Chemical profiling, anticonvulsant and anxiolytic effects of the smoke constituents isolated from Leonotis leonurus (L.) R.Br.

ETHNOPHARMACOLOGICAL RELEVANCE: The use of medicinal plants for central nervous system (CNS)-related ailments, such as epilepsy and anxiety, is prevalent in South Africa. Plants from the Lamiaceae family are commonly used for their therapeutic benefits. Leonotis leonurus (L.) R.Br. has been reported in ethnobotanical literature to have anticonvulsant and anxiolytic effects through the inhalation of pyrolysis products obtained by combustion of the aerial parts. AIM AND OBJECTIVES: To explore the chemical profiles and CNS activity of the smoke extract and isolated constituents of L. leonurus in zebrafish larvae, through anticonvulsive and anxiolytic activity assays. MATERIALS AND METHODS: The smoke extract of L. leonurus was obtained through the combustion of the aerial parts of the plant using a custom-built smoke recovery apparatus. The chemical profile of the smoke constituents was determined using Ultra-Performance Liquid Chromatography coupled with Mass Spectrometry (UPLC-MS). Targeted compounds were subjected to preparative High-Performance Liquid Chromatography for separation before structure elucidation using Nuclear Magnetic Resonance (NMR). The maximum tolerated concentrations, as well as the anxiolytic activity of the smoke extract were determined in five days post fertilisation zebrafish larvae. Reverse-thigmotaxis and locomotor activity of larvae in the light/dark transition assay were used to determine anxiolytic activity. Zebrafish larvae at six days post fertilisation (dpf) were subjected to several concentrations of the smoke constituents of L. leonurus. The baseline locomotor activity of the larvae was tracked for 30 min, prior to addition of pentylenetetrazole (PTZ) to induce seizure-like behaviour in the larvae, after which the locomotor activity of the larvae was once again tracked for an additional 30 min. RESULTS: The UPLC-MS profiles of the smoke extract revealed the presence of two main compounds, leoleorin A and leoleorin B, which were targeted and isolated. Upon subjection to NMR spectroscopy for structure elucidation, the compounds were confirmed to be labdane diterpenoids. Both leoleorin A and leoleorin B, and the smoke extract displayed suppression of the PTZ induced seizure-like behaviour in zebrafish larvae. Under light and dark conditions, the smoke extract and compounds displayed potential anxiolytic activity at different concentrations. CONCLUSION: Our results suggest that the smoke constituents of L. leonurus may exert anticonvulsant and anxiolytic effects which align with the traditional indications and the mode of administration.

Characterisation of Sclerocarya birrea (marula) seed oil and investigation of the geographical origin by applying similarity calculations, differential NMR and hierarchical cluster analysis.

INTRODUCTION: The marula fruit is an important indigenous African fruit since various commercial products are produced from the pulp and the seed oil. The increased demand requires methods for authentication, quality control and determination of geographical origin. OBJECTIVE: The study aimed to establish a fast and reliable method for characterisation and authentication of marula seed oil. Furthermore, to identify marker compounds that can distinguish marula seed oils from other commercial oils and indicate regional differences. MATERIALS AND METHODS: Metabolic profiling of 44 commercial marula seed oils was performed using proton nuclear magnetic resonance (1 H NMR). For rapid classification similarity calculations were compared with principal component analysis. Differential NMR was used to determine marker compounds. RESULTS: Marula seed oil was found to be similar to macadamia and olive oils and was distinguished from these oils by the detection of minor components. Marula seed oil is differentiated from the other two oils by the absence of α-linolenic acid, relatively high levels of monoglycerides and diglycerides, and an approximately 1:1 ratio of 1,2- and 1,3-diglycerides. When comparing marula seed oils from various regions using hierarchical cluster analysis, clustering of the marula seed oils from Namibia and Zimbabwe was observed and was related to the quantities of linoleic acid and monoglycerides and diglycerides. Some samples displayed deviations in their composition which might indicate adulteration or contamination during the production process. CONCLUSION: The study demonstrates the potential of NMR as a tool in the quality control of marula seed oil. This technique requires very little sample preparation, circumvents derivatisation of the oil components with fast run-times. In addition, samples with chemical profiles that differ from the general signature profile can easily be identified.

Permeation of Phytochemicals of Selected Psychoactive Medicinal Plants across Excised Sheep Respiratory and Olfactory Epithelial Tissues.

The intranasal route of drug administration offers an opportunity to bypass the blood-brain barrier and deliver compounds directly into the brain. Scientific evidence exists for medicinal plants (e.g., Centella asiatica and Mesembryanthemum tortuosum) to treat central nervous system conditions such as anxiety and depression. The ex vivo permeation of selected phytochemicals (i.e., asiaticoside and mesembrine) has been measured across excised sheep nasal respiratory and olfactory tissue. Permeation studies were conducted on individual phytochemicals and C. asiatica and M. tortuosum crude extracts. Asiaticoside exhibited statistically significantly higher permeation across both tissues when applied alone as compared to the C. asiatica crude extract, while mesembrine permeation was similar when applied alone or as M. tortuosum crude extract. Permeation of all the phytocompounds was similar or slightly higher than that of the drug atenolol across the respiratory tissue. Permeation of all the phytocompounds was similar to or slightly lower than that of atenolol across the olfactory tissue. In general, the permeation was higher across the olfactory epithelial tissue than across the respiratory epithelial tissue and therefore showed potential for direct nose-to-brain delivery of the selected psychoactive phytochemicals.

Commercially Available Viola odorata Oil, Chemical Variability and Antimicrobial Activity.

Viola odorata L. oil is frequently recommended in the aromatherapeutic literature for treating respiratory, urinary, and skin infections; however, antimicrobial evidence is lacking. In addition, in aromatherapy, combinations of essential oils are predominantly utilized with the goal of achieving therapeutic synergy, yet no studies investigating the interaction of essential oil combinations with V. odorata oil exists. This study thus aimed to address these gaps by investigating the antimicrobial activity of three Viola odorata oil samples, sourced from different suppliers, independently and in combination with 20 different commercial essential oils, against micro-organisms involved in respiratory, skin, and urinary tract infections associated with global resistance trends. These pathogens include several of the ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa and Enterobacter spp.) The chemical profile of the oils was determined using gas chromatography coupled with mass spectrometry. The minimum inhibitory concentrations (MIC) were determined using the broth micro-dilution method. The interactive profiles for the combinations were assessed by calculating the fractional inhibitory concentration index (ΣFIC). The main compounds varied across the three samples, and included phenethyl alcohol, isopropyl myristate, 2-nonynoic acid, methyl ester, α-terpineol, α-cetone, and benzyl acetate. The V. odorata oil samples displayed overall poor antimicrobial activity when tested alone; however, the antimicrobial activity of the combinations resulted in 55 synergistic interactions where the combination with Santalum austrocaledonicum resulted in the lowest MIC values as low as 0.13 mg/mL. The frequency of the synergistic interactions predominantly occurred against Klebsiella pneumoniae, Pseudomonas aeruginosa, Acinetobacter baumannii, and Enterococcus faecium with noteworthy MIC values ranging from 0.25-1.00 mg/mL. This study also reports on the variability of V. odorata oils sold commercially. While this warrants caution, the antimicrobial benefit in combination provides an impetus for further studies to investigate the therapeutic potential.

Optimizing the Antimicrobial Synergism of Melaleuca alternifolia (Tea Tree) Essential Oil Combinations for Application against Respiratory Related Pathogens.

Antimicrobial research into the use of Melaleuca alternifolia essential oil has demonstrated broad-spectrum activity; however, much of the research published focuses on identifying the potential of this essential oil individually, rather than in combination for an enhanced antimicrobial effect. This study aimed to determine the antimicrobial activity of four essential oil combinations, all inclusive of M. alternifolia, against nine pathogens associated with the respiratory tract. The minimum inhibitory concentration assay was used to determine the antimicrobial activity of four essential oil combinations, M. alternifolia in combination with Cupressus sempervirens, Origanum majorana, Myrtus communis, and Origanum vulgare essential oils. The interactions between essential oil combinations were analyzed using isobolograms and SynergyFinder 2.0 software to visualize the synergistic potential at varied ratios. The antimicrobial activity of the different combinations of essential oils all demonstrated the ability to produce an enhanced antimicrobial effect compared to the essential oils when investigated independently. The findings of this study determined that isobolograms provide a more in-depth analysis of an essential oil combination interaction; however, the value of that interaction should be further quantified using computational modelling such as SynergyFinder. This study further supports the need for more studies where varied ratios of essential oils are investigated for antimicrobial potential.

The African cherry: A review of the botany, traditional uses, phytochemistry, and biological activities of Prunus africana (Hook.f.) Kalkman.

ETHNOPHARMACOLOGICAL RELEVANCE: Prunus africana (Hook.f.) Kalkman (Rosaceae), commonly known as "Pygeum" or "African cherry", occurs in mainland montane forests scattered across sub-Saharan Africa, Madagascar, and some surrounding islands. Traditionally, decoctions of the stem-bark are taken orally for the treatment of a wide variety of conditions, such as benign prostatic hyperplasia (BPH), stomach ache, chest pain, malaria, heart conditions, and gonorrhoea, as well as urinary and kidney diseases. The timber is used to make axe handles and for other household needs. The dense wood is also sawn for timber. AIM: The fragmented information available on the ethnobotany, phytochemistry, and biological activities of the medicinally important P. africana was collated, organised, and analysed in this review, to highlight knowledge voids that can be addressed through future research. MATERIALS AND METHODS: A bibliometric analysis of research output on P. africana was conducted on literature retrieved, using the Scopus® database. The trend in the publications over time was assessed and a network analysis of collaborations between countries and authors was carried out. Furthermore, a detailed review of the literature over the period 1971 to 2021, acquired through Scopus, ScienceDirect, SciFinder, Pubmed, Scirp, DOAJ and Google Scholar, was conducted. All relevant abstracts, full-text articles and various book chapters on the botanical and ethnopharmacological aspects of P. africana, written in English and German, were consulted. RESULTS: A total of 455 documents published from 1971 to 2021, were retrieved using the Scopus search. Analysis of the data showed that the majority of these documents were original research articles, followed by reviews and lastly a miscellaneous group comprising conference papers, book chapters, short surveys, editorials and letters. Data were analysed for annual output and areas of intense research focus, and countries with high research output, productive institutions and authors, and collaborative networks were identified. Prunus africana is reported to exhibit anti-inflammatory, analgesic, antimicrobial, anti-oxidant, antiviral, antimutagenic, anti-asthmatic, anti-androgenic, antiproliferative and apoptotic activities amongst others. Phytosterols and other secondary metabolites such as phenols, triterpenes, fatty acids, and linear alcohols have been the focus of phytochemical investigations. The biological activity has largely been ascribed to the phytosterols (mainly 3-ß-sitosterol, 3-ß-sitostenone, and 3-ß-sitosterol-glucoside), which inhibit the production of prostaglandins in the prostate, thereby suppressing the inflammatory symptoms associated with BPH and chronic prostatitis. CONCLUSIONS: Many of the ethnobotanical assertions for the biological activity of P. africana have been confirmed through in vitro and in vivo studies. However, a disparity exists between the biological activity of the whole extract and that of single compounds isolated from the extract, which were reported to be less effective. This finding suggests that a different approach to biological activity studies should be encouraged that takes all secondary metabolites present into consideration. A robust technique, such as multivariate biochemometric data analysis, which allows for a holistic intervention to study the biological activity of a species is suggested. Furthermore, there is a need to develop rapid and efficient quality control methods for both raw materials and products to replace the time-consuming and laborious methods currently in use.

Flavonoid glycosides and ellagic acid cognates from defatted African mango (Irvingia gabonensis) seed kernel.

Seventeen compounds of diverse classes including four flavonoid glycosides, five ellagic acid derivatives, and eight other metabolites were isolated from the methanolic extract of the defatted seed kernel of Irvingia gabonensis. Among the isolates, quercetin 3-O-methyl-4'-[α-L-rhamnopyranosyl-(1→3)]-O-α-L-rhamnopyranoside (1) and 3,3'-di-O-methyl-4'-O-α-L-rhamnopyranosylellagic acid 4-sulfate ester (5) were found to be previously undescribed. Structure elucidation was mainly achieved by the interpretation of 1D and 2D NMR and HRESIMS spectral data. Though compound 6 was previously reported, its 13C NMR data is being reported herein for the first time. To the best of our literature search knowledge, this is the first phytochemical report on I. gabonensis seed kernels.

Bioautography-guided HPTLC-MS as a rapid hyphenated technique for the identification of antimicrobial compounds from selected South African Combretaceae species.

INTRODUCTION: Many species within Combretaceae are traditionally used for the treatment of bacterial infections. The similarity in chemistry and antimicrobial activities within the family pose a challenge in selecting suitable species for herbal drug development. OBJECTIVE: This study aimed at rapidly identifying antimicrobial compounds using bioautography-guided high-performance thin-layer chromatography coupled with mass spectrometry (HPTLC-MS). METHODS: Hierarchical cluster analysis of ultra-performance liquid chromatography-mass spectrometry data from the methanol extracts of 77 samples, representing four genera within Combretaceae, was carried out. Based on groupings on the dendrogram, 15 samples were selected for bioautography analysis against four pathogens (Staphylococcus aureus, Bacillus cereus, Escherichia coli and Salmonella typhimurium). Active compounds were identified using HPTLC-MS analysis of bands corresponding to the inhibition zones. RESULTS: Bioautography revealed 15 inhibition zones against the four pathogens, with the most prominent present for Combretum imberbe. Analysis of the active bands, using HPTLC-MS indicated that flavonoids, triterpenoids and combretastatin B5 contributed to the antibacterial activity. The compounds corresponding to molecular ions m/z 471 (Combretum imberbe) and 499 (Combretum elaeagnoides) inhibited all four pathogens, and were identified as imberbic acid and jessic acid, respectively. Chemotaxonomic analysis indicated that arjunic acid, ursolic acid and an unidentified triterpenoid (m/z 471) were ubiquitous in the Combretaceae species and could be responsible for their antibacterial activities. CONCLUSION: Application of HPTLC-MS enabled the rapid screening of extracts to identify active compounds within taxonomically related species. This approach allows for greater efficiency in the natural product research workflow to identify bioactive compounds in crude extracts.

Antibacterial Screening, Biochemometric and Bioautographic Evaluation of the Non-Volatile Bioactive Components of Three Indigenous South African Salvia Species.

Salvia africana-lutea L., S. lanceolata L., and S. chamelaeagnea L. are used in South Africa as traditional medicines to treat infections. This paper describes an in-depth investigation into their antibacterial activities to identify bioactive compounds. Methanol extracts from 81 samples were screened against seven bacterial pathogens, using the microdilution assay. Biochemometric models were constructed using data derived from minimum inhibitory concentration (MIC) and ultra-performance liquid chromatography-mass spectrometry data. Active molecules in selected extracts were tentatively identified using high-performance thin layer chromatography (HPTLC), combined with bioautography, and finally, by analysis of active zone eluates by mass spectrometry (MS) via a dedicated interface. Salvia chamelaeagnea displayed notable activity towards all seven pathogens, and the activity, reflected by MICs, was superior to that of the other two species, as confirmed through ANOVA. Biochemometric models highlighted potentially bioactive compounds, including rosmanol methyl ether, epiisorosmanol methyl ether and carnosic acid. Bioautography assays revealed inhibition zones against A. baumannii, an increasingly multidrug-resistant pathogen. Mass spectral data of the eluted zones correlated to those revealed through biochemometric analysis. The study demonstrates the application of a biochemometric approach, bioautography, and direct MS analysis as useful tools for the rapid identification of bioactive constituents in plant extracts.

Investigating the Antituberculosis Activity of Selected Commercial Essential Oils and Identification of Active Constituents Using a Biochemometrics Approach and In Silico Modeling.

Tuberculosis (TB) is a disease caused by Mycobacterium tuberculosis which has become prevalent due to the emergence of resistant M. tuberculosis strains. The use of essential oils (EOs) as potential anti-infective agents to treat microbial infections, including TB, offers promise due to their long historical use and low adverse effects. The current study aimed to investigate the in vitro anti-TB activity of 85 commercial EOs, and identify compounds responsible for the activity, using a biochemometrics approach. A microdilution assay was used to determine the antimycobacterial activity of the EOs towards some non-pathogenic Mycobacterium strains. In parallel, an Alamar blue assay was used to investigate antimycobacterial activity towards the pathogenic M. tuberculosis strain. Chemical profiling of the EOs was performed using gas chromatography-mass spectrometry (GC-MS) analysis. Biochemometrics filtered out putative biomarkers using orthogonal projections to latent structures discriminant analysis (OPLS-DA). In silico modeling was performed to identify potential therapeutic targets of the active biomarkers. Broad-spectrum antimycobacterial activity was observed for Cinnamomum zeylanicum (bark) (MICs = 1.00, 0.50, 0.25 and 0.008 mg/mL) and Levisticum officinale (MICs = 0.50, 0.5, 0.5 and 0.004 mg/mL) towards M. smegmatis, M. fortuitum, M. gordonae and M. tuberculosis, respectively. Biochemometrics predicted cinnamaldehyde, thymol and eugenol as putative biomarkers. Molecular docking demonstrated that cinnamaldehyde could serve as a scaffold for developing a novel class of antimicrobial compounds by targeting FtsZ and PknB from M. tuberculosis.

Optimization of Antioxidant Synergy in a Polyherbal Combination by Experimental Design.

Culinary herbs and spices are known to be good sources of natural antioxidants. Although the antioxidant effects of individual culinary herbs and spices are widely reported, little is known about their effects when used in combination. The current study was therefore undertaken to compare the antioxidant effects of crude extracts and essential oils of some common culinary herbs and spices in various combinations. The antioxidant interactions of 1:1 combinations of the most active individual extracts and essential oils were investigated as well as the optimization of various ratios using the design of experiments (DoE) approach. The 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2-azinobis (3-ethylbenzothiazoline-6-sulphonic acid) (ABTS), and ferric reducing antioxidant power (FRAP) assays were used to determine the antioxidant activity, and MODDE 9.1® software (Umetrics AB, Umea, Sweden) was used to determine the DoE. The results revealed synergism for the following combinations: Mentha piperita with Thymus vulgaris methanol extract (ΣFIC = 0.32 and ΣFIC = 0.15 using the DPPH and FRAP assays, respectively); Rosmarinus officinalis with Syzygium aromaticum methanol extract (ΣFIC = 0.47 using the FRAP assay); T. vulgaris with Zingiber officinalis methanol extracts (ΣFIC = 0.19 using the ABTS assay); and R. officinalis with Z. officinalis dichloromethane extract (ΣFIC = 0.22 using the ABTS assay). The DoE produced a statistically significant (R2 = 0.905 and Q2 = 0.710) model that was able to predict extract combinations with high antioxidant activities, as validated experimentally. The antioxidant activities of the crude extracts from a selection of culinary herbs and spices were improved when in combination, hence creating an innovative opportunity for the future development of supplements for optimum health.

Propolis: chemical diversity and challenges in quality control.

Propolis is a resinous natural product produced by honeybees using beeswax and plant exudates. The chemical composition of propolis is highly complex, and varies with region and season. This inherent chemical variability presents several challenges to its standardisation and quality control. The present review was aimed at highlighting marker compounds for different types of propolis, produced by the species Apis mellifera, from different geographical origins and that display different biological activities, and to discuss strategies for quality control. Over 800 compounds have been reported in the different propolises such as temperate, tropical, birch, Mediterranean, and Pacific propolis; these mainly include alcohols, acids and their esters, benzofuranes, benzopyranes, chalcones, flavonoids and their esters, glycosides (flavonoid and diterpene), glycerol and its esters, lignans, phenylpropanoids, steroids, terpenes and terpenoids. Among these, flavonoids (> 140), terpenes and terpenoids (> 160) were major components. A broad range of biological activities, such as anti-oxidant, antimicrobial, anti-inflammatory, immunomodulatory, and anticancer activities, have been ascribed to propolis constituents, as well as the potential of these compounds to be biomarkers. Several analytical techniques, including non-separation and separation methods have been described in the literature for the quality control assessment of propolis. Mass spectrometry coupled with separation methods, followed by chemometric analysis of the data, was found to be a valuable tool for the profiling and classification of propolis samples, including (bio)marker identification. Due to the rampant chemotypic variability, a multiple-marker assessment strategy considering geographical and biological activity marker(s) with chemometric analysis may be a promising approach for propolis quality assessment. Supplementary Information: The online version contains supplementary material available at 10.1007/s11101-022-09816-1.

Mesembryanthemum tortuosum L. alkaloids modify anxiety-like behaviour in a zebrafish model.

ETHNOPHARMACOLOGICAL RELEVANCE: Mesembryanthemum tortuosum L. (previously known as Sceletium tortuosum (L.) N.E. Br.) is indigenous to South Africa and traditionally used to alleviate anxiety, stress and depression. Mesembrine and its alkaloid analogues such as mesembrenone, mesembrenol and mesembranol have been identified as the key compounds responsible for the reported effects on the central nervous system. AIM OF THE STUDY: To investigate M. tortuosum alkaloids for possible anxiolytic-like effects in the 5-dpf in vivo zebrafish model by assessing thigmotaxis and locomotor activity. MATERIALS AND METHODS: Locomotor activity and reverse-thigmotaxis, recognised anxiety-related behaviours in 5-days post fertilization zebrafish larvae, were analysed under simulated stressful conditions of alternating light-dark challenges. Cheminformatics screening and molecular docking were also performed to rationalize the inhibitory activity of the alkaloids on the serotonin reuptake transporter, the accepted primary mechanism of action of selective serotonin reuptake inhibitors. Mesembrine has been reported to have inhibitory effects on serotonin reuptake, with consequential anti-depressant and anxiolytic effects. RESULTS: All four alkaloids assessed decreased the anxiety-related behaviour of zebrafish larvae exposed to the light-dark challenge. Significant increases in the percentage of time spent in the central arena during the dark phase were also observed when larvae were exposed to the pure alkaloids (mesembrenone, mesembrenol, mesembrine and mesembrenol) compared to the control. However, mesembrenone and mesembranol demonstrated a greater anxiolytic-like effect than the other alkaloids. In addition to favourable pharmacokinetic and physicochemical properties revealed via in silico predictions, high-affinity interactions characterized the binding of the alkaloids with the serotonin transporter. CONCLUSIONS: M. tortuosum alkaloids demonstrated an anxiolytic-like effect in zebrafish larvae providing evidence for its traditional and modern day use as an anxiolytic.

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.

An acute dose-ranging evaluation of the antidepressant properties of Sceletium tortuosum (Zembrin®) versus escitalopram in the Flinders Sensitive Line rat.

ETHNOPHARMACOLOGICAL RELEVANCE: Sceletium tortuosum (L.) N.E.Br. (ST) has been used by the Khoisan people of South Africa as a mood elevator. Its various pharmacological mechanisms of action suggest distinct potential as an antidepressant. Clinical studies in healthy individuals suggest beneficial effects on mood, cognition, and anxiety. AIM OF THE STUDY: To obtain a chromatographic fingerprint of a standardized extract of S. tortuosum (Zembrin®), and to evaluate the acute antidepressant-like properties of Zembrin® versus the reference antidepressant, escitalopram, in the Flinders Sensitive Line (FSL) rat, a genetic rodent model of depression. MATERIALS AND METHODS: The chemical profile of Zembrin® was determined by ultra-performance liquid chromatography-mass spectrometry (UPLC-MS) chromatogram method using alkaloid standards. Twelve saline treated FSL and six Flinders Resistant Line (FRL) control rats were used to confirm face validity of the FSL model using the forced swim test (FST). Thereafter, FSL rats (n = 10) received either 5, 10, 25, 50 or 100 mg/kg of Zembrin®, or 5, 10 or 20 mg/kg escitalopram oxalate (ESC), both via oral gavage, and subjected to the open field test (OFT) and FST. RESULTS: Four main ST alkaloids were identified and quantified in Zembrin® viz. mesembrenone, mesembrenol, mesembrine, and mesembranol (47.9%, 32%, 13.2%, and 6.8% of the total alkaloids, respectively). FSL rats showed significantly decreased swimming and climbing (coping) behaviours, and significantly increased immobility (despair), versus FRL controls. ESC 5 mg/kg and Zembrin® 25 mg/kg and 50 mg/kg showed significant dose-dependent reversal of immobility in FSL rats and variable effects on coping behaviours. Zembrin® 50 mg/kg was the most effective antidepressant dose, showing equivalence to ESC 5. CONCLUSIONS: Zembrin® (25 and 50 mg/kg) and ESC (5 mg/kg) are effective antidepressants after acute treatment in the FST, as assessed in FSL rats. Moreover, Zembrin® 50 mg/kg proved equivalent to ESC 5. Further long-term bio-behavioural studies on the antidepressant properties of Zembrin® are warranted.

Evaluation of the wound healing properties of South African medicinal plants using zebrafish and in vitro bioassays.

ETHNOPHARMACOLOGICAL RELEVANCE: In South Africa, medicinal plants have a history of traditional use, with many species used for treating wounds. The scientific basis of such uses remains largely unexplored. AIM OF THE STUDY: To screen South African plants used ethnomedicinally for wound healing based on their pro-angiogenic and wound healing activity, using transgenic zebrafish larvae and cell culture assays. MATERIALS AND METHODS: South African medicinal plants used for wound healing were chosen according to literature. Dried plant material was extracted using six solvents of varying polarities. Pro-angiogenesis was assessed in vivo by observing morphological changes in sub-intestinal vessels after crude extract treatment of transgenic zebrafish larvae with vasculature-specific expression of a green fluorescent protein. Subsequently, the in vitro anti-inflammatory, fibroblast proliferation and collagen production effects of the plant extracts that were active in the zebrafish angiogenesis assay were investigated using murine macrophage (RAW 264.7) and human fibroblast (MRHF) cell lines. RESULTS: Fourteen plants were extracted using six different solvents to yield 84 extracts and the non-toxic (n=72) were initially screened for pro-angiogenic activity in the zebrafish assay. Of these plant species, extracts of Lobostemon fruticosus, Scabiosa columbaria and Cotyledon orbiculata exhibited good activity in a concentration-dependent manner. All active extracts showed negligible in vitro toxicity using the MTT assay. Lobostemon fruticosus and Scabiosa columbaria extracts showed noteworthy anti-inflammatory activity in RAW 264.7 macrophages. The acetone extract of Lobostemon fruticosus stimulated the most collagen production at 122% above control values using the MRHF cell line, while all four of the selected extracts significantly stimulated cellular proliferation in vitro in the MRHF cell line. CONCLUSIONS: The screening of the selected plant species provided valuable preliminary information validating the use of some of the plants in traditional medicine used for wound healing in South Africa. This study is the first to discover through an evidence-based pharmacology approach the wound healing properties of such plant species using the zebrafish as an in vivo model.

Essential Oil Blends: The Potential of Combined Use for Respiratory Tract Infections.

This study investigated the potential efficacy of 369 commercial essential oil combinations for antimicrobial, anti-toxic and anti-inflammatory activity with the aim of identifying synergy among essential oils commonly used in combination by aromatherapists for respiratory purposes. Essential oil combinations were assessed for their antimicrobial activities using a panel of Gram-positive, Gram-negative, and yeast strains associated with respiratory tract infections. The antimicrobial activity was measured by determining the minimal inhibitory concentration (MIC) of microbial growth. The fractional inhibitory concentration index (ΣFIC) was calculated to determine the antimicrobial interactions between the essential oils in the combination. The toxicity of the essential oil combinations was tested in vitro using the brine shrimp lethality assay, the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay on RAW 264.7 mouse macrophage cells and A549 lung cancer cell lines. In addition, an inflammatory response was evaluated measuring nitric oxide production. The essential oils, when in combination, demonstrated an increased antimicrobial effect, a reduction in toxicity and provided improved anti-inflammatory outcomes. Five distinct combinations [Cupressus sempervirens (cypress) in combination with Melaleuca alternifolia (tea tree), Hyssopus officinalis (hyssop) in combination with Rosmarinus officinalis (rosemary), Origanum marjorana (marjoram) in combination with M. alternifolia, Myrtus communis (myrtle) in combination with M. alternifolia and Origanum vulgare (origanum) in combination with M. alternifolia] were found to be the most promising, demonstrating antimicrobial activity, reduced cytotoxicity and improved anti-inflammatory effects. With the increased prevalence of respiratory tract infections and the growing antimicrobial resistance development associated with antimicrobial treatments, this study provides a promising complementary alternative for the appropriate use of a selection of essential oil combinations for use in the respiratory tract.

The use of chemometric modelling to determine chemical composition-antimicrobial activity relationships of essential oils used in respiratory tract infections.

The antimicrobial effects of essential oils are commonly cited within aromatherapeutic texts for use in respiratory tract infections. These essential oils are inhaled or applied to the skin to treat infections and manage symptoms associated with these conditions. A limited number of these essential oils have been scientifically studied to support these claims, specifically, against respiratory pathogens. This study reports on the minimum inhibitory concentration (MIC) of 49 commercial essential oils recommended for respiratory tract infections, and identifies putative biomarkers responsible for the determined antimicrobial effect following a biochemometric workflow. Essential oils were investigated against nine pathogens. Three essential oils, Amyris balsamifera (amyris), Coriandrum sativum (coriander) and Santalum austrocaledonicum (sandalwood) were identified as having greater activity (MIC value = 0.03-0.13 mg/ml) compared to the other essential oils investigated. The essential oil composition of all 49 oils were determined using Gas Chromatography coupled to Mass Spectroscopy (GC-MS) analysis and the GC-MS data analysed together with the antimicrobial data using chemometric tools. Eugenol was identified as the main biomarker responsible for antimicrobial activity in the majority of the essential oils. The ability of a chemometric model to accurately predict the active and inactive biomarkers of the investigated essential oils against pathogens of the respiratory tract was 80.33%.

Volatile phenolics: A comprehensive review of the anti-infective properties of an important class of essential oil constituents.

Historically, essential oils and their lead molecules have been extensively recognised for their anti-infective properties. In this context, certain volatile phenolics (VPs) have emerged as important antimicrobial compounds with excellent inhibitory activity against pathogenic bacteria and fungi, which further extends to drug-resistant and biofilm-forming micro-organisms. In this review, we aim to collate and discuss a number of published papers on the anti-infective activities of naturally occurring VPs with special emphasis on eugenol, isoeugenol, thymol and carvacrol, using Scopus Web of Science and PubMed databases. The biosynthesis and extraction of these VPs are discussed, while particular attention is given to their broad-spectrum antimicrobial activity and the mechanisms of action. We highlight combinational studies of the VPs with other phytocompounds and with commercially available drugs, which may be a promising and a rewarding future approach to combat antimicrobial resistance. These VPs alone, or concomitantly with other compounds or drugs, have the potential to be incorporated into different formulations for biomedical applications. An in-depth assessment of 2310 articles retrieved from the Scopus database spanning a 35-year period indicated 23.1% increase in global publication growth in VPs anti-infective research, with authors from Italy, Portugal and Austria dominating the research landscape. The dominant areas of investigations are identified as antimicrobial activity, antibacterial mechanism of action, antifungal mechanism of action, extraction methods and phytochemistry, use in the food industry, and for oral and dental anti-infective activity. Specific research areas, which require future attention include; antituberculosis research, nanoparticle formulation of antimicrobial active VP molecules, preclinical and clinical trials. The antimicrobial testing of isoeugenol was found to be the least studied of the VPs and this requires further attention.

Emodin - A natural anthraquinone derivative with diverse pharmacological activities.

Emodin (1,3,8-trihydroxy-6-methyl-anthraquinone) is a natural anthraquinone derivative that is present in numerous globally renowned herbal medicines. It is recognised as a protein tyrosine kinase inhibitor and as an anticancer drug, active against various tumour cells, including lung, breast, liver, and ovarian cancer cells. Recently, its role in combination chemotherapy with various allopathic medicines, to minimize their toxicity and to enhance their efficacy, has been studied. The use of emodin in these therapies is gaining popularity, due to fewer associated side effects compared with standard anticancer drugs. Emodin has a broad therapeutic window, and in addition to its antineoplastic activity, it displays anti-ulcer, anti-inflammatory, hepatoprotective, neuroprotective, antimicrobial, muscle relaxant, immunosuppressive and antifibrotic activities, in both in vitro and in vivo models. Although reviews on the anticancer activity of emodin have been published, none coherently unite all the pharmacological properties of emodin, particularly the anti-oxidant, antimicrobial, antidiabetic, immunosuppressive and hepatoprotective activities of the compound. Hence, in this review, all of the available data regarding the pharmacological properties of emodin are explored, with particular emphasis on the modes of action of the molecule. In addition, the manuscript details the occurrence, biosynthesis and chemical synthesis of the compound, as well as its toxic effects on biotic systems.