In vitro cytotoxicity of Siphonochilus aethiopicus in combination with selected fillers for tableting
The use of herbal medicines is currently experiencing a revival of sorts. However, this resurgence in popularity is casting a much needed light on safety issues relevant to the use thereof, since many herbal products remain untested with regards to their toxicological properties. Siphonochilus aethiopicus, or African ginger, is one of the most desired medicinal plants at South African muthi markets. Traditional uses vary from malaria to various inflammatory related conditions such as asthma and dysmenorrhoea. Though African ginger has shown auspicious potential in several pharmacological studies - possessing anti-inflammatory, anti-trypanosomal and antimalarial activity (to name only a few) — information relating to its cytotoxicity is limited. When considering the wide range of commercially available products containing this plant, this raises concern for consumer safety. The main aim of the current study is to investigate the in vitro cytotoxicity of several S. aethiopicus extracts, alone and in combination with chitosan and Pharmacel® 101 fillers for tableting, on human hepatocellular liver carcinoma (HepG2) and human epithelial colorectal adenocarcinoma (Caco-2) cell lines since they represent the oral route of administration. Organic extracts were prepared by solvent extraction and compared with an aqueous extract and traditional infusion. These extracts were, subsequently, characterised with ultra-performance liquid chromatography quadruple time of flight mass spectrometry (UPLC-Q-TOF/MS) through the identification of AG 1—4, previously identified marker compounds. Standard cytotoxicity assays with different endpoints were selected and included the tetrazolium reduction (MTT) and lactate dehydrogenase (LDH) assays, as well as flow cytometry with fluorescent annexin V / propidium iodide (PI). UPLC chromatograms of extracts revealed organic extracts (ethanol and diethyl ether) to contain all four isolated African ginger compounds (AG 1—4). Aqueous extracts only contained AG 1 and AG 2 in small amounts and did not contain AG 3 or AG 4. MTT assays proved organic extracts to reduce cell viability, despite interference, whereas aqueous extracts did not cause interference; nor did it reduce cell viability. LDH data also indicated only organic extracts to cause LDH release on both cell lines. During both assays Caco-2 cells proved to be less sensitive to the effects of the extracts, compared to HepG2 cells, since cell viability only decreased at considerably higher concentrations. Fillers seldom, if at all, caused a significant (p ≤ 0.05) alteration of the effects caused by extracts. Subsequently, flow cytometric analysis was performed on organic extracts alone. Results indicated a definite decrease in cell viability of both cell lines following exposure, with a concomitant increase in apoptotic and necrotic cell populations. Therefore, it can be concluded that aqueous extracts do not possess cytotoxic properties, whereas organic extracts caused apoptotic and necrotic cell death. Considering the difference in phytochemical composition of these extracts, one cannot help to consider the possibility of AG 3 and AG 4, compounds only present in organic extracts, to be partly responsible for the observed cytotoxicity. Consequently, it is suggested to further isolate the major compounds present in S. aethiopicus and investigate their individual cytotoxicity. Selectivity of crude extracts and isolated compounds should, moreover, be investigated. Finally, development of polymer nanoparticle formulations are furthermore recommended as it might reduce toxicity.
- Health Sciences