Natural health products (NHP) have been experiencing strong growth in consumer demand, both domestically and in foreign markets. The nature of NHP and the small sector of the population that used them in the past, has allowed them to slip between the cracks of regulatory bodies. As NHPs have become mainstream and have been marketed and distributed through major agri-food supply chains, governments have had to become more active regulators. New Natural Health Product Regulations came into force in Canada on January 1, 2004 to regulate these product which had been generally regulated under the Food and Drug Act. Canada is not alone in its regulatory reform and other countries have begun to create new and often more rigorous regulations for NHP. It as often the case that domestic regulations have unintended and sometimes trade restricting side effects. The current restructuring and focus on regulations of NHP, and the potential importance of trade within this sector, suggests that a better understanding of the non-tariff barriers that may arise could be important for the development of the industry in Canada and elsewhere. Analysis of trade effects arising from standards and regulations is not an easy task. Non-tariff barriers to trade tend to have product-specific effects making it difficult to find general results. Using a review of current approaches to address technical barriers, an analytical framework has been selected and applied to case studies. The cases studies examined the welfare effects of regulations as they pertain to three products with different characteristics; flax omega-3 supplement, elk velvet, and a probiotic supplement. The case studies identified a range of non-tariff barriers arising from international regulatory divergence. The results suggest that trade barriers are likely to arise in the NHP industry and that they will differ from product to product. As a result, there is unlikely to be a single policy prescription that will facilitate the removal of barriers to international market access. Suggestions are made as to how barriers could be eliminated or reduced through formal trade negotiations or less formal bilateral discussions.

Natural health products (NHP) have been experiencing strong growth in consumer demand, both domestically and in foreign markets. The nature of NHP and the small sector of the population that used them in the past, has allowed them to slip between the cracks of regulatory bodies. As NHPs have become mainstream and have been marketed and distributed through major agri-food supply chains, governments have had to become more active regulators. New Natural Health Product Regulations came into force in Canada on January 1, 2004 to regulate these product which had been generally regulated under the Food and Drug Act. Canada is not alone in its regulatory reform and other countries have begun to create new and often more rigorous regulations for NHP. It as often the case that domestic regulations have unintended and sometimes trade restricting side effects. The current restructuring and focus on regulations of NHP, and the potential importance of trade within this sector, suggests that a better understanding of the non-tariff barriers that may arise could be important for the development of the industry in Canada and elsewhere. Analysis of trade effects arising from standards and regulations is not an easy task. Non-tariff barriers to trade tend to have product-specific effects making it difficult to find general results. Using a review of current approaches to address technical barriers, an analytical framework has been selected and applied to case studies. The cases studies examined the welfare effects of regulations as they pertain to three products with different characteristics; flax omega-3 supplement, elk velvet, and a probiotic supplement. The case studies identified a range of non-tariff barriers arising from international regulatory divergence. The results suggest that trade barriers are likely to arise in the NHP industry and that they will differ from product to product. As a result, there is unlikely to be a single policy prescription that will facilitate the removal of barriers to international market access. Suggestions are made as to how barriers could be eliminated or reduced through formal trade negotiations or less formal bilateral discussions.

Objective: To reach consensus on entry-to-practice natural health product (NHP)-related competency statements for Canadian pharmacy students. Methods: Four rounds of a modified Delphi method were conducted. Participants; pharmacy educators,and representatives from Canadian pharmacy organizations (n=17), ranked their level of agreement using a 5-point Likert scale. Results: Consensus occurred when all participants ranked a statement 4 or 5. Three core NHP-related competencies were identified: 1) the ability to incorporate NHP knowledge when providing pharmaceutical care; 2) the ability to access and critically appraise NHP-related information sources, and 3) the ability to provide appropriate education to patients and other health care providers on the effectiveness and potential adverse effects and drug interactions of NHPs. Two additional NHP-related competency statements emerged as important, but consensus was not achieved. Conclusions: If the developed core NHP-related competency statements are widely implemented, Canadian pharmacists will be able to fulfill their NHP-related professional responsibilities upon entry-to-practice.

Natural health products (NHPs) are increasingly sold in pharmacies. The purpose of this study was to determine Canadian pharmacists’ attitudes about their professional responsibilities regarding natural health products. A cross-sectional, random sample survey design was used: 3356 questionnaires were mailed to a random sample of licensed pharmacists in all provinces/territories in Canada (except Nunavut). Survey responses were evaluated using descriptive statistics and logistic regression analyses. Response rate was 61%. Overall, respondents were most likely to endorse pharmacist responsibilities related to vitamins and minerals, followed by herbal medicines, then homeopathic medicines. Responsibilities regarding knowledge of NHPs and counseling on NHPs were more frequently endorsed than responsibilities associated with NHP-related documentation. Self-reported knowledge was positively associated with endorsement of NHP-related responsibilities for all NHP types.

Natural health products (NHPs) are increasingly sold in pharmacies. The purpose of this study was to determine Canadian pharmacists’ attitudes about their professional responsibilities regarding natural health products. A cross-sectional, random sample survey design was used: 3356 questionnaires were mailed to a random sample of licensed pharmacists in all provinces/territories in Canada (except Nunavut). Survey responses were evaluated using descriptive statistics and logistic regression analyses. Response rate was 61%. Overall, respondents were most likely to endorse pharmacist responsibilities related to vitamins and minerals, followed by herbal medicines, then homeopathic medicines. Responsibilities regarding knowledge of NHPs and counseling on NHPs were more frequently endorsed than responsibilities associated with NHP-related documentation. Self-reported knowledge was positively associated with endorsement of NHP-related responsibilities for all NHP types.

Objective: To reach consensus on entry-to-practice natural health product (NHP)-related competency statements for Canadian pharmacy students. Methods: Four rounds of a modified Delphi method were conducted. Participants; pharmacy educators,and representatives from Canadian pharmacy organizations (n=17), ranked their level of agreement using a 5-point Likert scale. Results: Consensus occurred when all participants ranked a statement 4 or 5. Three core NHP-related competencies were identified: 1) the ability to incorporate NHP knowledge when providing pharmaceutical care; 2) the ability to access and critically appraise NHP-related information sources, and 3) the ability to provide appropriate education to patients and other health care providers on the effectiveness and potential adverse effects and drug interactions of NHPs. Two additional NHP-related competency statements emerged as important, but consensus was not achieved. Conclusions: If the developed core NHP-related competency statements are widely implemented, Canadian pharmacists will be able to fulfill their NHP-related professional responsibilities upon entry-to-practice.

The research presented in this thesis addressed knowledge gaps for three medicinal plant taxa, Souroubea spp. (Marcgraviaceae) and Platanus (Platanaceae) as well as Echinacea spp. (Asteraceae). The primary pharmacological mechanism of Souroubea sympetala and Platanus occidentalis were well established, with pentacyclic triterpenes identified as major active principles. My results indicate that major triterpenoids, and crude plant extracts, selectively inhibited monoacyglycerol lipase (MAGL) activity but not fatty acid amide hydrolase (FAAH) activity. These data suggest a possible secondary anxiolytic mechanism of action through the endocannabinoid system (ECS). My study of herb-drug interactions of Souroubea and Plantanus products showed some potential risk when combined with a classic benzodiazepine class drug, diazepam, and I proposed a mechanism through in vitro CYP450 enzyme inhibition. The pharmacokinetic study revealed the difficulty of detecting betulinic acid in animal blood. To support the development a commercial botanical composed of these medicinal plants, an extraction method and a highly sensitive and selectivity HPLC-APCI-MS based quantification method was successfully developed and validated. Part II of this thesis focused on the impact of phytochemical variation and hepatic metabolism on the ECS activity of Echinacea spp. and explored the potential for new applications of Echinacea spp. as a natural health product. My research indicated that considerable variability in the content of phenolic and alkylamide (AKA) compounds reflected similar variability in in vitro bioactivity at ECS-related pharmacological targets. Following biochemometric analysis, several phenolic compounds and AKAs in Echinacea spp. were found to be significant independent variables determining FAAH inhibition and CB receptor activation. Hepatic metabolism was also found to affect the FAAH inhibition of AKA, as increased FAAH inhibitory effects were observed after CYP450-mediated metabolism of both individual AKAs and crude extracts of E angustifolia and E. purpurea, suggesting a “pro-drug” mechanism. Dose dependent activities were observed with oral administration of both E angustifolia and E. purpurea root extract in rat paw model of inflammation and pain. Further tests indicated these activities can be partially blocked by co-administration of CB1 and CB2 receptor antagonists AM251 and AM630, respectively. This evidence suggests activity for peripheral pain was at least partially mediated through the ECS.

Oxidative stress is characterized by the cellular accumulation of reactive oxygen species (ROS). Increased production of ROS, such as the superoxide anion (O2.-), or a deficiency in their clearance by antioxidant defenses, mediates the cellular pathology. Non-alcoholic fatty liver disease (NAFLD) is a broad spectrum liver disorder commonly manifesting in milieu of the metabolic syndrome. Oxidative stress is an important pathogenic mediator in NAFLD, and in its associated morbidities like atherosclerosis. The objective of my research was to investigate the regulation of oxidative stress and the antioxidant actions of natural health products (NHPs) in the context of NAFLD and its associated disorders. The O2.- generating NADPH oxidase contributes to atherogenesis by facilitating macrophage induced vascular injury. In manuscript I, the plant alkaloid berberine effectively abolished NADPH oxidase mediated O2.- production in lipopolysaccharide stimulated macrophages. Real-time PCR analysis and siRNA transfection studies revealed that berberine mediated its effects through down-regulation of the oxidase’s catalytic subunit gp91phox. Berberine also restored the activity of the O2.- clearing enzyme superoxide dismutase (SOD). High fat diet (HFD) fed rodents are a popular model for investigating NAFLD pathogenesis. In manuscript II, folic acid supplementation significantly reduced HFD-induced hepatic oxidative stress and liver injury in mice. Folic acid decreased NF-kB/DNA binding, down-regulated NADPH oxidase gene expression, and inhibited the oxidase. The antioxidant activities of SOD and catalase were restored and the reduced to oxidized glutathione ratio (GSH:GSSG) was re-established with folic acid supplementation. Folic acid’s hepatoprotective antioxidant effects were associated with a marked improvement in liver histology. Homocysteine (Hcy) levels are perturbed in NAFLD, but the etiology is unclear. In manuscript III, HFD fed mice exhibited decreased Hcy levels. Real-time PCR and Western Immunoblotting analysis revealed that Hcy catabolising enzymes cystathionine-b-synthase (CBS) and cystathionine-g-lyase (CSE) were increased in the liver of these animals. The transsulfuration activities of these enzymes were elevated and coincided with enhanced hepatic hydrogen sulfide biosynthesis. Glutathione was maintained despite increased hepatic oxidative stress. Taken together, NHPs such as berberine and folate, and Hcy catabolising enzymes CBS and CSE, might have therapeutic potential for managing oxidative stress in NAFLD and its associated co-morbidities. / October 2015

Common observations of reduced gonad size and spawning inhibition in wild and laboratory raised fish exposed to pulp mill effluents indicate that reproductive neuroendocrine signalling pathways may be upset. This thesis supported the neuroendocrine disruption of reproduction hypothesis by identifying potential disruptors and targets where these impacts may occur. A mechanistic study of the in vivo fathead minnow (FHM) spawning assay used by industry to assess effluent quality showed that ovulation, but not milt production, was impaired. This finding supported the hypothesis that the neuroendocrine cascade that triggers ovulation may be disrupted. I hypothesized that neuroactive constituents previously described in effluents were originating in wood feedstocks and neuroactive extracts of hardwood and conifer feedstocks were identified. Phytochemicals associated with effluents were neuroactive. Structurally similar phenolic phytochemicals showed monoamine oxidase (MAO) inhibition, and resin acid diterpenes displayed glutamic acid decarboxylase (GAD) inhibition. Inhibitors of these enzymes may have impacts on the control of reproduction since MAO metabolizes dopamine, an inhibitor of the neuroendocrine reproductive axis, while GAD synthesizes -aminobutyric acid (GABA), a stimulator of this axis. Bioassay-guided fractionations of effluents and wood feedstocks identified that medium polar extracts of primary- and secondary-treated effluents and balsam fir feedstock contained high GAD inhibitory activity. This activity was associated with chemically complex fractions rather than single active principles. Advanced metabolomic comparison of medium polar extracts of feedstock and treated effluent identified 15 common plant metabolites, demonstrating that phytochemicals entering the mill in wood are surviving pulp production and effluent treatment processes and may be responsible for observed GAD inhibition. Discriminant metabolomics analysis identified 4-acetylpyridine, a novel compound to be described in effluents, as well as two other tentatively identified compounds, as chemical markers of GAD inhibitory effluent fractions. Five tentatively identified chemical markers and (+)-lariciresinol were found in inhibitory balsam fir feedstock fractions. Neuroendocrine pathways that control reproduction in fish, such as dopamine and GABA pathways, are also important drug targets for the treatment of neurological disorders in mammals; therefore these results also have implications for the development of natural health products from phytochemicals and tree extracts common to Canadian forests. By using an interdisciplinary approach (phytochemistry, neuroendocrinology, ecotoxicology), I was able to explore the various implications of my research on the fields of natural health products chemistry and aquatic toxicology.

This thesis applied a novel interdisciplinary approach for pharmacovigilance to examine the use of complementary and integrative medicine (CIM), focusing on herbal remedies, to manage pediatric attention-deficit hyperactivity disorder (ADHD). The safety and potential risk of herb-drug interactions in ADHD management were first evaluated through an assessment of available information on the safety and efficacy of natural health products (NHPs) commonly used by ADHD patients as a means of identifying knowledge gaps. A clinical questionnaire was administered to caregivers of pediatric patients with ADHD to determine the factors and related outcomes of CIM use, including adverse events. A systematic search was conducted to further identify clinical adverse events involving herbal remedies and ADHD drugs to determine causal links to herb-drug interactions. In vitro analysis of identified herbal remedies was conducted to determine their potential for pharmacokinetic interactions, specifically on carboxylesterase-1 (CES1) mediated metabolism. The presented research builds on otherwise scarce evidence of the safety of herbal remedies for ADHD, particularly with respect to herb-drug interactions and adverse events (AEs) associated with concurrent use of NHPs and ADHD prescription drugs. Beyond studies conducted on the pharmacokinetic safety of herbal remedies through the cytochrome P450 pathways that metabolize some ADHD drugs, including amphetamine, atomoxetine and guanfacine, few data were available for CES1, which metabolizes methylphenidate, the first line of drug used to manage ADHD. The clinical questionnaire revealed that 40% of patients had used CIM and confirmed the use of a variety of CIM. Moreover, the majority of CIM users were also concurrently taking ADHD medication, and eight mild adverse events were self-reported. The systematic search on the adverse event reporting system highlighted a potential NHP-drug interaction between methylphenidate and St. John’s wort, and the overall poor quality of NHP-related adverse event reports. As a follow-up from the adverse event results, various commercial St. John’s wort products showed variable inhibition of recombinant human CES1 in vitro. Although the concentration of marker phytochemicals was not correlated to inhibition, hyperforin showed stronger activity than hypericin and quercetin. The preliminary in vitro investigation revealed that the herbal remedies used by ADHD patients have the potential to interact with CES1 mediated metabolism, with Rhodiola rosea identified as the most potent inhibitor. Further investigation on various commercial products of Rhodiola rosea revealed both reversible and irreversible inhibition of recombinant CES1. However, the inhibition was not dependent on the concentration of marker phytochemicals, and rosarin, rosavin, rosin, and salidroside were not potent inhibitors of recombinant CES1. Moreover, a commercial Rhodiola rosea extract showed concentration-dependent inhibition of human liver microsome meditated metabolism of methylphenidate. Overall, results from this thesis suggest potential risk from use of NHPs concurrently with conventional medicine used to manage ADHD. Improved evidence and pharmacovigilance for the use of NHPs in a pediatric population is warranted.