Browsing by Author "Abdulla, Zohra Bibi"

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    Investigating the pharmacokinetics of a grape seed-derived proanthocyanidin extract in healthy human subjects.
    (Stellenbosch : Stellenbosch University, 2024-12) Abdulla, Zohra Bibi; Petersen-Ross, Kelly; Kellermann, Tracy; Smith, Carine; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Medicine. Division of Clinical Pharmacology.
    ENGLISH ABSTRACT: Introduction: Oxidative stress and inflammation have been implicated in the ageing phenotype associated with the development of non-communicable diseases (NCDs). Several pre-clinical and clinical studies have demonstrated the beneficial properties of grape seed-derived polyphenols in mediating oxidative stress. However, limited information on the bioavailability and pharmacokinetics of these products is available. This study consisted of the development and validation of a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to quantify catechin, epicatechin and procyanidin B1 as constituents of a grape seed derived supplement, OxiprovinTM in human plasma. Subsequently, the method was applied to an early phase clinical trial investigating the pharmacokinetics of these analytes following the administration of a single dose of 140 mg. Methods: Positive electrospray ionisation (ESI) was used to detect catechin, epicatechin and procyanidin B1 on a Shimadzu-8040 mass spectrometer with (+)-catechin-2,3,4-13C3 as an internal standard. A double liquid-liquid extraction (LLE) method was used to extract the analytes from human plasma using 50 mM ammonium bicarbonate in water and ethyl acetate in the first extraction step and ethyl acetate: acetone (10:1, v/v) in the second. Chromatographic separation was achieved on an Agilent XDB-C8 column (4.6 x 100 mm, 5 µm) with a gradient method at a flow rate of 0.600 mL/min over a total run time of 10 minutes. The mobile phases consisted of water + 0.1% formic acid (A) and methanol + 0.1% formic acid (B). The analyte to internal standard peak area ratios were plotted against the nominal concentrations to generate a calibration curve with a quadratic regression weighted by 1/c2, over the range 19.5-2 500 ng/mL for catechin and epicatechin and 39.1-2 500 ng/mL for procyanidin B1. In an early phase clinical trial, sixteen healthy volunteers (four male and 12 female) were recruited. Eight participants received a placebo and the other eight were administered a dose of 140 mg of OxiprovinTM. Blood samples were collected at 0 hours (baseline) and at 1, 2, 3, 6 and 24 hours post administration of the supplement, and the resulting plasma was stored at -80°C until analysis. Results and Discussion: During inter-day validations, the average accuracy of calibration standards ranged from 93.5-106.3% (% CV 2.6-7.1%) for catechin, 96.7-103.1% (% CV 4.2-11.7%) for epicatechin and 90.0-103.5% (% CV 3.8-9.9%) for procyanidin B1. For quality controls, the average accuracy ranged from 97.7-106.8% (% CV = 6.9-14.7%), 93.1-101.6% (% CV = 4.9-14.7%) and 96.0-110.1% (% CV = 10.0-19.8%) for catechin, epicatechin and procyanidin B1, respectively. Endogenous matrix effects were shown to have negligible effects on the quantification of the analytes when plasma samples from six different sources were analysed. The average recovery of catechin, epicatechin and procyanidin B1 was 78.7%, 70.6% and 52.4%, respectively. Furthermore, catechin, epicatechin and procyanidin B1 were found to be stable in plasma when subjected to three freeze-thaw cycles, on bench at room temperature for up to six hours and at -80°C for 27 days. All three analytes were also found to be stable in whole blood for up to 2 hours on bench at room temperature. Lastly, 2% haemolysis did not affect the reproducibility of the method. The validated method was used to quantify catechin, epicatechin and procyanidin B1 in the human plasma samples obtained from the trial. During the trial, no adverse events were reported, and the supplement was well-tolerated. Upon analysis, no analytes were detected in the plasma samples obtained at each time point. Conclusion: A LC-MS/MS method was successfully developed and validated according to United Staes Food and Drug Administration (FDA) and European Medicines Agency (EMA) guidelines for the quantification of catechin, epicatechin and procyanidin B1 in human plasma. The analytes were undetectable in the plasma samples collected at each time point during the trial. Due to impacts of metabolism, secondary metabolites may be detected at higher concentrations and using a more sensitive instrument may allow for lower assay ranges. Furthermore, future studies on urine or faecal samples may provide insight to the route of these parent compounds.