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This paper describes an investigation into the accumulation and distribution of Cu and Pb in Helichrysum splendidum, to assess the potential use of the plant in the phytoremediation of Cu- and Pb-contaminated soils. Plants of the same age were exposed to soils artificially contaminated with 10, 50, 100, 500 and 1000 mg/L Cu(II) and Pb(II). Although plant growth was not affected by the presence of Cu, high concentrations of Pb (> 500 mg/L) stunted the plants. Chlorophyll production was slightly reduced when plants were exposed to high concentrations of Cu (> 500 mg/L), whereas all the plants growing in Pb-contaminated soils indicated low levels of chlorophyll. Significant amounts of Cu and Pb were accumulated in the roots, with corresponding bioconcentration factors of > 1. Translocation of Cu to the shoots was restricted for plants growing in > 500 mg/L Cu-treated soils, whereas Pb was translocated to the shoots for all treatments, with translocation factors > 1. The Cu and Pb tolerance mechanism was ascribed to chelation and free radical scavenging, through the production of phenolic compounds. Helichrysum splendidum is able to adapt to and ameliorate soils contaminated with low to high levels of Cu(II) and Pb(II) and has potential for application in phytoextraction and phytostabilization of contaminated soils.

Chromones are naturally occurring phenolic compounds that are universally present in a healthy human diet. To date, thousands of chromone derivatives, including chromone glycosides have been discovered, both from natural and synthetic origin. Many plant genera, including Aloe, Aquilaria, Cassia, Hypericum and Polygonum, are rich sources of chromones. Several genera of fungi, such as Aspergillus, Mycoleptodiscus, Orbiocrella, and Penicillium produce bioactive chromones. This class of compounds is mainly associated with anti-oxidant, antimicrobial, anticancer and anti-inflammatory activities. Various in vitro, in vivo and clinical studies have established the role of chromones in alleviating allergies, reducing oxidative damage, inhibiting cancer, infections and inflammation, and for treating neurological and psychiatric disorders. This review focuses mainly on natural chromones, and elaborates on their nutritional benefits, pharmacological activities, and mechanisms of action. Various scientific databases, including PubMed, Scopus and Google Scholar, were used to find relevant literature on chromones covering all available literature up to, and including the year 2019. This critical analysis of completed research regarding the nutraceutical properties of various chromone derivatives may provide new insight to scientists working in related fields.

Fumonisins are common contaminants of maize. The neutral red assay, using the RTL-W1 trout liver cell line, and the fish embryo test (FET), using zebrafish, were selected to assess the effect of pH on the cytotoxicity, acute toxicity and teratogenicity of fumonisin B1 (FB1). The results demonstrated that FB1 exerts low cytotoxicity towards RTL-W1 cells without pH adjustment (IC50 1 746 μM), and no cytotoxicity after pH-adjustment to physiological conditions. The LC50 value for FB1 in the FET (1 058 μM at 48 h) confirmed low acute toxicity. Adjusting the pH to physiological conditions reduced the acute toxicity of FB1 towards zebrafish embryos, emphasising the importance of acidity/basicity of the medium in toxicity testing. Hydrolysed FB1 was less toxic than FB1 (LC50 2 690 μM at 48 h), and neither were teratogenic towards zebrafish embryos. Results confirm that the FET may account for effects not observable in cell cultures.

Acacetin (5,7-dihydroxy-4-methoxyflavone) is a naturally occurring flavonoid, known to possess numerous pharmacological properties, including neuroprotective, cardioprotective, anticancer, anti-inflammatory, antidiabetic, and antimicrobial activities. The molecule is widespread throughout the Plant Kingdom, but is particularly abundant in the Asteraceae, Euphorbiaceae, Passifloraceae, Lamiaceae and Malvaceae families. It has been shown to exhibit strong inhibitory effects against glutathione reductase, cyclo-oxygenase, acetylcholinesterase, aldose reductase and xanthine oxidase enzymes. A comprehensive overview of the potential therapeutic applications of acacetin was prepared after collating selected peer-reviewed papers, accessed from online scientific databases, specifically PubMed, Web of Science and Scopus. The mechanism of action, dose forms and structure-activity relationships of the molecule are some of the aspects discussed in the review, to facilitate further research and use of this molecule.

Organochlorine pesticides (OCPs) and phthalates are amongst the most emphasized man-made environmental contaminants, due to their prevalence, persistence and potential to induce adverse effects in organisms. In addition to prevailing industrial and domestic activities, their presence in the environment is exacerbated by leaching from associated materials, run-off and emissions. The Loskop Dam in the Mpumalanga Province, South Africa, is located on the Olifants River, which flows through a hub of industrial and agricultural activities. Research aimed at monitoring the levels of OCPs and phthalate contamination in South Africa, particularly in the Olifants catchment, has been limited and is restricted to short-term monitoring. In this study, the spatial and temporal variations of 21 OCPs and 7 phthalates in water and sediments from Loskop Dam were evaluated over a 3-year period (2015–2017). Annual average OCP levels ranged from 0.013±0.006 to 0.36±0.13 μg/L in water and from below the limit of detection to 2.4±1.2 µg/kg in sediments. Phthalates were present at average concentrations ranging from 0.023±0.041 to 2.1±1.5 μg/L and 20±0.63 to 55±6.9 µg/kg in water and sediments, respectively. The levels of OCPs and phthalates were greatest in autumn, attributed to higher input sources and concentration effects due to lower water levels. Phthalates were present at greater quantities at the inlet, while OCP accumulation was observed at the upper reaches of the dam. This may be an indication of the differences in input sources and translocation of these compounds, related to the topography of the dam and variability in water flow rates. These findings are important in highlighting the environmental and health concerns that may arise in the Olifants catchment area. After considering the outcomes of this study, we propose the implementation of regular and stringent monitoring strategies, which include surveys of OCPs and phthalates for Loskop Dam and similar water systems globally.

The South African Salvia species, Salvia africana-lutea, S. lanceolata and S. chamelaeagnea, are widely used to treat fever and inflammation associated with skin and lung infections. The aim of this study was to explore the non-volatile secondary metabolites and the phytochemical variation within these lesser known species, to support product development and commercialisation. Chemical profiles of the methanol extracts of 81 wild-harvested samples were obtained using ultra performance-quadrupole-Time-of-Flight-mass spectrometry (UPLC-qToF-MS). Forty-one compounds, including caffeic acid, rosmarinic acid, carnosol, carnosic acid and ursolic acid, were detected and confirmed across the three species. Nineteen compounds were tentatively identified of which 14 have not been reported in these species. Principal component analysis revealed distinct clusters corresponding to the three species, confirming chemical differences. Marker compounds for each species were revealed using orthogonal projection to latent structures-discriminant analysis. Further chemometric analysis reflected a degree of intraspecies variation, although the chemistry within populations was mostly conserved. Potential chemotypes for each species were identified through unique compounds associated with each group. The concentrations of medicinally important metabolites, namely, rosmarinic acid, carnosol, carnosic acid and ursolic acid, were determined, using validated UPLC-PDA methods. Ursolic acid was present at levels up to 38.2 mg/g, confirming that these species are a rich source of this compound. No similar studies combining liquid chromatography with chemometric analysis, and utilising a large sample size from various habitats, have been reported for these three Salvia species. The results will guide selection of cultivars with the best attributes for the intended therapeutic application, thereby protecting wild populations from over-exploitation.

Terminalia sericea is used throughout Africa for the treatment of a variety of conditions and has been identified as a potential commercial plant. The study was aimed at establishing a high-performance thin layer chromatography (HPTLC) chemical fingerprint for T. sericea root bark as a reference for quality control and exploring chemical variation within the species using HPTLC metabo3lomics. Forty-two root bark samples were collected from ten populations in South Africa and extracted with dichloromethane: methanol (1:1). An HPTLC method was optimized to resolve the major compounds from other sample components. Dichloromethane: ethyl acetate: methanol: formic acid (90:10:30:1) was used as the developing solvent and the plates were visualized using 10% sulfuric acid in methanol as derivatizing agent. The concentrations of three major bioactive compounds, sericic acid, sericoside and resveratrol-3-O-&beta, rutinoside, in the extracts were determined using a validated ultra-performance liquid chromatography-photodiode array (UPLC-PDA) detection method. The rTLC software (written in the R-programming language) was used to select the most informative retardation factor (Rf) ranges from the images of the analysed sample extracts. Further chemometric models, including principal component analysis (PCA) and hierarchical cluster analysis (HCA), were constructed using the web-based high throughput metabolomic software. The rTLC chemometric models were compared with the models previously obtained from ultra-performance liquid chromatography coupled with mass spectrometry (UPLC-MS). A characteristic fingerprint containing clear bands for the three bioactive compounds was established. All three bioactive compounds were present in all the samples, although their corresponding band intensities varied. The intensities correlated with the UPLC-PDA results, in that samples containing a high concentration of a particular compound, displayed a more intense band. Chemometric analysis using HCA revealed two chemotypes, and the subsequent construction of a loadings plot indicated that sericic acid and sericoside were responsible for the chemotypic variation, with sericoside concentrated in Chemotype 1, while sericic acid was more abundant in Chemotype 2. A characteristic chemical fingerprint with clearly distinguishable features was established for T. sericea root bark that can be used for species authentication, and to select samples with high concentrations of a particular marker compound(s). Different chemotypes, potentially differing in their therapeutic potency towards a particular target, could be distinguished. The models revealed the three analytes as biomarkers, corresponding to results reported for UPLC-MS profiling and thereby indicating that HPTLC is a suitable technique for the quality control of T. sericea root bark.

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.

Terminalia sericea Burch. ex. DC. (Combretaceae) is a popular remedy for the treatment of infectious diseases. It is widely prescribed by traditional healers and sold at informal markets and may be a good candidate for commercialisation. For this to be realised, a thorough phytochemical and bioactivity profile is required to identify constituents that may be associated with the antibacterial activity and hence the quality of raw materials and consumer products. The aim of this study was to explore the phytochemistry and identify the antibacterial constituents of T. sericea root bark, using a metabolomic approach. The chemical profiles and antibacterial activities of 42 root bark samples collected from three districts in the Limpopo Province, South Africa, were evaluated. Dichloromethane:methanol (1:1) extracts were analysed using ultraperformance liquid chromatography (UPLC)-mass spectrometry (MS), and chemometric models were constructed from the aligned data. The extracts were tested against Bacillus cereus (ATCC 11778), Staphylococcus epidermidis (ATCC 12223), Staphylococcus aureus (ATCC 25923), Escherichia coli (ATCC 8739), Klebsiella pneumoniae (ATCC 13883), Pseudomonas aeruginosa (ATCC 27853), Shigella sonnei (ATCC 9292) and Salmonella typhimurium (ATCC 14028), using the minimum inhibition microdilution assay. Nine compounds, sericic acid, sericoside, resveratrol-3-O-&beta, rutinoside, ellagic acid, flavogallonic acid dilactone, methyl-flavogallonate, quercetin-3-(2&prime, &prime, galloylrhamnoside), resveratrol-3-(6&prime, galloyl)-O-&beta, d-glucopyranoside and arjunetin, were isolated from the root bark. All the compounds, with the exception of sericic acid, sericoside and resveratrol-3-O-&beta, rutinoside, were isolated for the first time from the root bark of T. sericea. Chemometric analysis revealed clustering that was not population specific, and the presence of three groupings within the samples, characterised by sericic acid, sericoside and an unidentified compound (m/z 682/4.66 min), respectively. The crude extracts from different populations displayed varied antibacterial activities against S. typhimurium (minimum inhibitory concentrations (MICs) 0.25&ndash, 1.0 mg/mL), but similar activity towards Bacillus cereus (1.0 mg/mL). Several compounds present in the root bark were highly active towards all or most of the pathogens tested, but this activity was not reflected by the chemical profiles of extracts prepared from the individual samples. Among the pure compounds tested, only flavogallonic acid dilactone and methyl-flavogallonate exhibited broad-spectrum activity. A biochemometric analysis indicated that there was no consistent association between the levels of phytochemicals and the activity of the active or non-active extracts. Although it was deduced that the major constituents of T. sericea root bark contributed to the chemotypic variation, further investigation of the interactions of compounds present in the root bark may provide antibacterial efficacies not evident when examining compounds singularly. The data reported herein will provide information that is fundamentally important for the development of quality control protocols.

Sceletium tortuosum (Aizoaceae) is widely recognised for the treatment of stress, anxiety and depression, as well as for obsessive compulsive disorders. A comprehensive intraspecies chemotypic variation study, using samples harvested from two distinct regions of South Africa, was done using both proton nuclear magnetic resonance (1H-NMR) spectroscopy of methanol extracts (N = 145) and ultra performance liquid chromatography-mass spectrometry (UPLC-MS) of acid/base extracts (N = 289). Chemometric analysis of the 1H-NMR data indicated two main clusters that were region-specific (Northern Cape and Western Cape provinces). Two dimensional (2D) NMR was used to identify analytes that contributed to the clustering as revealed by the S-plot. The sceletium alkaloids, pinitol and two alkylamines, herein reported for the first time from S. tortuosum, were identified as markers that distinguished the two groups. Relative quantification of the marker analytes conducted by qNMR indicated that samples from the Northern Cape generally contained higher concentrations of all the markers than samples from the Western Cape. Quantitative analysis of the four mesembrine alkaloids using a validated UPLC-photo diode array (PDA) detection method confirmed the results of qNMR with regard to the total alkaloid concentrations. Samples from the Northern Cape Province were found to contain, on average, very high total alkaloids, ranging from 4938.0 to 9376.8 mg/kg dry w. Regarding the Western Cape samples, the total yields of the four mesembrine alkaloids were substantially lower (averages 16.4–4143.2 mg/kg). Hierarchical cluster analysis of the UPLC-MS data, based on the alkaloid chemistry, revealed three branches, with one branch comprising samples primarily from the Northern Cape that seemed somewhat chemically conserved, while the other two branches represented mainly samples from the Western Cape. The construction of an orthogonal projections to latent structures-discriminant analysis model and subsequent loadings plot, allowed alkaloid markers to be identified for each cluster. The diverse sceletium alkaloid chemistry of samples from the three clusters may facilitate the recognition of alkaloid profiles, rather than individual compounds, that exert targeted effects on various brain receptors involved in stress, anxiety or depression.

Ethnopharmacological relevance Khat, the leaves of Catha edulis, is used as a “natural amphetamine-like” stimulant in eastern and southern Africa, as well as in the Arabian Peninsula. Leaves are masticated to elicit a state of euphoria. Although the psychostimulatory effects of the leaves are attributed to the presence of phenylpropylamino alkaloids ( i.e. cathinone, cathine and norephedrine), the extent of permeation of these alkaloids across the oral and intestinal mucosa has not been established. Materials and methods Cathinone was isolated in the form of the oxalate salt from young buds, following acid-base extraction. High performance countercurrent chromatography (HPCCC) was used to isolate cathine and norephedrine, following borohydride reduction of a mixture of the three alkaloids. The in vitro permeability of these three alkaloids in their pure form, as well as in a crude extract, was evaluated across Caco-2 cell monolayers and across excised porcine intestinal, sublingual and buccal tissues. Results The purities of the isolated cathine and norephedrine were in excess of 90%, thereby proving that HPCCC can be applied for efficient separation of these alkaloids from extracts of Khat. The apparent permeability (P app ) coefficients for the Khat alkaloids in their pure form were all above 1.0×10 −6 cm/s, indicating that the transport of the three alkaloids across the selected biological membranes is comparable to that of the highly permeable reference compound, caffeine. Although readily transported across the various membranes, the alkaloids were transported to a lesser extent when present in a leaf extract, suggesting that other phytochemicals present in the extract influence their permeation. Conclusions These results provide evidence that chewing of Khat contributes to the buccal and sublingual absorption of the psychoactive alkaloids in the bloodstream directly across the oral mucosal membranes. In addition, it confirms that these metabolites will be readily absorbed from the gastrointestinal tract when swallowed.

Contamination of maize by fumonisin mycotoxins is of major concern to consumer health. This study was designed to improve our understanding of the binding of fumonisin B (FB) to maize and its major macrocomponents, during the preparation of porridge. None of the processing treatments (pH 4, 7 and 10) or temperatures (30 and 90 °C) used resulted in a significant decrease in the recovery of FB1 from spiked starch, zein or maize meal. These results confirm the stability of FB1 in maize porridge, and the absence of modified forms of FB. The bioaccessibility of fumonisins in the gastrointestinal tract, following consumption of contaminated stiff maize porridge, was determined using the TNO Gastrointestinal Model (tiny-TIM). The porridge was digested in the tiny-TIM using parameters simulating human digestion. The concentrations of fumonisins were determined in the dialysates at fixed time intervals using ultra performance liquid chromatography-quadrupole Time-of-Flight high definition mass spectrometry (UPLC-QToF-HDMS). The results proved that fumonisins B1, B2 and B3 are released rapidly from stiff porridge prepared from contaminated maize meal. This confirms that the low bioavailability of fumonisins cannot be attributed to strong association of the toxin with the intestinal content. This study presents new insights into the stability of fumonisins in maize porridge and the kinetics of fumonisin-release from porridge through simulated human digestion, suggesting that consumers using maize as a staple may be at risk.

Indigenous Salvia species from southern Africa are popular traditional medicines for the treatment of a variety of conditions. They produce fragrant volatiles that can be isolated as essential oils. Some of these volatile organic compounds may play a role in the biological activities of the extracts. Three indigenous Salvia species, Salvia africana-lutea, S. lanceolata and S. chamelaeagnea, were selected for this study as they are commonly used in traditional medicine in South Africa, and the essential oils from these species have potential for commercialisation. Although some studies have described the essential oil compositions and some biological activities, only single composite samples were used. The aim of this study was to investigate the intra- and interspecies variation of the essential oils, sampled over a wide geographical area and using a representative sample size, to encourage commercialisation of the essential oil. Essential oils were isolated from individual plants using conventional hydrodistillation of the aerial parts, harvested from several localities. Gas chromatography coupled simultaneously to mass spectrometry/flame ionisation detection (GC-MS/FID) was used to identify and quantify the volatile constituents. The essential oils of S. africana-lutea consisted mainly of terpinene-4-ol + β-caryophyllene (1.4 – 29.0%), T-cadinol (1.2 – 20.0%), α-eudesmol (trace – 23.0%) and β-eudesmol (trace – 26.0%), those of S. lanceolata comprised mainly terpinene-4-ol + β-caryophyllene (4.3 – 31.0%), α-humulene (2.3 – 15.0%), bicyclogermacrene (trace – 37.0%) and spathulenol (trace – 25.0%), while the essential oils of S. chamelaeagnea were characterised by δ-3-carene (trace – 18.0%), limonene (1.6 – 36.0%), viridiflorol (9.8 – 61.0%) and 1,8-cineole (not detected – 11.0%). The compounds identified in the essential oils of the three selected Salvia species have been identified in other Salvia essential oils. To add to the novelty of this study, the superior resolving power of two-dimensional gas chromatography was demonstrated through analysis of selected essential oils. Many additional compounds were identified, and previously co-eluting compounds were clearly separated. Chemometric modelling of the GC-MS data using SIMCA P+ 14 software allowed distinct clustering patterns to be discerned. The unsupervised principal component analysis model revealed separate clusters for the three species, confirming substantial chemical differences between their essential oils. Quantitative, rather than qualitative differences were evident when individual essential oil samples representing the same species, were compared. For each species, two chemically distinct groups were observed and unique marker compounds could be identified. This study has contributed detailed information on the major and minor volatile compounds present in the essential oils of the three Salvia species investigated.

Piper methysticum Forst., commonly referred to as kava, has been used medicinally and recreationally by inhabitants of the South Pacific Islands for centuries. Kavalactones present in roots and aerial parts are regarded as the bioactive compounds responsible for the relaxant effects, and for mitigating anxiety and stress-related conditions. The development of pharmaceutical products containing root extracts led to a boom in kava sales in Europe in 1998. However, reported cases of toxicity led to the subsequent banning of kava products in several countries. This study was initiated to develop rapid, robust and alternative spectroscopic methods for quality control that can be implemented at the point of export, to authenticate the use of kava roots as legislated by the Vanuatu Amended Kava Act no. 6 (2015). Roots, peeled stems, and stump peelings samples (n = 47) were sourced from Fiji, Hawaii, Samoa, the Solomon Islands and Tonga. The sample extracts were analysed using ultra performance liquid chromatography coupled to a photodiode array detector and mass spectrometer (UHPLC-PDA/MS), while powdered material was analysed using spectroscopic techniques. These included single-point (near-infrared (NIR) and mid-infrared (MIR) spectroscopy), as well as an imaging (hyperspectral imaging). Principal component analysis of both the raw UPLC-MS and the quantitative UPLC-PDA data revealed chemical differences between the root and non-root samples. Kavain, methysticin and yangonin were identified as the compounds largely responsible for the chemical differences between the plant parts. Discriminant analysis models (OPLS-DA and PLS-DA) were developed for all the techniques, to reliably discriminate kava roots from non-roots. All the discriminant models indicated a good prediction ability (Q2XCum ≥ 60 %) and were successfully used to accurately identify external roots and non-root samples. However, hyperspectral imaging yielded superior results, with a prediction ability above 90 %. This technique can be automated and is capable of continuously scanning multiple samples, making it ideal for quality control.

The African wild olive (Olea europaea subsp. africana) is traditionally used as a hypotensive agent. Herb-drug interactions may result from the concurrent use of herbal medicines and conventional prescription drugs. This aspect was investigated by determining the effect of the extract on the in vitro intestinal epithelial permeation of selected hypotensive drugs using the Caco-2 cell culture model. The phytochemical profiles of leaf extracts of African wild olive from different localities in South Africa were compared, since efficacy is determined by the chemical composition. Extracts were analysed using ultra-performance liquid chromatography. The oleuropein concentration varied considerably from below the detection limit (4.94 µg/mL) to 59.4 mg/g dry weight. Chemometric models constructed from the aligned chromatographic data indicated only quantitative differences between the profiles. The leaf extract was found to increase the permeability of propranolol in the absorptive direction (Papp = 8.93 × 10−6 cm/s) across Caco-2 cell monolayers, but considerably decreased transport in the secretory direction (Papp = 3.68 × 10−6 cm/s). The permeation of diltiazem was enhanced by the extract in both the absorptive (Papp = 7.33 × 10−6 cm/s) as well as in the secretory direction (Papp = 7.16 × 10−6 cm/s), but a decrease in the efflux ratio was observed. The extract therefore caused a net increase in the transport of both drugs in the absorptive direction due to an inhibition effect on their efflux. This suggests a potential increase in the blood levels of these drugs when taken simultaneously with African wild olive leaf extract, indicating potential adverse effects that must be verified in vivo.

A molecularly imprinted polymer (MIP) was prepared using (-)-norephedrine as the template, methacrylic acid as the functional monomer, ethylene glycol dimethacrylate as the cross-linker and chloroform as the porogen. The MIP was used as a selective sorbent in the molecularly imprinted solid-phase extraction (MIP-SPE) of the psychoactive phenylpropylamino alkaloids, norephedrine and its analogs, cathinone and cathine, from Khat (Catha edulis Vahl. Endl.) leaf extracts prior to HPLC-DAD analysis. The MIP was able to selectively extract the alkaloids from the aqueous extracts of Khat. Loading, washing and elution of the alkaloids bound to the MIP were evaluated under different conditions. The clean baseline of the Khat extract obtained after MIP-SPE confirmed that a selective and efficient sample clean-up was achieved. Good recoveries (90.0-107%) and precision (RSDs 2.3-3.2%) were obtained in the validation of the MIP-SPE-HPLC procedure. The content of the three alkaloids in Khat samples determined after treatment with MIP-SPE and a commercial Isolute C18 (EC) SPE cartridge were in good agreement. These findings indicate that MIP-SPE is a reliable method that can be used for sample pre-treatment for the determination of Khat alkaloids in plant extracts or similar matrices and could be applicable in pharmaceutical, forensic and biomedical laboratories. Copyright © 2015 John Wiley & Sons, Ltd.

The alkaloid compositions of 99 green coffee (Coffea arabica L.) bean samples comprising eight varieties (Harar, Jimma, Kaffa, Wollega, Sidama, Yirgachefe, Benishangul and Finoteselam) from the major production regions of Ethiopia were investigated. High performance liquid chromatography was applied for the simultaneous determination of four coffee alkaloids in the aqueous extracts of the beans. The limits of detection for the method were established as 13 mg kg−1 for trigonelline, 7 mg kg−1 for theobromine, 8.5 mg kg−1 for caffeine and 4 mg kg−1 for theophylline in the dry coffee beans. Theophylline was not detected in any of the samples. The determined concentrations (% w/w dry coffee beans) ranged from 0.98 to 1.32 % for trigonelline, 0.0048 to 0.0094 % for theobromine and 0.87 to 1.38 % for caffeine. The concentrations of the alkaloids varied significantly, depending on the geographical origin of the beans. Theobromine was not detected in coffee beans from the East (Harar coffees), and its absence in samples can be used to ascertain whether the coffee originates from this region. Coffee beans from the Northwest were characterized by higher concentrations of caffeine. Application of linear discriminant analysis provided 75 % correct classification of samples into the respective production regions, with a 74 % prediction success rate. The moderate classification efficiency obtained when using alkaloid data demonstrates the potential of using this class of compounds in discriminant models for determination of the geographical origin of green coffee beans from Ethiopia.

Gingerols are the major pungent compounds present in the rhizomes of ginger (Zingiber officinale Roscoe) and are renowned for their contribution to human health and nutrition. Medicinal properties of ginger, including the alleviation of nausea, arthritis and pain, have been associated with the gingerols. Gingerol analogues are thermally labile and easily undergo dehydration reactions to form the corresponding shogaols, which impart the characteristic pungent taste to dried ginger. Both gingerols and shogaols exhibit a host of biological activities, ranging from anticancer, anti-oxidant, antimicrobial, anti-inflammatory and anti-allergic to various central nervous system activities. Shogaols are important biomarkers used for the quality control of many ginger-containing products, due to their diverse biological activities. In this review, a large body of available knowledge on the biosynthesis, chemical synthesis and pharmacological activities, as well as on the structure-activity relationships of various gingerols and shogaols, have been collated, coherently summarised and discussed. The manuscript highlights convincing evidence indicating that these phenolic compounds could serve as important lead molecules for the development of therapeutic agents to treat various life-threatening human diseases, particularly cancer. Inclusion of ginger or ginger extracts in nutraceutical formulations could provide valuable protection against diabetes, cardiac and hepatic disorders.

Butein (2′,3,4,4′-tetrahydroxychalcone), a simple chalcone derivative, occurs in many unrelated genera including Butea Dahlia, Coreopsis and Searsia. It is a reputed food additive and a common ingredient of botanicals used in herbal medicine formulations, particularly in Asian countries. Although a simple polyphenol, this molecule exhibits a range of pharmacological properties, most notably acting as a potent protein tyrosine kinase inhibitor and as an antineoplastic agent. Researchers have convincingly demonstrated that butein inhibits the epidermal growth factor receptor in HepG2 cells and the tyrosine-specific protein kinase activities of the epidermal growth factor receptor. In addition, it also exhibits promising anti-inflammatory, antidiabetic, antinephritic, antithrombin, anti-angiogenic and hepatoprotective activities in various animal models. Although this molecule is endowed with an impressive list of biological properties, which have acted as scientific support for its commercialization, there are no review articles that coherently discuss various aspects of this chalcanoid. This review aims to explore the pharmacological relevance of butein, together with its structure–activity relationships and mechanisms of action. In addition, the occurrence, chemical synthesis and biosynthesis of butein are discussed.

The well-known anti-infective properties of propolis are determined by its chemical composition, which in turn is influenced by geographical factors and reflects the botanical diversity in the vicinity of the beehive. Although there are several reports on the anti-infective properties of crude propolis, few are aimed at identifying specific compound(s) responsible for the observed activities. Using South African propolis as an example, the application of high performance thin layer chromatography-bioautography in tandem with mass spectrometry was investigated for the rapid identification of antimicrobial and anti-quorum sensing (anti-QS) compounds. Pinocembrin was found to be responsible for the observed antifungal activity of the propolis against Candida albicans. Three compounds were found to be active against all of the evaluated Gram-positive and Gram-negative bacteria. The identity of the first was confirmed as pinobanksin, one remains unidentified, while the third corresponds to either pinobanksin 3- O -pentanoate or 2-methylbutyrate. The identification of caffeic acid as the anti-QS component was confirmed quantitatively using the violacein inhibitory assay.

Embelin(2,5-dihydroxy-3-undecyl-p-benzoquinone)is known for its potent anthelmintic activity, but also for wound-healing, antidiabetic, anticonvulsant, antitumour, anti-inflammatory, analgesic, hepatoprotective, antioxidant, antibacterial andantispermatogenic activities. A high performance countercurrent chromatography method was developed for the purification of embelin from an extract of the seeds of Embelia schimperi fruit. The optimised solvent system (n-hexane:ethylacetate:ethanol:water (7:3:7:3)) resulted in the isolation of 13.9 mg of embelin, directly from 100 mg of crude extract, in a single step within a short retention time (40 min). Although the compound appeared to be completely pure when analysed by ultra performance liquid chromatography (UPLC) with photo diode array detection (PDA), the purity was established as approximately 90% by UPLC-mass spectrometry (MS). Furthermore, we report the fatty acid composition of the seeds of Embelia schimperi fruit. Nine fatty acids were quantified from the fruit seed extract by gas chromatography-mass spectrometry, with linoleic (46.4%), palmitic (21.5%), and oleic (19.6%) acids making up the largest proportions.

Infection of mango panicles and young leaf shoots by Fusarium results in significantly lower fruit yields. Some mango cultivars are known to be tolerant to blossom malformation, while others are highly susceptible. Previous researchers linked the ability of certain cultivars to confine Fusarium infection to the production of high levels of mangiferin. This study further elucidates the role of phenolic metabolites contributing to tolerance. Phenolic profiles of mature leaf extracts from cultivars with varying degrees of resistance to Fusarium infection were obtained by ultrahigh performance liquid chromatography–time-of-flight-mass detection. Chemometric models constructed from the data revealed a clear distinction between the profiles of tolerant and susceptible cultivars and permitted the identification of biomarkers associated with susceptibility to infection. The presence of elevated levels of mangiferin, maclurin and maclurin O-galloyl-glucoside in the leaves were associated with a higher degree of tolerance. In contrast, high levels of iriflophenone glucosides were linked to increased susceptibility of the cultivars to infection. These models can serve as an important tool to investigate appropriate cultivars, prior to their introduction to areas prone to the disease.

Ethnopharmacological relevance The use of Lawsonia inermis L. (henna) for medicinal and cosmetic purposes is inextricably linked to ancient and modern cultures of North Africa and Asia. Literature and artwork indicates that Lawsonia inermis played an important holistic role in the daily lives of some ancient cultures, providing psychological and medicinal benefits, as well as being used for personal adornment. Although henna was historically applied to the hands and feet to protect against fungal pathogens and to hair to combat lice and dandruff, other traditional uses include the treatment of liver and digestive disorders, reduction of tissue loss in leprosy, diabetic foot disorders and ulcers. Phytochemistry: Almost 70 phenolic compounds have been isolated from various parts of the plant. Naphthaquinones, which include the dyeing principle lawsone, have been linked to many of the pharmacological activities. The terpene, β-ionone is largely responsible for the pungent odour of the essential oil isolated from the flowers. In addition to other volatile terpenes, some non-volatile terpenoids, a single sterol, two alkaloids and two dioxin derivatives have also been isolated from the plant. Bioactivity: Henna is a pharmacologically important plant with significant in vitro and in vivo biological activities. Although a myriad of pharmacological activities have been documented, the antioxidant and antimicrobial activities are the most thoroughly investigated. Some incidents of adverse reactions following application to the skin have been reported, but these are mainly confined to cases involving individuals with glucose-6-phosphate dehydrogenase deficiency and reactions to adulterants added to henna products. Conclusions: Adulteration of henna is very common and may have resulted in unwarranted scientific findings. Phytochemical profiling studies of the plant, which are crucial for the establishment of proper quality control protocols, are lacking and hamper the development of medicinal products. Although many in vitro studies have been conducted to evaluate the pharmacological activities and many in vivo studies have focussed on the toxicity of extracts, more in vivo studies to validate pharmacological activities are needed. The roles of specific compounds and their synergies have not been comprehensively investigated.

Phytosterols of manketti (Schinziophyton rautanenii) nut oil extracted by Soxhlet, mechanical shaking using hexane, screw press and supercritical carbon dioxide, were analyzed by gas chromatography with flame ionization detection and identified by gas chromatography–mass spectrometry. The presence of several phytosterols (campesterol, stigmasterol, β-sitosterol, ∆5-avenasterol, 22-dihydrospinasterol and ∆7-avenasterol) previously reported in manketti oil, was confirmed. In addition, another fourteen phytosterols (lanosterol, ∆5,23-stigmastadienol, ∆7-campesterol, clerosterol, obtusifoliol, ∆5,24(25)-stigmastadienol, α-amyrin, gramisterol, cycloeucalenol, cycloartenol, stigmasta-8,24-dienol-3-β-ol, 28-methylobtusifoliol, 24-methylenecycloartenol and citrostadienol) were identified. The phytosterols, β-sitosterol, ∆5-avenasterol and campesterol, had the highest concentrations in oils extracted by all the methods, whereas stigmasterol and cycloartenol were abundant in oils extracted by mechanical shaking and supercritical carbon dioxide. Total phytosterols and the quantities of individual phytosterols differed significantly (p ≤ 0.05) in oils from the four extraction methods. Mechanical shaking extracted the highest levels of total sterols (22,100 mg/100 g oil), followed by supercritical fluid extraction (9,550 mg/100 g oil). Screw press and Soxhlet extracted oils contained the lowest levels of total sterols, 3,810 mg/100 g oil and 3,350 mg/100 g oil, respectively.

The biological properties of verbascoside are numerous and include antimicrobial activities. As part of a drive to identify alternatives to synthetic fungicides for crop protection, this study was aimed at investigating methods of preparing verbascoside-rich plant extracts from viable sources of the compound and monitoring its stability under various conditions. Leaf extracts of Lippia javanica Spreng. and Lantana camara Linne (Verbenaceae) were partially purified using column chromatography and high speed centrifugal countercurrent chromatography, the latter yielding fractions with higher purity (71%) than those from a single column chromatographic separation (38–44% pure). Verbascoside remained stable upon heating, but was completely decomposed after four hours exposure of the extract to sunlight. Compared to the other storage conditions, the compound was best preserved in a dry form in the dark. Analysis by high performance liquid chromatography revealed that the verbascoside content of plant parts of L. camara from natural populations was highly variable, both within and between populations. However, several specimens produced high levels of the compound (Hazyview: Plant 3 (83.0 mg/g dry weight), Magoebaskloof 2: Plant 5 (64.8 mg/g) and White River: Plant 2 (64.0 mg/g)), suggesting that L. camara is an excellent source of verbascoside. Extracts of the plant displayed effective in vivo inhibition of Penicillium digitatum on oranges.

Henna (Lawsonia inermis) is applied to stain keratin, present in hair, skin and fingernails, a red-orange or rust colour. Producers of temporary tattoos mix the aromatic amine compound, para-phenylenediamine (PPD) into natural henna to create 'black henna' that rapidly stains the skin black. However, PPD may cause severe delayed hypersensitivity reactions following skin contact. This study proposes a rapid direct-analysis method to detect and identify PPD using an atmospheric solids analysis probe (ASAP) coupled to a Q-ToF mass spectrometer (MS). Since laborious, multistep methods of analysis to determine PPD are undesirable, due to the instability of the compound in solution, a screening method involving no sample preparation steps was developed. Experiments were carried out to optimise the corona current, sample cone voltage, source temperature, and desolvation gas temperature to determine ideal ASAP-Q-ToF-MS analysing conditions. Eleven of the 109 henna samples, originating from various countries, tested positive for PPD when henna products were screened using ASAP-MS, without any form of sample preparation other than grinding. Ultra-performance liquid chromatography electrospray ionisation-mass spectrometry (UPLC-Q-ToF-MS) was subsequently used to confirm the results from ASAP and to determine the concentrations of PPD in henna products. The allergen was detected in the same eleven samples, with concentrations ranging from 0.05-4.21% (w/w). It can be concluded that the sensitivity of the ASAP-MS technique is sufficient (limit of detection=0.025% w/w) to allow screening of henna samples for the presence of PPD. This relatively new technique can be applied to commercial products without extraction, sample treatment or chromatographic separation.

The development of natural crop protective products as alternatives to synthetic fungicides is currently in the spotlight. In vitro experiments are valuable precursors to more costly in vivo trials, allowing the identification of effective essential oils and establishing the concentrations required for inhibition of a specific, or spectrum of decay pathogens. In this study, the antifungal properties of eighteen essential oils were evaluated in vitro by addition to the fungal growth medium of five pathogens ( Lasiodiplodia theobromae , Colletotrichum gloeosporioides , Alternaria citrii , Botrytis cinerea and Penicillium digitatum ) isolated from mango, avocado, citrus, grapes and cactus pear. The inhibitory properties of some of the major compounds of the oils, identified and quantified by gas chromatography–mass spectroscopy and gas chromatography–flame ionization detection were also determined. Most of the oils were selected on the basis of commercial availability and for containing a predominant compound. Visual inspection of fungal growth was done and the lowest concentration where fungal growth was completely inhibited on all replicates was recorded. Thyme oil proved to be the most effective inhibitor, totally inhibiting all of the pathogens tested at concentrations of 1000 μl/l and lower, with the exception of a resistant Penicillium strain. Cinnamon oil, rich in eugenol (81.2%), demonstrated good fungicide potential, while the carvone-rich oils displayed promising activity against the citrus pathogens. Oils characterized by high concentrations of S-carvone were less effective than those containing the R-enantiomer. Essential oil of Lippia citriodora was active against all of the pathogens, excluding L. theobromae from avocado. These essential oils, applied alone or in combination, are good candidates for further in vivo testing and for investigations concerning their modes of action.

Lippia scaberrima Sond. (Verbenaceae) is an aromatic shrub, indigenous to South Africa. A camphorous essential oil was obtained from bulk aerial parts of L. scaberrima by steam distillation. Gas chromatography with flame ionization detection revealed that d-limonene, 1,8-cineole and (R)-(-)-carvone were the main oil constituents. Strong in vitro antifungal activity of the essential oil was observed against common postharvest spoilage pathogens of mango, avocado and citrus fruit. R-carvone was found to be largely responsible for this antifungal activity. These results were confirmed by semi-commercial and commercial trials using fruit treated with coatings, amended with L. scaberrima and Mentha spicata (spearmint) essential oil. The control treatments were done using standard commercial fungicide applications, while the experimental treatments all consisted of coating amended with the oil or single terpenoid constituents, in the absence of synthetic fungicides. Effective disease control was obtained using all of the amended treatments. Quality parameters measured indicated that the overall quality of the fruits was maintained. Essential oil amended coatings could eliminate environmentally harmful toxic waste originating from standard packhouse procedures. Alternatively, the use of essential oils alone or in combination with synthetic fungicides, could lengthen the useful lifespan of existing fungicides.

Proliferation of fungal pathogens, such as Mycogone perniciosa , can severely affect the yields of cultivated mushrooms, including that of the button mushroom, Agaricus bisporus. A reduction in the number of fungicidal products approved for commercial application is currently providing new challenges to the mushroom industry . Forty essential oils, seven pure terpenoids and one phenylpropanoid were screened in vitro to determine the abilities of these substances to inhibit the growth of M. perniciosa . The fungal growth medium of both A. bisporus and M. perniciosa was supplemented with each test substance at a concentration of 50 μL/L. Ten essential oils were further investigated at lower concentrations ranging from 5 to 40 μL/L. The main components of these oils were determined by GC–FID and GC–MS. Lemon verbena ( Lippia citriodora ), lemongrass ( Cymbopogon citratus ) and thyme ( Thymus vulgaris ) oils were found to substantially inhibit the growth of the pathogen, while demonstrating lower toxicity towards A. bisporus than any of the other oils tested. A preliminary in vivo trial using M. perniciosa -inoculated casings revealed that the preventative use of lemon verbena or thyme oils was able to control the development of the disease. A commercial trial using these oils, as well as two of their main components (nerol and thymol), at a concentration of 40 μL/L, revealed that none of these treatments were detrimental to the growth of the A. bisporus and an overall yield similar to that following application of a commercial fungicide (Chronos 450 SC) was obtained. These results suggest that essential oils or mixtures of selected pure components of essential oils may in future find application in button mushroom production, either as a substitute for synthetic fungicides or as an additional protective measure.

Mycobiocides are attracting research interest worldwide as possible postharvest pathogen control measures to replace synthetic fungicides. In this study, the application of two essential oils as fungicides was evaluated. Initially, the in vitro antifungal effects of Lippia scaberrima essential oil and three of the major oil components, ( d )-limonene, R-(−)-carvone, and 1,8-cineole, as well as that of S-(+)-carvone, were investigated against Colletotrichum gloeosporioides , Lasiodiplodia theobromae , and an Alternaria isolate. The oil and terpenoids caused significant inhibition of the mycelial growth of all the pathogens when applied at a concentration of 2000 μL L −1 . The most potent volatile component of L . scaberrima essential oil, able to inhibit all the pathogens tested, proved to be R -(−)-carvone. The efficacy of the essential oil (1000 and 2000 μL L −1 ) incorporated into the commercial coating was confirmed on fruit inoculated with two of the pathogens. A simulated export trial was done using Lippia essential oil, in addition to Mentha spicata (spearmint) essential oil, as supplements for fruit coatings. Results indicate that essential oils rich in R-(−)-carvone could be valuable alternatives to synthetic fungicides for the postharvest management of avocado fruit. The combination of essential oils with a commercial coating, acceptable to the organic market, offers additional protection to this vulnerable commodity.

The mango gall fly (Procontarinia) is a growing concern in many countries that import mango trees. Mangoes flush (produce opening buds) twice a year: once in spring, and the main flush, which occurs in early autumn flush and corresponds to the photoperiod necessary for the gall fly to emerge. Gall flies attack only very young mango flush leaves. Three cultivars displaying varying susceptibility were used in this study: Heidi, which typically is heavily infected; Keitt, which displays antixenosis properties; and Sensation, which is resistant to gall fly attack. Headspace profiles of the three cultivars were obtained over two full growing seasons. It was found that flush leaves emit higher levels of volatile organic compounds than mature leaves. Volatile emissions peaked during the main flush, after which they declined as the tree went into the winter dormant period. The levels of α- and β-pinene increased in gall fly-susceptible cultivars as the season progressed, while very low levels of these terpenes were emitted by resistant cultivars. The changes in the profiles of these emissions suggest that α- and β-pinene may be linked to gall fly susceptibility. In addition, chemometric analysis confirmed the correlation between volatile emissions and gall fly susceptibility. Copyright © 2010 John Wiley & Sons, Ltd.

Lippia rehmannii H.Pearson (Verbenaceae) is an aromatic bush, indigenous to the northern parts of South Africa. As far as could be ascertained, the essential oil composition has not been previously reported and forms the subject of this investigation. Aerial parts of the shrub were collected from two localities in Gauteng, South Africa, and the isolated essential oils were analysed by gas chromatography. Citral, a mixture of the E- and Z-isomers, was found to be the main constituent of the oils, while borneol, camphor, neryl acetate, isocaryophyllene, p -cymene, β-caryophyllene and β-caryophyllene oxide were other major compounds present. Oil compositions, within and between the two localities, did not differ significantly. The in vitro antifungal activity of L. rehmannii essential oil was compared to that of Cympopogon citratus (lemongrass) and pure citral, against a number of pre- and postharvest fungal food pathogens. At a concentration of 3000 µL/L, lemongrass oil and pure citral caused complete growth inhibition of all the pathogens tested. Lippia rehmannii , containing less citral than lemongrass oil, was effective at this concentration against the majority of pathogens, but only partially restricted the growth of Lasiodiplodia theobromae and Botrytis cinerea. This finding suggests that citral may be largely responsible for the observed antifungal activities. Essential oil from L. rehmannii appears to be a good candidate for the in vitro control of Fusarium oxysporum and Rhizoctonia solani and application of these oils in the field should be investigated.

A new approach to the control of postharvest pathogens, while maintaining fruit quality, has been implemented by the application of essential oil amended coatings to citrus. This approach eliminates the need for synthetic fungicides, thereby complying with consumer preferences, organic requirements and reducing environmental pollution. In vitro studies indicated that the essential oils and some of the terpenoid components tested were active against Penicillium digitatum. In a series of subsequent semi-commercial and commercial trials, Mentha spicata and Lippia scaberrima essential oils, as well as pure (d)-limonene and R-(−)-carvone were incorporated into a variety of commercial citrus coatings. These amended coatings were applied postharvest to ‘Tomango’ oranges in the absence of the standard fungicide dip. Excellent disease control was achieved with the amended coatings, while measured quality parameters indicated that overall fruit quality was maintained. Moreover, moisture loss was decreased significantly in fruit treated with essential oil enriched coatings. The efficacy of amended coatings as a viable alternative or supplement to existing fruit protection strategies was demonstrated in a commercial trial.

Introduction . Penicillium digitatum is a commercially important postharvest pathogen of citrus that is responsible for significant annual global losses. Strains of the fungus, which exhibit strong resistance to widely used synthetic fungicides, are of major concern to the industry. The aim of the study was to investigate the antifungal activities of polar extracts and compounds from Lippia species, indigenous to South Africa, against a Guazatine® -resistant strain of P. digitatum. Materials and methods . In vitro tests were done by incorporating the compounds and plant extracts into Malt Extract Agar at concentrations ranging from (0.2 to 1.0) mg·mL–1 . An in vivo (curative) assay was conducted using the checkerboard technique on ‘Valencia’ oranges. Results and discussion . A strong correlation between the in vitro and in vivo results was observed. All the compounds and extracts were able to inhibit fungal growth at concentrations above 0.6 mg·mL–1 . Verbascoside was the most active compound, while extracts of Lippia javanica and Lippia rehmannii caused significant inhibition of mycelial growth. The observed activity was largely ascribed to the presence of verbascoside in the plant extracts. Conclusion . Polar extracts of Lippia species have potential as environmentally friendly alternatives for the control of P. digitatum on citrus.

Lippia scaberrima , an aromatic shrub with medicinal properties, is indigenous to southern Africa. Essential oil obtained from aerial plant parts was assessed in vitro and in vivo for antifungal activity against Botryosphaeria parva and Colletotrichum gloeosporioides isolated from mango fruit. Exposure to oil vapour for 2 and 4 days caused inhibition of mycelial growth of both isolates. Three major terpenoids identified in the oil, limonene, R -(−)-carvone, and 1,8-cineole, as well as S -(+)-carvone, were individually evaluated in vitro . The two enantiomers of carvone, along with L. scaberrima oil, were found to have the highest fungistatic activity. Oil and carvone enantiomer fungicidal activities were less pronounced. The use of commercial wax coatings enriched with essential oils led to reduced fungal infection by both pathogens. Results of the in vitro and preliminary in vivo studies suggest that employing L. scaberrima oil to control postharvest pathogens is worthy of further investigation.

Leaves of khat ( Catha edulis ) are masticated to elicit their psycho-stimulating properties, resulting from the presence of the phenylpropylamino alkaloids. The determination of these alkaloids is important in pharmacological, phytochemical, forensic and law enforcement environments. In this study, the use of strong cation exchange-solid phase extraction (SCX-SPE) was investigated as an alternative means of sample purification prior to the determination of cathinone, cathine and norephedrine by reversed phase (C 18 ) high performance liquid chromatography (HPLC). Extraction parameters for SCX, including loading capacity and washing solvents, were established. An existing liquid-liquid extraction (LLE) method was improved in terms of recoveries obtained, by using ethyl acetate as extractant. For pure standards of the khat alkaloids, recoveries ranged from 83 to 97%. Preconcentration, using a Genevac evaporator after the addition of acidified water to the sample, restricted analyte losses when compared to concentration under nitrogen. Although comparable recoveries were obtained when preconcentration was achieved in the presence of acidified water by rotary evaporation, this method is not suitable for large sample numbers. Best recoveries, ranging from 94 to 102%, were obtained by SCX from spiked samples. Although the extraction efficiencies of LLE were lower (87-90%), LLE yielded less complex chromatograms, indicating a purer extract. KEY WORDS : Khat alkaloids, Solid phase extraction, Liquid-liquid extraction, HPLC, Genevac Bull. Chem. Soc. Ethiop. 2015 , 29(3), 331-339. DOI: http://dx.doi.org/10.4314/bcse.v29i3.1

Lippia scaberrima (Verbenaceae) is an aromatic, medicinal shrub which is indigenous to southern Africa. Quantitative and qualitative analyses of the essential oil were conducted using gas chromatog...

Hypoxis species (Hypoxidaceae) are amongst the most widely used medicinal plants in southern Africa. Although the phytochemistry of Hypoxis hemerocallidea has been extensively investigated, little is known regarding the secondary metabolites of the other indigenous species, including Hypoxis colchicifolia and Hypoxis galpinii. Two new phenolic glycosides, 3-hydroxy-4-O-β-d-glucopyranosylbenzaldehyde and 1,5-bis(3,4-dihydroxyphenyl)-1,2-dihydroxy-4-pentyne-2-p-coumaroyl-β-d-glucopyranoside were isolated from corms of H. colchicifolia and H. galpinii, respectively. The norlignan glycosides (hypoxoside, dehydroxy hypoxoside and bis-dehydroxy hypoxoside) were isolated from H. colchicifolia for the first time, using high performance countercurrent chromatography and elucidated by 1D and 2D NMR as well as high resolution MS. In addition, geraniol glycoside and β-sitosterol, previously described in Hypoxidaceae, were isolated from the same species. The presence of hypoxoside in all three species investigated indicated that the interchangeable use of Hypoxis species by communities has some merit. Establishing the biological properties of the new constituents will provide more insight into the medicinal value of the genus.

The leaves of Leucosidea sericea (Rosaceae) are used medicinally by some indigenous South African people as a vermifuge and astringent. No information on the biological activity and phytochemistry of the plant could be found in the literature. Bioassay-guided fractionation was used to identify the active compound. Partial purification of the petroleum ether extract of the leaves and flowers was followed by fractionation using chromatographic methods. This process yielded the Phloroglucinol derivatives, aspidinol and desaspidinol, which were previously reported to be present in a Dryopteris species. Aspidinol was further isolated and structurally elucidated. This is the first report of the presence of these compounds in Leucosidea sericea. The disk diffusion method used in our study, indicated that the plant has antimicrobial activity against Staphylococcus aureus , Bacillus subtilis and Candida albicans .

Myricetin is a common plant-derived flavonoid and is well recognised for its nutraceuticals value. It is one of the key ingredients of various foods and beverages. The compound exhibits a wide range of activities that include strong anti-oxidant, anticancer, antidiabetic and anti-inflammatory activities. It displays several activities that are related to the central nervous system and numerous studies have suggested that the compound may be beneficial to protect against diseases such as Parkinson's and Alzheimer's. The use of myricetin as a preserving agent to extend the shelf life of foods containing oils and fats is attributed to the compound's ability to protect lipids against oxidation. A detailed search of existing literature revealed that there is currently no comprehensive review available on this important molecule. Hence, the present work includes the history, synthesis, pharmaceutical applications and toxicity studies of myricetin. This report also highlights structure-activity relationships and mechanisms of action for various biological activities.

Sceletium tortuosum is an indigenous South African plant that has traditionally been used for its mood-enhancing properties. Recently, products containing S. tortuosum have become increasingly popular and are commonly administered as tablets, capsules, teas, decoctions, or tinctures, while traditionally the dried plant material has been masticated. This study evaluated the in vitro permeability of the four major S. tortuosum alkaloids (i.e., mesembrine, mesembrenone, mesembrenol, and mesembranol) across porcine intestinal, sublingual, and buccal tissues in their pure form and in the form of three different crude plant extracts, namely water, methanol, and an acid-base alkaloid-enriched extract. The permeability of mesembrine across intestinal tissue was higher than that of the highly permeable reference compound caffeine (which served as a positive control for membrane permeability) both in its pure form, as well as in the form of crude extracts. The intestinal permeability of mesembranol was similar to that of caffeine, while those of mesembrenol and mesembrenone were lower than that of caffeine, but much higher than that of the poorly permeable reference compound atenolol (which served as a negative control for membrane permeability). In general, the permeabilities of the alkaloids were lower across the sublingual and the buccal tissues than across the intestinal tissue. However, comparing the transport of the alkaloids with that of the reference compounds, there are indications that transport across the membranes of the oral cavity may contribute considerably to the overall bioavailability of the alkaloids, depending on pre-systemic metabolism, when the plant material is chewed and kept in the mouth for prolonged periods. The results from this study confirmed the ability of the alkaloids of S. tortuosum in purified or crude extract form to permeate across intestinal, buccal, and sublingual mucosal tissues. http://www.thieme.com/books-main/biochemistry/product/3494-planta-medica http://dx.doi.org/10.1055/s-0031-1280367 The authors would like to thank the National Research Foundation (South Africa) for providing financial support for the study.

Lippia scaberrima Sond. is an aromatic shrub of semi-arid, summer-rainfall grassland and occurs in the northern provinces of South Africa. The plant is a common natural coloniser of gold mine tailings and contaminated soils, and is one of a number of similar species that has been tested for use in tailings stabilisation. Mine rehabilitation strategies worldwide involve the use of native and locally tolerant plants. Where such plants have commercially valuable attributes, their use to stabilise contaminated soils and tailings also provides opportunities for local beneficiation and the development of new industries post- mining. The production of aromatic plants is considered a particularly safe option for mine waste since they are generally unpalatable to herbivores and the steam distillation methods used for essential oil isolation yield products free from metal and radioactive contaminants. In the initial part of this study the essential oil of bulk samples of L. scaberrima growing in a natural habitat was found to exhibit strong in vitro antifungal activity against several commercially important fruit post- harvest spoilage pathogens. Gas chromatography with flame ionisation detection (GC-FID) analysis of the oil identified limonene, 1,8-cineole and R-carvone as the main constituents. The antifungal activity of the oil was attributed to the presence of R-carvone. These results were confirmed by several semi-commercial and commercial trials on mango, avocado and citrus fruit. The potential application of L. scaberrima essential oil, rich in R-carvone, as a post-harvest fungicide of fruit, may necessitate the large-scale production of the oil. However, essential oil profiles of Lippia spp. in general have been shown to exhibit high intraspecific variability due to genetic differences. In this investigation the essential oil profiles of L. scaberrima specimens from populations growing in mine- contaminated substrata were compared to those of specimens growing in uncontaminated soils to identify a R-carvone chemotype selected for the harsh growing conditions existing in and around mine tailings. The findings indicate that L. scaberrima does not bioaccumulate metals present in contaminated soils, and essential oils prepared using steam distillation were free of metals and uranium.

A short-term cancer initiation/promotion bioassay was established to screen 10 toxic strains of Fusarium moniliforme for their cancer promoting activity in rats. The assay consisted of a four week 'promoting' treatment, effected by incorporating culture material (5%) of each strain into the diet, commencing one week after an initiation treatment with diethylnitrosamine (DEN, 200 mg/kg). The appearance of gamma-glutamyltranspeptidase-positive (GGT+) foci was used as an indication of promoting activity. Three out of 10 strains of F. moniliforme obtained from corn from a high risk area for esophageal cancer in Transkei, southern Africa, had significant cancer promoting activity. A highly significant correlation was found between toxicity expressed as reduction in body weight gain and cancer promoting activity. This finding suggests that the compounds responsible for the hepatotoxicity and hepatocarcinogenicity of F. moniliforme could be identical.

Lippia javanica and Lippia scaberrima are used as herbal remedies and are commercially traded as health teas in southern Africa under the brands “Mosukujane” and “Musukudu”, respectively. This study evaluates the relationship between the presence of phenolic compounds and the antioxidant activities of infusions prepared from four Lippia species ( L. javanica , L. scaberrima , L. rehmannii and L. wilmsii ) indigenous to South Africa. The antioxidant activities of the infusions, determined by the 2,2-diphenylpycrylhydrazyl (DPPH) method, were also compared to those of popular black, green and herbal tea brands. Of the four indigenous species, infusions of L. javanica and L. wilmsii exhibited the highest antioxidant activities (EC 50 : 358 and 525 µg/ml, respectively) and contained the most phenolic compounds (14.8 and 14.5 mg/ml of dry weight gallic acid equivalent, respectively). Antibacterial activities of methanolic extracts of the four Lippia species were determined against four human pathogens ( Staphylococcus aureus , Enterococcus faecalis , Escherichia coli and Pseudomonas aeruginosa ). The extract of L. javanica was the most active against all the pathogens tested. Those Lippia species ( L. javanica and L. wilmsii ) previously reported to produce higher levels of the pharmacologically active phenylethanoid glycosides verbascoside and isoverbascoside, portrayed stronger antioxidant and antibacterial activities. This study gives credence to the use of infusions of these Lippia species for their general health benefits.

Anecdotal evidence indicates that Sceletium tortuosum has been used as a psychoactive preparation for several centuries. The psychological activity is attributed to the production of mesembrine-type alkaloids (mesembrenol, mesembranol, mesembrenone and mesembrine) by the plant. This investigation was aimed at developing validated reversed phase ultra performance liquid chromatography with photodiode array detector (RP‐UHPLC PDA) and gas chromatography coupled to mass spectrometry (GC–MS) methods for quantitative assessment of mesembrine-type alkaloids in S. tortuosum raw materials and products. Both methods were validated for linearity, repeatability and recovery. Limits of detection (LOD) and limits of quantification (LOQ) for the alkaloids were also established. The LOD range for the RP‐UHPLC PDA analysis of compounds was 0.91–1.21 μg ml − 1 , while the LOQ range was 2.72–3.71 μg ml − 1 . For GC–MS analysis, the LOD and LOQ ranges for the compounds were 11.78–22.76 μg ml − 1 and 35.34–68.29 μg ml − 1 respectively. Regression analyses indicated good linear relationships for the investigated compounds. The regression coefficients ( R 2 ) from RP‐UHPLC PDA analyses ranged from 0.9977 to 0.9991, while those derived from GC–MS analyses were within the range of 0.9979 to 0.9995. Intra- and inter-day precision values for all the alkaloids were below 3.44% for RP‐UHPLC PDA and below 3.15% for GC–MS analysis, indicating excellent repeatability. Percentage recovery ranges for the four alkaloids as determined by RP‐UHPLC PDA and GC–MS analyses ranged from 80.00 to 89.83% and from 80.77 to 90.00%, respectively. Plots of RP‐UHPLC PDA data of selected raw materials and products versus GC–MS data, showed good linear correlation ( R 2 = 0.9820–0.9987). The chromatographic methods were found to be repeatable, precise and appropriate for use in the routine quality control of the psychoactive alkaloids in S. tortuosum raw materials and products.

Lippia javanica (N.L.Burm.) Spreng. is an aromatic, multipurpose medicinal plant from which a number of volatile compounds have been identified, together with toxic triterpenoids and iridoid glycosides. Two additional phenylethanoid glycosides, verbascoside and isoverbascoside, were isolated from L. javanica and characterized. High performance liquid chromatography analyses of polar extracts of three other Lippia species (L. scaberrima, L. rehmannii and L. wilmsii), indigenous to South Africa, revealed the presence of both isomers. When compared to the other indigenous Lippia species, the leaves of L. javanica were found to contain the highest concentrations of both isomers. In addition, the intraspecies variation of the verbascoside/isoverbascoside content of L. javanica, harvested from the same and different localities, was investigated. The concentrations of the two phenylethanoids remained fairly consistent within and between different populations, even when geographically separated. While these compounds are produced by many genera, they may now be added to the list of iridoid glucosides employed as chemotaxonomic markers for Lippia species.

Listeria monocytogenes is a Gram-positive bacterium, able to survive and grow in water, soil, agricultural products, various foods and the foodprocessingenvironment.Theubiquitousnatureoftheorganism,coupledwithitsabilitytocolonisefood-processingsurfacesbyformingbiofilms,causes it to be of a major concern to the industry. Increased foodborne pathogen resistance and negative consumer perceptions regarding the use of synthetic bacteriocides, has resulted in natural antimicrobials being sourced from the plant kingdom. The listerial antibiofilm activities of Syzygium aromaticum (clove),Menthaspicata(spearmint),LippiarehmanniiandCymbopogoncitratus(lemongrass)essentialoilsandtheirmajorcomponentswereevaluated using the crystal violet assay and confocal scanning laser microscopy. Listerial biofilms treated with S. aromaticum or M. spicata essential oils, or the pure compounds nerol and citral, exhibited a similar biofilm biomass to the positive control. However, the essential oils of lemongrass, L. rehmannii, eugenol and R-carvone caused biofilm enhancement, rather than inhibition. L. rehmannii and lemongrass essential oils did not display any antibiofilm properties. Results obtained were confirmed by microscopic observations indicating either a reduction (inhibition) or an increase (enhancement) in biofilm biomass when exposed to the essential oils or pure compounds. The present study revealed that M. spicata and S. aromaticum essential oils as well as pure citral and nerol are good candidates for further development of ecofriendly disinfectants. © 2010 SAAB. Published by Elsevier B.V. All rights reserved.