recognized as causes of adverse effects in humans. However, apart from of the effects of noxious weed species, the literature on herbal toxicity in domestic animals is sparse. Thus, the forensic evaluation of cases of suspected poisoning in domestic animals should also encompass an accurate description the type and dose of any herbal preparations that may have been recently administered.

The caul and its relation to drowning, lawyers and sorcerers.

The caul is a fragment of amniotic membrane that is occasionally observed adherent to the head and face of a newborn child. A dried caul has been in The University of Adelaide Pathology Archive for a number of years. Beliefs in the supernatural powers of these detached membranes are found in the historical traditions of a wide variety of cultures, resulting in cauls being regularly dried out and sold as talismans. It was thought that lawyers could sway opinions in court more readily if they possessed a caul, and that such a token could either be used to facilitate black magic rituals or alternatively to thwart the machinations of sorcerers and witches. A pervasive belief was that a caul would guarantee that the owner would never drown resulting in cauls being sold for between £10 to £30 in the seventeenth century at a time when considerable naval warfare and maritime exploration were occurring. Thus, rather than illuminating pathophysiological processes occasional specimens in pathology archives may remind us of folk traditions and superstitions that have long since been forgotten in modern times.

Paracetamol (acetaminophen) hepatotoxicity increases in the presence of an added herbal compound.

Hepatotoxicity from paracetamol/acetaminophen has occasionally been reported at lower than expected doses. As herbal preparations may interact with pharmaceutical drugs the following in vitro study was undertaken to determine whether the toxic effects of paracetamol on liver cell growth in culture would be exacerbated by the addition of psoralen, a furanocoumarin compound that is present in Psoralea corylifolia, a common Chinese herb. The following study utilising a liver carcinoma cell line (HepG2) showed that Psoralea corylifolia was significantly toxic from 0.3 mg/ml to 5 mg/ml (p < 0.05), whereas paracetamol was not toxic below 50 mM (p = 0.0026). Interactions between previously non-toxic levels of 0.1 mg/ml of Psoralea corylifolia and increasing concentrations of paracetamol (0-50 mM), however, were observed, with a significant increase in toxicity compared to paracetamol alone (30% cell death vs. 72% cell death with Psoralea corylifolia). A significant synergistic interaction was observed at 40 mM paracetamol with 0.1 mg/ml of Psoralea (p = 0.038). This study has, therefore, shown significantly increased hepatotoxicity in cell cultures exposed to paracetamol when herbal compounds containing furanocoumarins were added. Fulminant acute liver failure occurring after the ingestion of low doses of paracetamol may not, therefore, always be due to an occult idiosyncratic response to paracetamol, but instead possibly to the combined effects of paracetamol and herbal preparations. Given the widespread use of both paracetamol and herbal preparations this possibility should be considered in cases of unexplained hepatic necrosis and liver failure that present for medicolegal investigation.

Potential forensic implications of kerokan therapy - a traditional Indonesian therapy associated with neck bruising.

Kerokan is a traditional Indonesian treatment involving abrading the skin over various parts of the body with a blunt object such as a coin or a piece of ginger which may create suspicious injuries. Here, the case of a 30-year-old woman who had undergone kerokan therapy is reported. She was complaining of retro-sternal pain and epigastric discomfort and subsequently became markedly short of breath and died. The observation of injuries predominantly located around the neck initiated a police investigation and forensic autopsy. Scattered abrasions, blisters and bruises were present on the forehead, the bridge of the nose extending to the ala, the upper and anterior aspect of the chin and the sternal notch, with discrete fresh and scabbed abrasions around the neck, over the sternum and on the backs of both hands. There were, however, no facial or conjunctival petechiae and no bruising of the underlying strap muscles, or fractures of the hyoid bone or thyroid cartilage. Histology revealed that myocarditis was the cause of death. Traditional practices may cause unusual bruises, abrasions, blisters and burns that may raise suspicion of inflicted injury from an assault. Knowledge of the manifestations of socio-ethnic practices such as kerokan is important in the evaluation of trauma in specific cultural groups.

Acute Respiratory Obstruction due to Accidental Inhalation of Perlite: A Novel Mechanism for Upper Airway Occlusion with Cast Formation.

A 56-year-old man died following a fall resulting in complete submersion into a deep pit containing insulation material, expanded perlite. The most striking finding at autopsy was of impacted, moist pale yellow perlite that extended from the epiglottis into the main bronchi resulting in complete obstruction of the larger and smaller airways with cast formation. Perlite inhalation differs from inhalation of inert materials such as sand and wheat due to its hygroscopic properties and ability to expand forming an occlusive cast. Subsequent analyses of perlite from the worksite and within the airways indicated that hydration of perlite and adsorption of organic molecules into the perlite glass flakes had formed an interlocking, three-dimensional structure that was likely responsible for triggering a coagulation-flocculation process causing strong cohesion between the aggregates of glass flakes. This unique mechanism was likely responsible for the formation of the plug of perlite that obstructed the upper airway.

Combined Liquid Chromatography-mass Spectrometry and Next-generation DNA Sequencing Detection of Adulterants and Contaminants in Analgesic and Anti-inflammatory Herbal Medicines.

INTRODUCTION: Methods for assessing the quality of herbal medicine preparations have advanced significantly in recent years in conjunction with increases in herbal medicine use and reports of adulteration and contamination. OBJECTIVE: This study examined the quality of analgesic and anti-inflammatory herbal medicine preparations available on the Australian market by detecting the presence of listed ingredients, adulterants and contaminants. METHODS: Forty-nine analgesic and anti-inflammatory herbal medicine preparations were randomly sourced from Australian capital cities. They were audited using a dual approach of liquid chromatography-mass spectrometry (LC-MS) combined with next-generation DNA sequencing. Once screened, a comparison of listed ingredients with verified ingredients was conducted to determine the accuracy of labelling, and the extent of adulteration and contamination. RESULTS: Twenty-six of 49 (53%) herbal medicines were adulterated or contaminated with undeclared ingredients. LC-MS revealed the presence of pharmaceutical adulterants including atropine and ephedrine. DNA sequencing uncovered concerning levels of herbal substitution, adulteration and contamination, including the use of fillers (alfalfa, wheat and soy), as well as pharmacologically relevant species (Centella asiatica, Panax ginseng, Bupleurum and Passiflora). Pig/boar and bird DNA was found in some preparations, inferring substandard manufacturing practices. Of the 26 contaminated samples, 19 (73%) were manufactured in Australia, and 7 (27%) were imported from other countries (6 from China, 1 from New Zealand). In 23 of 49 (47%) herbal medicine samples, no biological ingredients were detected at all. These were predominantly pain and anti-inflammatory preparations such as glucosamine and eicosapentaenoic and docosahexaenoic acids found in krill and fish oils, so DNA would not be expected to survive the manufacturing process. CONCLUSION: The high level of contamination and substitution of herbal medicine preparations sourced from Australian dispensaries supports the need for more stringent pharmacovigilance measures in Australia and abroad.

Toxicological screening and DNA sequencing detects contamination and adulteration in regulated herbal medicines and supplements for diet, weight loss and cardiovascular health.

Use of herbal medicines and supplements by consumers to prevent or treat disease, particularly chronic conditions continues to grow, leading to increased awareness of the minimal regulation standards in many countries. Fraudulent, adulterated and contaminated herbal and traditional medicines and dietary supplements are a risk to consumer health, with adverse effects and events including overdose, drug-herb interactions and hospitalisation. The scope of the risk has been difficult to determine, prompting calls for new approaches, such as the combination of DNA metabarcoding and mass spectrometry used in this study. Here we show that nearly 50% of products tested had contamination issues, in terms of DNA, chemical composition or both. Two samples were clear cases of pharmaceutical adulteration, including a combination of paracetamol and chlorpheniramine in one product and trace amounts of buclizine, a drug no longer in use in Australia, in another. Other issues include the undeclared presence of stimulants such as caffeine, synephrine or ephedrine. DNA data highlighted potential allergy concerns (nuts, wheat), presence of potential toxins (Neem oil) and animal ingredients (reindeer, frog, shrew), and possible substitution of bird cartilage in place of shark. Only 21% of the tested products were able to have at least one ingredient corroborated by DNA sequencing. This study demonstrates that, despite current monitoring approaches, contaminated and adulterated products are still reaching the consumer. We suggest that a better solution is stronger pre-market evaluation, using techniques such as that outlined in this study.

Analysis of spontaneous adverse drug reactions to echinacea, valerian, black cohosh and ginkgo in Australia from 2000 to 2015.

OBJECTIVE: Assessing adverse drug reactions (ADRs) is a proven method to estimate the safety of medicines. The ADRs to herbal medicines in Australia (and by inference, the safety of herbal medicines in Australia) remain unknown. This study examines spontaneous ADR cases to four of the most popular herbs in Australia from 2000 to 2015: echinacea (Echinacea purpurea), valerian (Valeriana officinalis), black cohosh (Actaea racemosa) and ginkgo (Ginkgo biloba). METHODS: ADRs of echinacea, valerian, black cohosh and ginkgo reported to the Australian Therapeutic Goods Administration (TGA) between 2000 and 2015 were obtained from the TGA database. Data were collated and analysed according to age, sex, severity, type of ADR and body system affected. Statistics were calculated using GraphPad Prism software. RESULTS: Most ADRs were mild or moderate. However, every herbal medicine was associated with life-threatening ADRs. In each life-threatening case, the herbal medicine was taken concomitantly with prescription medications. Black cohosh was associated with a significant number of severe ADRs (30.3% of the total), with 39.4% of these ADRs being associated with abnormal hepatic function, hepatitis or hepatotoxicity. CONCLUSION: This study highlights the lack of public awareness with regard to herb-drug interactions, since most of the severe ADRs involved a herb-drug interaction.

Evidence for the efficacy and safety of herbal weight loss preparations.

Rising rates of obesity across the globe have been associated with an increase in the use of herbal preparations for weight control. However, the mechanisms of action for these substances are often not known, as is the potential for interaction with other herbal preparations or prescription pharmaceutical drugs. To investigate the reported efficacy and safety of herbal weight loss preparations, we conducted a review of the literature focusing on herbs that are most commonly used in weight loss preparations, specifically, Garcinia cambogia, Camellia sinensis, Hoodia gordonii, Citrus aurantium and Coleus forskohlii. There was no clear evidence that the above herbal preparations would cause sustained long-term weight loss in humans in the long term. Serious illness and even death have occasionally resulted from the use of herbal weight loss preparations. Few clinical trials have been undertaken to evaluate the efficacy and/or safety of herbal weight loss preparations. In addition, potential issues of herb-herb and herb-drug interactions are often not considered. Regulation of these products is much less rigorous than for prescription medications, despite documented cases of associated hepatotoxicity.

Unusual osteological findings in sacred relics from the LESJE monastery in central Serbia.

An adult male skeleton was submitted to the Department of Anatomy at the University of Belgrade for evaluation. It was believed to represent the remains of a second to third century Christian saint from the Lesje Monastery in central Serbia. Examination of the remains revealed an old crush fracture of a thoracic vertebra and an unusual, probably congenital, malformation of the atlanto-occipital joint with deformation of the left occipital condyle and resultant narrowing of the foramen magnum. Although the occipital malformations were most likely congenital, they may still have caused, or contributed to, death by compression of the underlying upper cervical spinal cord.

Potential adverse outcomes of herbal preparation use in childhood.

AIM: Complementary and alternative medicines are becoming increasingly popular worldwide with a variety of purported medicinal uses. These products are generally believed to be natural and therefore safe, with few adverse reactions. With this perception, parents are now taking their children to see practitioners prescribing these medicines as well as self-prescribing. Despite this, there are issues regarding safety, efficacy and regulation, with increasing numbers of reports of adverse reactions to these products. Therefore, a mini-review was conducted to ascertain the potential risks to children. METHODS: A overview of literature was conducted to highlight the current use of complementary and alternative medicines in children and the possible risks associated with their use. RESULTS: Infants and children may be more susceptible to harmful effects due to their immature physiology and metabolic pathways and different dosage requirements. Adverse reactions may also be caused by interactions with conventional medicines, contamination with heavy metals, and adulteration of filler products including other plant species or pharmaceutical agents. CONCLUSION: As complementary and alternative medicines become increasingly used alongside and with conventional drug therapy, there needs to be greater awareness and discussion among parents, complementary practitioners and medical practitioners to ensure the overall health and safety of children being exposed to these products.

Evaluation of an early twentieth century Afghan herbalist's preparations.

Mahomet Allum was a flamboyant philanthropist and herbalist who worked in South Australia in the early part of last century, whose herbal therapies generated some controversy at the time. Two of his preparations have survived to the present day, a general tonic and a treatment for liver and kidney dysfunction. Given the frequent use of pharmaceutical drugs in "tonics" at the time, toxicological analysis was undertaken at Forensic Science SA, Adelaide with liquid chromatography/quadrupole-time-of-flight mass-spectrometer (LC-QTOF MS), liquid-chromatography/ diode array detector (LC/UV) and gas chromatography/ nitrogen phosphorous- detector/mass-spectrometer (GC-NPD/MS), to look for common drugs. In addition DNA analysis was also undertaken at Trace and Environmental DNA (TrEnD) Laboratory (Curtin University) to evaluate the types of plant products used to make these remedies. The general tonic contained genera from the Triticeae (wheat) family as well as the Medicago family (includes alfalfa), possibly as fillers. Other genera found included Utrica (nettle) and Passiflora (passion flower). The preparation for liver and kidney disease also contained genera from the Medicago family as well as genera Arctostaphylos (bear berry) which has traditionally been used for the treatment of dysuria and bladder stones. No common drugs were found. Thus it appears that the two treatments prepared by Mahomet Allum contained only herbal substances and not adulterant pharmaceutical agents. The herbals identified provide an insight into herbalist practices in the early twentieth century.