Method Development and Selection of Verbascoside-Rich Acanthus Species and their Isolation Using Centrifugal Partition Chromatography

Abstract

Verbascoside possesses antioxidant, anti-inflammatory, anti-cancer, neuroprotective, and anti-depressant activities. This compound can be found in plants from the Acanthaceae family.  However, the production of this compound varies significantly, depending on several factors. This study aims to develop an HPLC method for verbascoside quantification in Acanthus species, its isolation from verbascoside-Rich plant is also optimized to obtain the high purity verbascoside. Acanthus ebracteatus and Acanthus ilicifolius were extracted by sonication. The extracts were subjected to HPLC-UV analysis. It revealed varying verbascoside content between A. ebracteatus and A. ilicifolius across 22 different locations, with A. ebracteatus from Samut Songkhram exhibiting the highest content (0.18% w/w), and A. ebracteatus from Samut Sakhon exhibiting the lowest content (0.01% w/w) during March to August collection periods. Chromatographic analysis indicated differing resolutions, with the best resolution (5.79) observed in A. ebracteatus from Muang Nakhon Si Thammarat despite low verbascoside content (0.05% w/w), while the runner-up was A. ebracteatus from Samut Songkhram (5.50). Thus, verbascoside was isolated from A. ebracteatus from Samut Songkhram using Centrifugal Partition Chromatography (CPC). It successfully isolated verbascoside within 2 hours 15 minutes, with some loss observed during the process. Mass spectrometry confirmed verbascoside presence using a reference standard. The study found that 11 mg of verbascoside with a purity of 79% was isolated from 1 g of extract of A. ebracteatus. This research underscores the effectiveness of plant material selection and CPC in obtaining high amounts of verbascoside from natural sources, suggesting avenues for further enhancement in isolation techniques to maximize yield.