Exploring four South African Croton species for potential anti-inflammatory properties: in vitro activity and toxicity risk assessment.

ETHNOPHARMACOLOGICAL RELEVANCE: The African Continent harbours approximately 26 Croton species. Many Croton species are used in traditional medicine in southern Africa to treat a variety of ailments including malaria, tuberculosis, microbial infection and inflammation. Considering the high diversity of the genus Croton, the ethnopharmacological information available on southern African species is rather limited. Furthermore, the potential for novel anti-inflammatory drug scaffolds has not previously been investigated. AIM OF THE STUDY: The aim of the study was to evaluate the potential of four South African Croton species extracts (Croton gratissimus, Croton pseudopulchellus, Croton sylvaticus, and Croton steenkampianus) for anti-inflammatory activity targeting the TLR4 signalling pathway and to assess the potential risk for hepatotoxicity and genotoxicity using an in vitro cellomics approach. MATERIAL AND METHODS: Leaf extracts of C. gratissimus, C. pseudopulchellus, C. sylvaticus and C. steenkampianus were prepared using methanol and chloroform (1:1, v/v). The anti-inflammatory activity was determined using LPS induced nitric oxide production in RAW 264.7 macrophages, while the hepatotoxicity and genotoxicity was evaluated using multi-parameter end point analysis in C3A and Vero cells, respectively. Mitochondrial membrane potential, mitochondrial mass, oxidative stress, lysosomal content and lipid accumulation were used as markers to assess the risk for hepatotoxicity. RESULTS: All four species attenuated nitric oxide production with negligible cytotoxicity. However, C. gratissimus yielded the most favorable profile. Cell density was significantly reduced in both C3A and Vero cells with the C. gratissimus extract providing a suitable toxicity profile amenable to further high content analysis. While there was no meaningful effect on mitochondrial dynamics, a strong dose dependent increase in lipid content, paralleled by an expansion of the lysosomal compartment, identifies a potential risk for steatosis. Risk for genotoxicity was investigated using the micronucleus assay which revealed a dose dependent increase in micronuclei formation. Changes in nuclear morphology and cell ploidy further strengthens the associated risk for genotoxicity and suggests the extract from C. gratissimus may function as an aneugen. Collectively, the data demonstrates that although the selected species possess anti-inflammatory components, the risk for possible hepatotoxic and genotoxic side effects may negate their prospect towards further drug development.

A review of biological activities and phytochemistry of six ethnomedicinally important South African Croton species.

ETHNOPHARMACOLOGICAL RELEVANCE: The genus Croton (Euphorbiaceae) encompasses 1300 species, which consist of a variety of trees and shrubs distributed across the world. About 26 species are harboured on the African continent. This genus plays an essential role in African folk medicine. Croton species are traditionally used for the treatment of many diverse conditions such as diabetes, malaria, sexually transmitted diseases, cancer, inflammation, fever, digestive problems and fungal infections. AIM OF THE REVIEW: To provide a comprehensive overview of the ethnobotany, traditional uses, phytochemistry and biological activities of six selected southern Africa Croton species; C.gratissimus Burch., C. megalobotrys Müll.-Arg., C. menyhartii Gȕrke, C. pseudopulchellus Pax, C. steenkampianus Gerstner and C. sylvaticus Schltdl.). MATERIALS AND METHODS: Various electronic databases, namely Google, Google Scholar, Scopus, ScienceDirect, Biomed Central and Pubmed, were used to search for information related to the traditional uses, chemistry and pharmacology of Croton species. Books were also consulted to collect all pertinent information. RESULTS AND DISCUSSION: The ethnopharmacology, phytochemistry and biological activities of southern African Croton species are reviewed. The literature revealed that Croton species are trusted traditional medicines for the treatment of microbial infections and malaria. The non-volatile components of Croton species include flavonoids, terpenoids and alkaloids, while the volatile constituents comprise mainly of monoterpenes (α-phellandrene, α-pinene and 1,8-cineole) and sesquiterpenes (caryophyllene oxide). Most of the reported biological activities (anti-oxidant, antimicrobial, anticancer and antimalarial) were based on in vitro assays and were accredited to various extracts. However, both in vitro and in vivo studies, linking the reported activities to specific compounds, are still lacking. CONCLUSIONS: Croton species are used in traditional medicine to treat a range of ailments, and various in vitro biological activities have been investigated, with some extracts exhibiting good activity that could be considered for further investigation. The in vitro activities obtained seem to justify the use of Croton species in traditional medicine. Data on in vivo studies are scarce and studies usually focused on a single collection. The need to establish a quality control protocols for the standardisation of these herbal drugs is also important.

Chemotypic variation of non-volatile constituents of Artemisia afra (African wormwood) from South Africa.

Artemisia afra (African wormwood) is a popular medicinal plant of southern Africa and is an excellent candidate for commercialisation. This current study was aimed at exploring the phytochemistry and chemical variation of non-volatile compounds within wild populations of A. afra, and developing chromatographic quality control protocols for raw materials based on the identification of marker compounds. Chromatographic data, from samples representing 12 distinct populations, were obtained using liquid chromatography-mass spectrometry. An untargeted chemometric approach revealed three clusters. Marker compounds for each cluster, revealed through discriminant analysis, were isolated and identified using NMR spectroscopy, as acacetin (1) (Group 1), chrysoeriol (2) (Group 2), and 3,5-di-O-caffeoylquinic acid (3) and scopoletin (4) (Group 3). In addition, (3) and rutin (5), (both reported for the first time from A. afra), and (1), (2), (4) and 4-caffeoylquinic acid (6) were established as reliable markers for species identification, since they were abundant in most samples. Quantitative analysis using a validated method established (4) as the dominant compound in the samples (1080-19,600 µg/g dry weight (d.w.)), followed by (5) (49.5-2490 µg/g d.w.). A high performance thin layer chromatography (HPTLC) method was developed. The Rf values and colours of the bands corresponding to the marker compounds were recorded so that these compounds could be easily identified for quality control purposes. Multivariate analysis of the data using the rTLC online application confirmed the presence of different chemical groupings within the samples. It was deduced that quantitative, rather than qualitative differences, characterised the samples. Future research should focus on comparing the efficacy of the various chemical clusters in multi-target biological assays aligned to the traditional use of the plant.

Constituents of cinnamon inhibit bacterial acetyl CoA carboxylase.

Cinnamon bark ( CINNAMOMUM ZEYLANICUM) is used extensively as an antimicrobial material and currently is being increasingly used in Europe by people with type II diabetes to control their glucose levels. In this paper we describe the action of cinnamon oil, its major component, TRANS-cinnamaldehyde, and an analogue, 4-hydroxy-3-methoxy- TRANS-cinnamaldehyde against bacterial acetyl-CoA carboxylase in an attempt to elucidate the mechanism of action of this well-known antimicrobial material. These natural products inhibited the carboxyltransferase component of ESCHERICHIA COLI acetyl-CoA carboxylase but had no effect on the activity of the biotin carboxylase component. The inhibition patterns indicated that these products bound to the biotin binding site of carboxyltransferase with TRANS-cinnamaldehyde having a K (i) value of 3.8 ± 0.6 mM. The inhibition of carboxyltransferase by 4-hydroxy-3-methoxy- TRANS-cinnamaldehyde was analyzed with a new assay for this enzyme based on capillary electrophoresis. These results explain, in part, the antibacterial activity of this well-known antimicrobial material.