Danielle Julie Carrier

Clinical assessment of CYP2D6-mediated herb–drug interactions in humans: Effects of milk thistle, black cohosh, goldenseal, kava kava, St. John's wort, and Echinacea

Cytochrome P450 2D6 (CYP2D6), an important CYP isoform with regard to drug-drug interactions, accounts for the metabolism of approximately 30% of all medications. To date, few studies have assessed the effects of botanical supplementation on human CYP2D6 activity in vivo. Six botanical extracts were evaluated in three separate studies (two extracts per study), each incorporating 16 healthy volunteers (eight females). Subjects were randomized to receive a standardized botanical extract for 14 days on separate occasions. A 30-day washout period was interposed between each supplementation phase. In study 1, subjects received milk thistle (Silybum marianum) and black cohosh (Cimicifuga racemosa). In study 2, kava kava (Piper methysticum) and goldenseal (Hydrastis canadensis) extracts were administered, and in study 3 subjects received St. Johns wort (Hypericum perforatum) and Echinacea (Echinacea purpurea). The CYP2D6 substrate, debrisoquine (5 mg), was administered before and at the end of supplementation. Pre- and post-supplementation phenotypic trait measurements were determined for CYP2D6 using 8-h debrisoquine urinary recovery ratios (DURR). Comparisons of pre- and post-supplementation DURR revealed significant inhibition (approximately 50%) of CYP2D6 activity for goldenseal, but not for the other extracts. Accordingly, adverse herb-drug interactions may result with concomitant ingestion of goldenseal supplements and drugs that are CYP2D6 substrates.

Assessing the clinical significance of botanical supplementation on human cytochrome P450 3A activity: Comparison of a milk thistle and black cohosh product to rifampin and clarithromycin

Phytochemical‐mediated modulation of cytochrome P450 enzymes (CYPs) may underlie many herb‐drug interactions. This studys purpose was to assess the effects of milk thistle and black cohosh supplementation on CYP3A activity and compare them to a clinically recognized inducer, rifampin, and inhibitor, clarithromycin. Healthy volunteers were randomly assigned to receive a standardized milk thistle (900 mg) or black cohosh (80 mg) supplement for 14 days. Subjects also received rifampin (600 mg) and clarithromycin (1000 mg) for 7 days as positive controls for CYP3A induction and inhibition, respectively. Midazolam was administered orally before and after each supplementation and control period. The effects of milk thistle, black cohosh, rifampin, and clarithromycin on midazolam pharmacokinetics were determined using noncompartmental techniques. Unlike those observed for rifampin and clarithromycin, midazolam pharmacokinetics was unaffected by milk thistle or black cohosh. Milk thistle and black cohosh appear to have no clinically relevant effect on CYP3A activity in vivo.

Pre- and post-harvest processing of medicinal plants

Herbal medicine is used worldwide either as a sole treatment method or as part of a comprehensive treatment plan alongside orthodox methods of diagnosis and treatment. A survey reported that, in the USA, nearly one-sixth of women took at least one herbal product in 2000. Despite their widespread use, numerous reports show that the herbal products available to consumers are of variable quality. This disparity in quality of herbal preparations can be attributed to the fact that their production is complicated. To produce high-quality herbal products, attention must be paid to, among others, phytochemical variations due to plant breed, organ specificity, stages of growth, cultivation parameters, contamination by microbial and chemical agents, substitution, adulteration with synthetic drugs, heavy metal contamination, storage and extraction. This review focuses on organ specificity, seasonal variations, the effect of drying and storage, and the extraction of phytochemical constituents. Special emphasis is placed on the four most frequently used herbal products in the USA: echinacea, Ginkgo biloba , ginseng and St Johns Wort.

Purification of Resveratrol, Arachidin-1, and Arachidin-3 from Hairy Root Cultures of Peanut (Arachis hypogaea) and Determination of Their Antioxidant Activity and Cytotoxicity

Antioxidant stilbenoids, such as resveratrol, arachidin‐1, and arachidin‐3, have demonstrated beneficial effects on human health. Although resveratrol is commercially available, arachidin‐1 and arachidin‐3 are not, resulting in an opportunity to explore purification methods and to confirm biological activity. Recently, Arachis hypogaea hairy root cultures (produced via Agrobacterium rhizogenes‐mediated transformation) were reported to secrete stilbenoids into liquid growth media upon elicitation in quantities sufficient for commercial production. The purpose of this study was to purify substantial quantities of resveratrol, arachidin‐1, and arachidin‐3 from A. hypogaea hairy root cultures using centrifugal partition chromatography (CPC), determine the antioxidant activity of these compounds using the thiobarbituric acid reactive substances (TBARS) assay, and determine the cytotoxicity of the compounds using the 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide (MTT) assay. In a single run of CPC, resveratrol, arachidin‐1, and arachidin‐3 were separated to a purity of 97.1%, 97.0%, and 91.8%, respectively. Lipid oxidation was inhibited by a 27 and 7 μM dose for reference standards of resveratrol and arachidin‐1, respectively, while oxidation was not inhibited up to a 27 μM dose for reference standard of arachidin‐3. Oxidation was inhibited at a 14, 7, and 14 μM doses for CPC‐purified resveratrol, arachidin‐1, and arachidin‐3, respectively. Arachidin‐1 and arachidin‐3 demonstrated cytotoxicity at 27 and 55 μM in RAW 264.7 and HeLa cell lines, respectively; while resveratrol exhibited no cytotoxicity to either cell line. These results demonstrate the integration of a production and purification system for the manufacturing of A. hypogaea‐derived stilbenoids.

Extraction of nutraceuticals from milk thistle: I. Hot water extraction.

Milk thistle contains compounds that display hepatoxic protection properties. We examined the batch extraction of silymarin compounds from milk thistle seed meal in 50, 70, 85, and 100°C water as a function of time. After 210 min of extraction at 100°C, the yield of taxifolin was 1.2 mg/g of seed, a 6.2-fold increase over the results obtained in a Soxhlet extraction with ethanol on pretreated (defatted) seeds. Similarly, the yield of silychristin was 5.0 mg/g of seed, a 3.8-fold increase. The yields of silybinin A and silybinin B were 1.8 and 3.3 mg/g of seed, respectively, or roughly 30% of the Soxhlet yield. The ratios of the extracted compounds, and particularly the ratios at long extraction times, showed that the more polar compounds (taxifolin and silychristin) were preferentially extracted at 85°C, while the less polar silybinin was favored at 100°C.

Silymarin extraction from milk thistle using hot water

Hot water is attracting attention as an extraction solvent in the recovery of compounds from plant material as the search for milder and “greener” solvents intensifies. The use of hot water as an extraction solvent for milk thistle at temperatures above 100°C was explored. The maximum extraction yield of each of the silymarin compounds and taxifolin did not increase with temperature, most likely because significant compound degradation occurred. However, the time required for the yields of the compounds to reach their maxima was reduced from 200 to 55 min when the extraction temperature was increased from 100 to 140°C. Severe degradation of unprotected (plant matrix not present) silymarin compounds was observed and first-order degradation kinetics were obtained at 140°C.

Biorefinery co-products : phytochemicals, primary metabolites and value-added biomass processing

DESCRIPTION In order to successfully compete as a sustainable energy source, the value of biomass must be maximized through the production of valuable co-products in the biorefinery. Specialty chemicals and other biobased products can be extracted from biomass prior to or after the conversion process, thus increasing the overall profitability and sustainability of the biorefinery. Biorefinery Co-Products highlights various co-products that are present in biomass prior to and after processing, describes strategies for their extraction , and presents examples of bioenergy feedstocks that contain high value products.

Milk Thistle, Silybum marianum (L.) Gaertn., Flower Head Development and Associated Marker Compound Profile

ABSTRACT Flower head development and marker compound changes were examined for milk thistle grown under dryland conditions during the 1998 and 1999 growing seasons. Flower heads harvested at the early flowering, mid-flowering, late flowering, and dehiscing (seed development) growth stages had average seed/ovule weights of 5 mg, 13 mg, 21 mg, and 26 mg, respectively. At the time of harvest, the plants contained flower heads at all stages of development. During the 1999 harvest, the average plant produced 2.0 g, 2.9 g, and 3.4 g of dry seeds per flower head, corresponding to mid-flowering, late flowering, and dehiscing development stages of the flower. The silymarin and fixed-oil content for the 1999 seeds were 1 mg, 14 mg, and 24 mg of silymarin and 15 mg, 168 mg, and 252 mg of lipid/g dry weight for mid-flowering, late flowering, and dehiscing development stages, respectively. The highest silymarin yield was obtained from late flowering and dehiscing flower heads in early September of both test years.

EXTRACTION OF NUTRACEUTICALS FROM MILK THISTLE: PART II. EXTRACTION WITH ORGANIC SOLVENTS

Seeds from milk thistle (Silybum marianum Gaert L.) contain flavanolignan and dihydroflavanol compounds that have interesting and important therapeutic activities. The recovery of these silymarin compounds generally involves a two-step defatting and extraction process using organic solvents. This study examined the batch, single-stage extraction of whole and defatted seeds using ethanol, methanol, acetonitrile, and acetone as the solvents. In extracting defatted milk thistle seeds with organic solvents, extraction with ethanol resulted in the highest silymarin yield, although some potential degradation was observed. The maximum yields of taxifolin, silychristin, silydianin, silybinin A, and silybinin B in ethanol were 0.6, 4.0, 0.4, 4.0, and 7.0 mg/g of defatted seed, respectively. However, if silybinin A were the diastereoisomer of choice, methanol would be the preferred extraction solvent because it yielded the highest silybinin A to silybinin B ratio. Interestingly, lipid removal is an important extraction step, because defatted material yields twice the silymarin concentration.

Switchgrass Water Extracts: Extraction, Separation and Biological Activity of Rutin and Quercitrin

Switchgrass (Panicum virgatum L.) has recently received significant attention as a possible feedstock for the production of liquid fuels such as ethanol. In addition, switchgrass may also be a source of valuable co-products, such as antioxidants, and our laboratory recently reported that switchgrass contains policosanols and alpha-tocopherol. Motivation for this work began when a switchgrass sample was extracted with water at 50 degrees C and was then tested for low-density lipoprotein (LDL) oxidation inhibition activity using the Thiobarbituric Acid Reactive Substances (TBARS) assay. The TBARS results showed that the switchgrass water extracts inhibited LDL oxidation by as much as 70% in comparison to the control. Liquid chromatography coupled with mass spectrometry (LC-MS) and high performance liquid chromatography (HPLC) were used to identify the compounds that were responsible for LDL oxidation inhibition activity as flavonoids: quercitrin (quercetin-3-O-rhamnoside) and rutin (quercetin-3-O-rutinoside). To maximize flavonoid concentrations, switchgrass was then extracted with water and 60% methanol at different temperatures. The 60% methanol treatment resulted in higher rutin and quercitrin yields when compared to water-only extraction; however, the use of this solvent would not be practical with current biorefinery technology. Centrifugal partition chromatography (CPC) was then used to purify rutin and quercitrin from the switchgrass water extract, which were then tested via the TBARS assay and shown to exhibit lipid peroxidation inhibition activity similar to that obtained with pure flavonoid standards. This is the first report on the presence of rutin and quercitrin in switchgrass. The results support the extraction of viable coproducts from switchgrass prior to conversion to liquid fuel.