2 - Agathosma betulina

Abstract

Agathosma betulina (P.J.Bergius) Pillans (Rutaceae), commonly known as ‘buchu’, is an evergreen, aromatic shrub that is naturally distributed in the Western Cape Province of South Africa. The plant has an extensive history of traditional use by indigenous South African cultures, where leaf infusions and tinctures are used medicinally for the treatment of stomach ailments, skin disorders, respiratory complaints and urinary tract infections. The essential oil of ‘buchu’ contains minor sulphur-containing compounds that contribute to its unique organoleptic properties, making it desirable for imparting blackcurrant notes used in the flavour and fragrance industries. Agathosma betulina has been extensively studied both in vitro and in vivo for potential antimicrobial, anti-inflammatory and anti-oxidant properties, among others. Herein, we document a quality control protocol for A. betulina based on the chromatographic profiling of the non-volatile and volatile fractions. Using a semi-automated high-performance thin-layer chromatography (HPTLC) system, ultra-performance liquid chromatography coupled to mass spectrometry and photodiode array detection (UPLC-MS-PDA), and gas chromatography coupled to mass spectrometry and flame ionisation detection (GC-MS-FID), the chemical profiles of six specimens of A. betulina were obtained. The aerial parts were harvested and divided into two portions; then methanol extraction of the first portion was performed to obtain the non-volatile fraction, and hydrodistillation of the second portion to yield the essential oil (volatile fraction). The non-volatile fractions were analysed using HPTLC and UPLC-MS-PDA, while the essential oils were profiled on HPTLC and GC-MS-FID. The crude methanol extracts, viewed under 366nm radiation following derivatisation, revealed the presence of hesperidin, rutin and diosmin in all the samples, using HPTLC. The presence of these marker compounds in A. betulina was confirmed by UPLC-MS-PDA, where peaks for rutin, diosmin and hesperidin were identified on the chromatograms. The HPTLC plate of the essential oils viewed under 254nm and white reflectance light following derivatisation, revealed the presence of diosphenol, which is the distinctive constituent in A. betulina essential oil. A typical A. betulina essential oil fingerprint characterised by high levels of diosphenol, pseudo-diosphenol, limonene, 1,8-cineole, menthone, isomenthone and trans-8-mercapto-p-menthan-3-one was obtained using GC-MS-FID.