Pharmacokinetics and Drug Interactions With Milk Thistle

BIOAVAILABILTY STUDY:

1. Clinical Protocol

Screening/Informed Consent: Sixteen subjects will be recruited for the clinical protocol. All protocols, consent forms, and HIPAA forms shall be IRB-approved prior to initiating any study procedure or recruitment. At a screening/informed consent visit, the study design, purposes and inclusion/exclusion criteria will be explained to each prospective subject. Written informed consent will be obtained prior to commencing any study procedures. After informed consent is obtained, medical examinations will be performed along with baseline serum chemistries, complete blood count, electrocardiogram, urinalysis, urine drug screen, nicotine/cotinine, and urine pregnancy test (females). Subjects will be asked about their diets including alcohol use and the use of herbal medications/dietary supplements. If a potential subject has been consuming any herbal or dietary supplements a two-week period of abstinence from supplement use will be necessary before beginning the study. Additionally, subjects will refrain from use of honey or propolis-containing products (due to presence of chrysin and potential assay interference), and artichokes, or artichoke-containing foods that also contain taxifolin (a constituent found in MT).

Single- and Multiple-Dose Silymarin Pharmacokinetics: Subjects will arrive at the MUSC GCRC in the morning where they will remain for approximately 10-hours for each of 4 separate blood drawing phases of the study. After checking in, and under medical supervision, a modification of a modified Treatment Emergent Symptoms Scale (TESS) will be administered and a registered nurse will place an indwelling venous catheter in each subject's arm to facilitate serial blood sampling. At 8 AM after urinary void and collection of an aliquot for creatinine analysis, subjects will take one 175 mg capsule of MT standardized to 80% SM (140 mg of SM). The rationale for product choice is discussed below. Subjects will be in a fasted state for 4 hours following MT administration to reduce any effect of food on absorption. A total of 12 blood samples (10 ml each) will be taken over the next 24-hour period. All samples will be drawn in heparinized tubes and stored on ice until centrifugation at 4°C. As flavonoids are notorious for their instability in biological matrices, no sample will remain on ice for longer than 15 minutes prior to centrifugation. Time points of blood collection will be immediately prior to the dose of MT (0 time point), and at 0.5 hours, 1.0, 1.5, 2.0, 3.0, 4.0, 6.0, 8.0, 12, and 24 hours. A second TESS will be administered to the subjects 2 hours post-dosing, at a time generally expected to represent silybin Cmax. Subjects will be asked to return for 4 brief visits for collection of single blood samples through a separate venipuncture at time points of 24, 36, and 48 hours. This extended sampling will provide adequate PK data to determine Cmax, AUC, terminal t1/2, and Clo of SM flavonoids. Standard meals will be served by the registered dietician at the GCRC and will not include any other sources of flavonoids. The composition and amount of food eaten throughout the day will be recorded. Plasma will be acidified with 10 ml of 1M acetic acid/ml plasma and frozen immediately at -70°C. This single dose assessment of silymarin pharmacokinetics will be conducted at three dosage levels of MT supplement in an attempt to span the usual therapeutic range of MT and will be performed on three distinct occasions separated by no less than a 7-day wash-out period. The doses of MT used will be 175 mg, 350 mg and 525 mg administered orally with 200 mL of water as one, two, and three capsules, respectively. Again, these doses as well as product could be changed if a more standardized supplement is made available through NIH/NCCAM or some other source. The highest dose of MT supplement proposed for oral administration, 525 mg (420 mg SM) does not pose any undue risk to subjects since SM doses of 420 mg to 800 mg has been administered to many patients with hepatic dysfunction in clinical trials with few side effects reported (NLM, 2000; Saller et al., 2001).

Following the last single dose assessment and an ensuing 7-day minimum wash-out period, subjects will initiate treatment of a 28-day treatment exposure to the MT supplement 175 mg (140 mg SM) at the generally recommended and clinically studied dosage of one capsule thrice daily (NLM, 2000). A 28-day course is selected to assure steady-state is reached and to surpass the duration of any other published pharmacokinetic studies of SM in human subjects which have generally been of brief duration. In addition, this extended dosing assessment will allow for the gathering of pilot data on the antioxidant potential of SM via measurements of urinary isoprostanes. Finally, an assessment of treatment-emergent adverse effects and tolerability will be made. Subjects will report to the outpatient GCRC on 4 separate occasions to pick up a 7-day supply of SM at which time a TESS will be administered. At the initial visit to the GCRC to obtain the first week's supply of MT, subjects will provide a urine sample for baseline measurement of urinary isoprostanes and creatinine. After 28 days of treatment subjects will report to the GCRC for the determination of steady-state concentrations of SMs in plasma and urine. A blood draw will be performed immediately prior to dosing with the MT (Ctrough) and 0.5, 1.0, 2.0, 3.0 and 4.0 and 8.0 following MT dosing. Based upon available published data, these timepoints should be sufficient to capture the (Cmax) for all flavonoids of interest despite interindividual variability. Additionally, subjects will provide a morning urine sample for analysis of urinary isoprostane and creatinine.