Serial Exhaustive Extraction Revealed Antimicrobial and Antioxidant Properties of Platycerium stemaria (Beauv) Desv.

Microbial infections are increasing worldwide, and the widespread emergence of antibiotic-resistant pathogens poses a severe threat to public health. Medicinal plants are well-known sources of bioactive ingredients. This study was designed to determine the antimicrobial and antioxidant activities of extracts from Platycerium stemaria. The serial exhaustive extraction method using a solvent of increasing polarity from nonpolar (hexane) to polar (water) was designed to prepare crude extracts; liquid-liquid partition was used to fractionate of active extracts. The extracts and fractions were screened for antimicrobial activity on bacteria and yeasts using the microdilution method. The antioxidant activity was done using DPPH and FRAP assays. Out of the sixteen extracts screened, four (PsHex, PsH2O(H), PsMeOH(EA), and PsMeOH) exhibited potency with minimal inhibitory concentration (MIC) values ranging from 31.25 to 500 µg/mL. Out of the four extracts, two, including PsMeOH and PsMeOH(EA), exhibited DPPH radical scavenging activity with the antiradical power of 8.94 × 10-5 and 47.96 × 10-5, respectively, and ferric reducing antioxidant power values ranging from 0.34 to 61.53 µg equivalent Vit C/g of extract. The phytochemical screening of the promising crude extracts revealed flavonoids, glycosides, phenols, tannins, terpenoids, saponins, and anthraquinones. This study reports the antimicrobial and antioxidant activities of P. stemaria for the first time. The results showed that the serial exhaustive extraction approach used in this study allowed capturing the antimicrobial and antioxidant metabolites beyond the single extraction, indicating the need for a rigorous choice of an appropriate solvent and method for extracting P. stemaria. Further investigation is needed to characterize the active ingredients present in the promising extracts.

Antibacterial and Mode of Action of Extracts from Endophytic Fungi Derived from Terminalia mantaly, Terminalia catappa, and Cananga odorata.

Emerging drug-resistant bacteria creates an urgent need to search for antibiotics drugs with novel mechanisms of action. Endophytes have established a reputation as a source of structurally novel secondary metabolites with a wide range of biological activities. In the present study, we explore the antibacterial potential of endophytic fungi isolated from different tissues of Terminalia mantaly, Terminalia catappa, and Cananga odorata. The crude ethyl acetate extracts of 56 different endophytic fungi were screened against seven bacterial strains using the broth microdilution method. The antibacterial modes of action of the most active extracts (04) were evaluated using E. coli ATCC 25922 and H. influenzae ATCC 49247 strains. Both the DPPH and FRAP assays were used to investigate their antioxidant activity, and their cytotoxicity against the Vero cell line was evaluated using the MTT assay. Out of the 56 crude extracts tested, about 13% were considered very active, 66% partially active, and 21% nonactive against all tested bacterial strains with MIC values ranging from 0.32 µg/mL to 25 µg/mL. The four more potent extracts (MIC <5 µg/mL) (from Aspergillus sp. N454, Aspergillus sp. N13, Curvularia sp. N101, and Aspergillus sp. N18) significantly lysed the bacteria cells, increased outer membrane permeability, reduced salt tolerance, and inhibited bacterial catalase activity. They exhibited a DPPH free radical scavenging activity with IC50 ranging from 150.71 to 936.08 µg/mL. Three of the four potent extracts were noncytotoxic against the Vero cells line (CC50 > 100 µg/mL). Results from this investigation demonstrated that endophytes from Cameroonian medicinal plants might content potent antibacterial metabolites. The bioguided fractionation of these potent extracts is ongoing to isolate and characterise potential active ingredients.

Antiplasmodial potential and GC-MS fingerprint of endophytic fungal extracts derived from Cameroonian Annona muricata.

ETHNOPHARMACOLOGICAL RELEVANCE: Annona muricata (Annonaceae) is a commonly used medicinal plants in Cameroonian traditional medicines to treat various diseases including malaria. Previous studies have shown that extracts from this plant have antiplasmodial activity. AIM OF THE STUDY: This study aimed to explore the endophyic fungi associated with some parts of this plant for their ability to produce antiplasmodial metabolites. MATERIALS AND METHODS: One hundred and fifty-two endophytic fungi isolated from twelve different organs of A. muricata were cultured and the ethyl acetate extracts of conditioned media screened for antiplasmodial activity using the 96-well microtiter plate format SYBR green florescence assay against Chloroquine-sensitive Pf3D7 and Chloroquine-resistant PfINDO/PfDd2 strains of Plasmodium falciparum. RESULTS: Twenty-seven (17.76%) of fungi tested were found to completely inhibit the growth of Plasmodium parasites at 10 µg/mL. The 5.8S rDNA sequencing data revealed the strongly active (IC50 < 2 µg/mL against at least 2 P. falciparum strains) isolates to be Trichoderma afroharzianum AMrb7, Penicillium citrinum AMrb11, Neocosmospora rubicola AMb22, Penicillium tropicum AMb3, Penicillium citrinum AMrb23, Aspergillus versicolor AMb7, and Fusarium sp AMst1. Of these, the extracts from Penicillium citrinum AMrb11 (IC50 0.84-0.93 µg/mL) and Neocosmospora rubicola AMb22 (IC50 0.39-1.92 µg/mL) showed the highest promise against all three plasmodial strains with selectivity indices ranging from 34.71 to 180.97. Dynamic head space GC-MS analysis of ethyl acetate extracts of promising fungi revealed broad-spectrum antimicrobial compounds such as Penicidin, Aromadendrene, Ethyl p-methoxycinnamate, 2-Coumaranone and 2-Methyl resorcinol. CONCLUSION: These results have opened new avenues for discovery of novel antimalarial lead compounds from endophytic fungi associated with Annona muricata - a medicinally important plant.

Conditioned media and organic elicitors underpin the production of potent antiplasmodial metabolites by endophytic fungi from Cameroonian medicinal plants.

Plasmodial resistance to artemisinin-based combination therapies emphasizes the need for new drug development to control malaria. This paper describes the antiplasmodial activity of metabolites produced by endophytic fungi of three Cameroonian plants. Ethyl acetate extracts of fungi cultivated on three different media were tested against Plasmodium falciparum chloroquine-sensitive (Pf3D7) and chloroquine-resistant (PfINDO) strains using the SYBR green florescence assay. Selected endophytes were further grown in potato dextrose broth supplemented with small organic elicitors and their extracts tested for activity. The effect of elicitors on de novo metabolite synthesis was assessed by reverse-phase HPLC. Activity screening of 81 extracts indicated that Aspergillus niger 58 (IC50 2.25-6.69 µg/mL, Pf3D7), Fusarium sp. N240 (IC50 1.62-4.38 µg/mL, Pf3D7), Phomopsis sp. N114 (IC50 0.34-7.26 µg/mL, Pf3D7), and Xylaria sp. N120 (IC50 2.69-6.77 µg/mL, Pf3D7) produced potent extracts when grown in all three media. Further culture of these endophytes in potato dextrose broth supplemented with each of the eight small organic elicitors and subsequent extracts screening indicated the extract of Phomopsis sp. N114 grown with 1% 1-butanol to be highly selective and extremely potent (IC50 0.20-0.33 µg/mL; SI > 666). RPHPLC profiles of extracts of Phomopsis sp. N114 grown with or without 1-butanol showed some peaks of enhanced intensities in the former without any qualitative change in the chromatograms. This study showed the ability of selected endophytes to produce potent and selective antiplasmodial metabolites in varied culture conditions. It also showed how the production of desired metabolites can be enhanced by use of small molecular weight elicitors.