Bioassay and RP-HPLC Method Development for the Analysis of Extracts and Herbal Products Containing Anthocleista nobilis

Abstract

Anthocleista nobilis is a common constituent in numerous conventional medications in West Africa. The stem bark of A. nobilis is known to contain brucine and is used to treat intestinal parasites, stomachaches, gonorrhoea, wounds, etc. The extensive use has led to high market value and adulteration of their herbal products. In this work, the antioxidant properties of extracts of A. nobilis are verified, and a validated RP-HPLC method is used to estimate the brucine content of extracts and products containing A. nobilis. Stem bark extracts from ethyl acetate, 96% v/v ethanol, 70% v/v ethanol, and water of A. nobilis were investigated for phytochemical content using standard methods and their antioxidant activity using DPPH and phosphomolybdenum assays. An RP-HPLC method was developed and validated using brucine as a reference standard. The optimized conditions of the developed RP-HPLC method include a Supelcosil C18 column with dimensions 150 × 4.6 mm and particle size of 3 μm, a mobile phase comprising of MeOH : 0.1% v/v HCOOH (65:35 %v/v ) which was injected at a flow rate of 0.8 mL/min, and an injection volume of 50 μL. The wavelength of detection was 274 nm. The developed method was validated as per ICH guidelines. The common phytochemicals among the various extracts were tannins and alkaloids. All extracts exhibited a reasonable antioxidant activity in the DPPH and phosphomolybdenum assays, with the ethanol extract recording the highest activity of 27.681 μg/mL and 696.7452 μg/g AAE, respectively. The content of brucine in the extracts was determined to be 0.0177–0.1259 × 10^–3% w/v, whereas the herbal products tested had a content of 0.8950−2.5013 × 10^–3% w/v. These levels were below the toxicity threshold of brucine. The developed method could be used for the routine quality control of A. nobilis extracts and formulations.