Bioassay-guided Fractionation Research Articles

Bioactive metabolites from endophytes of African club moss- Selaginella kraussiana (Kunze) A. Braun exhibit broad-spectrum pharmaceutical utilities

The search for potent natural compounds from plant and microbial sources is increasing logarithmically to combat various types of health issues of which multidrug-resistant microbes and problems related to oxidative stress are the prime ones. Novel secondary metabolites from endophytes have always been potent alternatives to contemporary antimicrobials and antioxidants. The present study shows that two potent endophytic fungal isolates Ascochyta medicaginicola SK16 and Phomopsis sp. SK5 synthesises non-proteinaceous, thermostable secondary metabolites with broad-spectrum antimicrobial, antioxidative, and cytotoxic activity. Bioassay-guided fractionation revealed that the Ethyl acetate (EA) extract of the cell-free culture extract (CFCE) of SK5 and SK16 constitutes ten and eleven secondary metabolites of which 2,4, -di tert butyl phenol, aziridine- 1,2-amino ethyl, actinobolin, bactobolin, azulene, menthyl acetate, phenol, p-[2-(methylamino)ethyl], and imidazole components were significant. SK16 exhibited strong antimicrobial activity against seventeen pathogens including Methicillin-resistant Staphylococcus aureus (MRSA), and C. albicans with a minimum microbicidal concentration of 100- 400 µg mL−1. These metabolites disrupt the microbial cell membranes causing lethal leakage of biomacromolecules- nucleic acid, protein, and potassium ions. Enzymes involved in the EMP pathway, TCA cycle, and gluconeogenesis were also blocked. Both the biofilm formation and transition of Candida cells from the yeast-to-hyphal form were inhibited. SK5 metabolites are potent radical scavengers with an IC50 of 34.25 ± 0.87 – 195.21 ± 5.73 µg mL−1 against DPPH, ABTS, FRAP, and H2O2 free radical generators and significantly increased antioxidant parameters in treated peritoneal macrophage cells of male Wistar rats. Also, SK5 metabolites were cytotoxic against breast (T47D), pancreatic (MIA Paca-2), lung (A549) and colon (HCT-116) cancer cells. The present findings suggest that metabolites from endophytes of African club moss hold potent pharmaceutical utilities.

Antiglycemic Activity of Vietnamese Scutellaria barbata Extract in Streptozotocin-Induced Diabetic Mice

Scutellariabarbata D. Don (S. barbata), a medicinal herb commonly used in traditional Chinese and Korean medicine, consists of bioactive neoclerodanes and flavonoids. However, its anti-hyperglycemic activity has not been reported. In this work, we used bioassay-guided fractionation to find effective anti-hyperglycemic fractions of S. barbata. The chemical components of these fractions were also determined structurally; they are scutellarin (1), luteolin (2), naringenin (3), tricin (4), baicalein (5), baicalin (6), apigenin (7), and wogonin (8). The most potent fraction SB6 consisting of flavonoids 5, 6, 7, and 8 showed 89% inhibition of α-glucosidase in the in vitro experiment. In further in vivo experiments on streptozotocin (STZ)-induced diabetic mice, SB6 exhibited significant anti-hyperglycemic activity comparable to metformin after 28 days of treatment. This fraction, however, did not show efficiency in reducing the weight gain of the treated mice.

Assessment of Anti-Prostate Cancer Activity among Four Seaweeds, with Focus on Caulerpa lentillifera J.Agardh.

In response to a global shift towards health-conscious and environmentally sustainable food choices, seaweed has emerged as a focus for researchers due to its large-scale cultivation potential and the development of bioactive substances. This research explores the potential anticancer properties of seaweed extracts, focusing on analyzing the impact of four common edible seaweeds in Taiwan on prostate cancer (PCa) cells' activity. The study used bioassay-guided fractionation to extract Cl80 from various seaweeds with androgen receptor (AR)-inhibitory activity. Cl80 demonstrated effective suppression of 5α-dihydrotestosterone (DHT)-induced AR activity in 103E cells and attenuated the growth and prostate-specific antigen (PSA) protein expression in LNCaP and 22Rv1 cells. Additionally, Cl80 exhibited differential effects on various PCa cell lines. Concentrations above 5 μg/mL significantly inhibited LNCaP cell proliferation, while 22Rv1 cells were more resistant to Cl80. PC-3 cell proliferation was inhibited at 5 μg/mL but not completely at 50 μg/mL. A clonogenic assay showed that at a concentration of 0.5 μg/mL, the colony formation in LNCaP and PC-3 cells was significantly reduced, with a dose-dependent effect. Cl80 induced apoptosis in all PCa cell types, especially in LNCaP cells, with increased apoptotic cells observed at higher concentrations. Cl80 also decreased the mitochondrial membrane potential (ΔΨm) in a dose-dependent manner in all PCa cell lines. Furthermore, Cl80 suppressed the migration ability of PCa cells, with significant reductions observed in LNCaP, 22Rv1, and PC-3 cells at various concentrations. These compelling findings highlight the promising therapeutic potential of C. lentillifera J.Agardh and its isolated compound Cl80 in the treatment of PCa.

Characterisation and expression analyses of two putative cerato-platanin proteins isolated from Ganoderma boninense Pat.

Cerato-platanin (CP) proteins are small extracellular proteins that play essential roles in the plant-pathogen interactions of some economically important filamentous fungi. At present, there is a lack of information about the roles of CP proteins secreted by Ganoderma boninense, the causal agent of basal stem rot disease in African oil palm, during the early host-pathogen interaction. This work studies the properties and roles of two CP proteins from G. boninense strain PER71. Using bioassay-guided fractionation and nano-LC-MS/MS analysis, we identified peptide matches for G. sinense ZZ0214-1 CP-like and other elicitor proteins in the culture filtrate of G. boninense strain PER71. We found that GbCP2 and GbCP6, two out of nine CP gene models identified in the fungal genome of G. boninense strain PER71, exhibited potential effector properties. We further validated their effector properties by isolating and characterising their coding regions. Interestingly, GbCP6 showed a significant expression level at 11 days post inoculation, indicating its significance in fungal pathogenesis. On the other hand, GbCP2 did not show differential expression at any sampling point, suggesting a different function or role in the pathogenicity of G. boninense. This study identifies GbCP6 as a potential biomarker for early detection of basal stem rot disease in oil palm.

Compounds isolation from Syzygium cumini leaf extract against the Vibrio species in shrimp through bioassay-guided fractionation

This study was conducted to isolate and identify the bioactive compounds from the ethanolic extract of Syzygium cumini leaf against Vibrio species through a bioassay-guided fractionation. The ethanol extract was exposed to silica gel chromatography followed by reversed phase HPLC to isolate the most effective fraction against V. parahaemolyticus. Using further UHPLC-orbitrap-ion trap mass spectrometry, five compounds were isolated with broad-spectrum potency against a range of Vibrio species viz. V. parahaemolyticus, V. alginolyticus, V. harveyi, V. vulnificus and V. anguillarum. The IC50 values for the compounds ranged from 8 to 48 µg/mL against the most sensitive species V. vulnificus and 58 to >400 µg/mL against V. alginolyticus. The results of the toxicity tests demonstrated that the compounds were not harmful for shrimp. The study’s findings indicate that S. cumini leaf extract may contain bioactive molecules that are able to be substituted for antibiotics to treat vibriosis in shrimp farming.

Antiprotozoal Activity of Plants Used in the Management of Sleeping Sickness in Angola and Bioactivity-Guided Fractionation of Brasenia schreberi J.F.Gmel and Nymphaea lotus L. Active against T. b. rhodesiense.

Folk medicine is widely used in Angola, even for human African trypanosomiasis (sleeping sickness) in spite of the fact that the reference treatment is available for free. Aiming to validate herbal remedies in use, we selected nine medicinal plants and assessed their antitrypanosomal activity. A total of 122 extracts were prepared using different plant parts and solvents. A total of 15 extracts from seven different plants exhibited in vitro activity (>70% at 20 µg/mL) against Trypanosoma brucei rhodesiense bloodstream forms. The dichloromethane extract of Nymphaea lotus (leaves and leaflets) and the ethanolic extract of Brasenia schreberi (leaves) had IC50 values ≤ 10 µg/mL. These two aquatic plants are of particular interest. They are being co-applied in the form of a decoction of leaves because they are considered by local healers as male and female of the same species, the ethnotaxon "longa dia simbi". Bioassay-guided fractionation led to the identification of eight active molecules: gallic acid (IC50 0.5 µg/mL), methyl gallate (IC50 1.1 µg/mL), 2,3,4,6-tetragalloyl-glucopyranoside, ethyl gallate (IC50 0.5 µg/mL), 1,2,3,4,6-pentagalloyl-β-glucopyranoside (IC50 20 µg/mL), gossypetin-7-O-β-glucopyranoside (IC50 5.5 µg/mL), and hypolaetin-7-O-glucoside (IC50 5.7 µg/mL) in B. schreberi, and 5-[(8Z,11Z,14Z)-heptadeca-8,11,14-trienyl] resorcinol (IC50 5.3 µg/mL) not described to date in N. lotus. Five of these active constituents were detected in the traditional preparation. This work provides the first evidence for the ethnomedicinal use of these plants in the management of sleeping sickness in Angola.

Rethinking the basic action modes of herbal medicine and pondering classical standardization

Over the past two decades, the secondary metabolite platform has determined the scientific direction of herbal medicines, while plant sources have been assumed to be the object of lead discoveries through bioassay-guided fractionation efforts. Nonetheless, the majority of purification programs have resulted in fractions and pure compounds with much lower efficacy than their parent extracts. It is then assumed that co-working action modes among chemical constituents occur in the herbal preparations. Primary metabolites (polysaccharides, peptides, and fatty acids) and mineral groups, on the other hand, have been neglected in the herbal effect contributions. This review aims to understand the interplay of secondary metabolites in herbal preparations, particularly how they interact with primary metabolites and mineral groups. Thus, by adhering to classical methods, it is possible to address certain aspects that modern standardization lacks, thereby facilitating a more comprehensive approach to these issues.

Ellagitannins from pomegranate (Punica granatum L.) flower with xanthine oxidase and α-glucosidase inhibitory activities

In the course of our research on utilizing edible flower resources to promote health, five ellagitannins (1–5) were isolated from pomegranate (Punica granatum L.) flower by bioassay-guided fractionation. Among them, mallotusinin (1) and tercatain (2) were first isolated from pomegranate flower, and punicatannin D (4) was an unprecedented compound. Compounds 1–3 exhibited significant xanthine oxidase (XO) inhibitory activity compared with allopurinol, and a potent inhibition on α-glucosidase that was superior to acarbose was observed for compounds 1–4. The XO inhibitory activity is positively correlated with the amount of galloyl moiety (3 > 2 > 1). Presenting tetrahydroxydibenzofuranoyl substituent or increasing the number of galloyl group enhanced the inhibition capacity of α-glucosidase (1 > 3 > 2). The inhibitory mechanism was further clarified in terms of kinetic studies and molecular docking. This study revealed that the ellagitannins present in pomegranate flower might be promising functional food ingredients with anti-hyperuricemia and anti-hyperglycemia properties.

Glucoregulatory Properties of a Protein Hydrolysate from Atlantic Salmon (Salmo salar): Preliminary Characterization and Evaluation of DPP-IV Inhibition and Direct Glucose Uptake In Vitro.

Metabolic disorders are increasingly prevalent conditions that manifest pathophysiologically along a continuum. Among reported metabolic risk factors, elevated fasting serum glucose (FSG) levels have shown the most substantial increase in risk exposure. Ultimately leading to insulin resistance (IR), this condition is associated with notable deteriorations in the prognostic outlook for major diseases, including neurodegenerative diseases, cancer risk, and mortality related to cardiovascular disease. Tackling metabolic dysfunction, with a focus on prevention, is a critically important aspect for human health. In this study, an investigation into the potential antidiabetic properties of a salmon protein hydrolysate (SPH) was conducted, focusing on its potential dipeptidyl peptidase-IV (DPP-IV) inhibition and direct glucose uptake in vitro. Characterization of the SPH utilized a bioassay-guided fractionation approach to identify potent glucoregulatory peptide fractions. Low-molecular-weight (MW) fractions prepared by membrane filtration (MWCO = 3 kDa) showed significant DPP-IV inhibition (IC50 = 1.01 ± 0.12 mg/mL) and glucose uptake in vitro (p ≤ 0.0001 at 1 mg/mL). Further fractionation of the lowest MW fractions (<3 kDa) derived from the permeate resulted in three peptide subfractions. The subfraction with the lowest molecular weight demonstrated the most significant glucose uptake activity (p ≤ 0.0001), maintaining its potency even at a dilution of 1:500 (p ≤ 0.01).

Effects of Naphthoquinones from the Roots of Onosma Armeniacum Klokov on Wound Healing.

In Turkish folk medicine, the roots of Onosma armeniacum Klokov are used to heal wounds, burns, hemorrhoids, hoarseness, dyspnea, stomach ulcers, and abdominal aches. The objective was to evaluate the plant's ethnopharmacological applications using in vivo pharmacological experimental models. In vivo linear incision and circular excision the wound models were used to assess the wound healing activity along with histopathological investigation. The active component(s) were isolated and identified after being exposed to several chromatographic separation procedures on the primary extract. The n-hexane-dichloromethane mixture extract was subjected to chromatographic separation after the wound-healing activity was confirmed. Deoxyshikonin (1), β,β-dimethylacrylshikonin (2), α-methyl-n-butylshikonin (3), isovalerylshikonin (4), acetylshikonin (5), β-hydroxyisovalerylshikonin (6), and 5,8-O-dimethylacetylshikonin (7) were identified as the structures of the isolated compounds. The efficacy of O. armeniacum to heal wounds was investigated in this study. Shikonin derivatives were identified as the primary active components of the roots by bioassay-guided fractionation and isolation procedures.

Bioassay-Guided Fractionation of Pittosporum angustifolium and Terminalia ferdinandiana with Liquid Chromatography Mass Spectroscopy and Gas Chromatography Mass Spectroscopy Exploratory Study

Bioprospecting native Australian plants offers the potential discovery of latent and novel bioactive compounds. The promising cytotoxic and antibacterial activity of methanolic extracts of Pittosporum angustifolium and Terminalia ferdinandiana led to further fractionation and isolation using our laboratory’s bioassay-guided fractionation protocol. Hence, the aim of this study was to further evaluate the bioactivity of the fractions and subfractions and characterize bioactive compounds using liquid chromatography mass spectroscopy (LC-MS/MS) and gas chromatography MS (GC-MS). Compounds tentatively identified in P. angustifolium Fraction 1 using LC-ESI-QTOF-MS/MS were chlorogenic acid and/or neochlorogenic acid, bergapten, berberine, 8′-epitanegool and rosmarinic acid. GC-MS analysis data showed the presence of around 100 compounds, mainly comprising carboxylic acids, sugars, sugar alcohols, amino acids and monoalkylglycerols. Furthermore, the fractions obtained from T. ferdinandiana flesh extracts showed no cytotoxicity, except against HT29 cell lines, and only Fraction 2 exhibited some antibacterial activity. The reduced bioactivity observed in the T. ferdinandiana fractions could be attributed to the potential loss of synergy as compounds become separated within the fractions. As a result, the further fractionation and separation of compounds in these samples was not pursued. However, additional dose-dependent studies are warranted to validate the bioactivity of T. ferdinandiana flesh fractions, particularly since this is an understudied species. Moreover, LC-MS/GC-MS studies confirm the presence of bioactive compounds in P. angustifolium Fraction 1/subfractions, which helps to explain the significant acute anticancer activity of this plant. The screening process designed in this study has the potential to pave the way for developing scientifically validated phytochemical/bioactivity information on ethnomedicinal plants, thereby facilitating further bioprospecting efforts and supporting the discovery of novel drugs in modern medicine.

Anti-diabetic and anti-inflammatory bioactive hits from Coriaria intermedia Matsum. stem and Dracontomelon dao (Blanco) Merr. & Rolfe bark through bioassay-guided fractionation and liquid chromatography-tandem mass spectrometry.

Women have been found to be at a higher risk of morbidity and mortality from type 2 diabetes mellitus (T2DM) and asthma. α-Glucosidase inhibitors have been used to treat T2DM, and arachidonic acid 15-lipoxygenase (ALOX15) inhibitors have been suggested to be used as treatments for asthma and T2DM. Compounds that inhibit both enzymes may be studied as potential treatments for people with both T2DM and asthma. This study aimed to determine potential anti-diabetic and anti-inflammatory bioactive hits from Coriaria intermedia Matsum. stem and Dracontomelon dao (Blanco) Merr. & Rolfe bark. A bioassay-guided fractionation framework was used to generate bioactive fractions from C. intermedia stem and D. dao bark. Subsequently, dereplication through ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) and database searching was performed to putatively identify the components of one bioactive fraction from each plant. Seven compounds were putatively identified from the C. intermedia stem active fraction, and six of these compounds were putatively identified from this plant for the first time. Nine compounds were putatively identified from the D. dao bark active fraction, and seven of these compounds were putatively identified from this plant for the first time. One putative compound from the C. intermedia stem active fraction (corilagin) has been previously reported to have inhibitory activity against both α-glucosidase and 15-lipoxygenase-1. It is suggested that further studies on the potential of corilagin as an anti-diabetic and anti-inflammatory treatment should be pursued based on its several beneficial pharmacological activities and its low reported toxicity.

Bioassay-guided fractionations of Cannabis sativa extract and HPLC-assisted purifications of anti-proliferative active fractions lead to the isolation of 16 known and one new phytomolecule and their in-silico analysis

Bioassay-guided fractionations of Cannabis sativa extract and HPLC-assisted purifications of anti-proliferative active fractions lead to the isolation of 16 known and one new phytomolecule and their in-silico analysis

Cytotoxicity, antifungal and antioxidant properties of Lonchocarpus capassa leaf extracts

Background: Lonchocarpus capassa is a medicinal plant used to treat diseases such as fungal infections, diarrhoea, oral candidiasis, and stomach complaints in South Africa.Aim: The study aimed to investigate the cytotoxicity and antifungal compounds isolated from L. capassa leaf extracts.Setting: The study was conducted in Muduluni village, Makhado Local Municipality, Limpopo province.Methods: Leaf extracts were screened for antifungal activity against fungal pathogens: Candida albicans, Cryptococcus neoformans, and Aspergillus fumigatus. Bioassay-guided fractionation using column chromatography of the acetone extract led to the isolation of six antifungal compounds. Nuclear Magnetic Resonance spectroscopy and Mass Spectrometry were used for the identification of compounds. The antioxidant activity of the plant extracts was investigated using 1,1-diphenyl-2-picrylhydrazyl (DPPH) assays. Cytotoxicity of isolated compounds was determined using the 3-(4,5-dimethylthiazol)-2,5-diphenyl tetrazolium bromide (MTT) assay against Vero monkey kidney cells.Results: The plant extract had an excellent minimum inhibitory concentrations (MIC) value of 40 μg/mL against the microorganisms. Compound 1 was identified as Lupeol, Compound 3 as Friedelin, and Compound 4 as 6-(γ,γ-Dimethylallyl)-3’,4’-dimethoxy-6”,6”-dimethylpyrano-[2”,3”:7,8]-flavanone (Compound 4). Compounds 2 and 5 were not identified because of the presence of mixtures of long-chain fatty acids. Friedelin was the most active radical scavenger in the DPPH assay. The compounds were not toxic with an LC50 value of ˃ 0.2 mg/mL.Conclusion: Screening of medicinal plants could provide lead to the discovery of novel antifungal agents.Contribution: The results support the traditional use of L. capassa to combat fungal infections in humans.

Identification of Lydicamycins as Main Antioomycete Compounds from the Biocontrol Agent Streptomyces sp. NEAU-S7GS2.

Oomycetes are well-known phytopathogens that seriously threaten many important crops worldwide. In this study, the endophytic actinobacterium Streptomyces sp. NEAU-S7GS2 demonstrated significant antagonistic activity against Phytophthora and Pythium and showed a potent biocontrol effect on suppression of soybean phytophthora root rot and pepper phytophthora blight. Two compounds were subsequently isolated as the main active components by bioassay-guided fractionation and identified as lydicamycins A and B. These two compounds showed high antioomycete activity against Phytophthora and Pythium with EC50 values of 0.73-2.67 μg/mL, which are equal to or lower than those of commercialized drug metalaxyl. In vivo bioassay using detached leaves demonstrated that lydicamycin A had a better control efficiency against soybean phytophthora root rot than metalaxyl. Taken together, these results suggest that the biocontrol agent Streptomyces sp. NEAU-S7GS2 and lydicamycins have the potential to be developed as promising pesticides to control diseases caused by oomycetes.

Cytotoxicity of luteolin, a flavonoid compound isolated from &lt;i&gt;Anthemis palestina&lt;/i&gt;

Purpose: To determine the active principle responsible for the cytotoxic effect of Anthemis palestina (Reut. ex Kotschy) Reut. ex Boiss. (Asteraceae).&#x0D; Methods: A bioassay-guided fractionation was used to isolate the active principle, luteolin, which structure was elucidated using 1H and 13C NMR. The cytotoxic effects of luteolin and doxorubicin (positive control) in the breast cancer cell lines MDA-MB-231 and MCF-7, and in normal fibroblasts, were determined using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay.&#x0D; Results: Luteolin was isolated from the ethyl acetate extract of the aerial parts of Anthemis palestina. With an IC50 value of 14.91 ± 5.77 µM for MDA-MB-231 cells, luteolin was less active than doxorubicin. However, with respect to MCF-7 cells, there was no significant difference in the cytotoxicity values of luteolin and doxorubicin, with IC50 value of 29.28 ± 11.85 µM for luteolin. However, with an IC50 value of 51.39 ± 18.51 µM against fibroblasts, luteolin was significantly safer than doxorubicin.&#x0D; Conclusion: Luteolin might be responsible for the cytotoxicity of Anthemis palestina. The high level of luteolin cytotoxicity indicates the potential benefits of Anthemis palestina, not only in terms of its taste, but also for its likely positive therapeutic impact on cancer, especially breast cancer.

Characterization by LC-MS/MS and Antioxidant Activity of Extract and its Different Fractions of Calotropis procera Young Leaves

Objective: Plants are extremely useful as a source of medication. Calotropis procera is a plant that is used to treat a variety of illnesses in traditional medical and folklore systems. In this work, bioassay guided liquid-liquid fractionation, phytochemical and molecular spectroscopic analysis and the antioxidant activity of young leaves of Calotropis procera were evaluated. Methods: Bioassay-guided fractionation was carried out on the basis of polarity of solvents and UV, FTIR, LCMS profiling were conducted by using the standard methods. The DPPH scavenging assay was used to determine the antioxidant activity. Results: The extractive value for the ethyl acetate fraction was 9.14 % w/w. The Total phenolic content was highest in the water fraction 83.63±0.0788 GAE/g of dry weight of extract. UV spectroscopy was used to standardize the fractions. The Presence of various functional groups in the extract/fraction was confirmed by FTIR spectroscopy. Based on LCMS, fifty compounds were identified in the hydro-ethanolic extract/fractions. Among all the fractions, the chloroform fraction showed the greatest antioxidant properties (IC50 values=8.03 mg/mL). Conclusion: On the basis of the result of the study it can be concluded that Calotropis procera have the potential to become a promising herb for the treatment of various ailments.

Efficient discovery of active isolates from Dioscorea spongiosa by the combination of bioassay-guided macroporous resin column chromatography and high-speed counter-current chromatography.

In the present study, twelve compounds from Dioscorea spongiosa were successfully purified by an efficient technique combined bioassay-guided fractionation macroporous resin column chromatography (MRCC) pretreatment and high-speed counter-current chromatography (HSCCC) separation for the first time. Then, D101 MRCC was used to fractionate the crude extract into five parts, which further applied the bioassay-guided fractionation strategy to screen the active fractions of 2 and 4. As for the separation, 200mg Fr.2 was purified by HSCCC using EtOAc/n-BuOH/H2 O (2:2:3, v/v), leading to annulatomarin (1), dioscoresides C (2), diosniponol C (3), methyl protodioscin (4), pseudoprotodioscin (5), protogracillin (6), as well as 200mg Fr.4 yielding montroumarin (7), dioscorone A (8), diosniponol D (9), protodioscin (10), gracillin (11), and dioscin (12) using CH2 Cl2 /MeOH/H2 O (3:3:2, v/v) with the purities over 95.0%. Finally, the isolates were assayed for their anti-inflammatory, urico-lowering, and anti-diabetic activities in vitro, which indicated that the steroidal saponins of 5, 6, and 11 showed all these three activities.

Evaluation of the potential of endophytic Trichoderma sp. isolated from medicinal plant Ampelopsis japonica against MRSA and bioassay-guided separation of the anti-MRSA compound.

Endophytic fungi have been recognized as a valuable source for the production of biologically active compounds with potential applications in various domains. This study aimed to isolate endophytic fungi from Ampelopsis japonica (Thunb.) Makino and assess their anti-MRSA activity. Meanwhile, chromatographic separation techniques were applied to analyze the constituents of endophytic fungal secondary metabolites. The isolate BLR24, which exhibited strong inhibition activity against MRSA, was identified as Trichoderma virens based on morphological characteristics and ITS sequence analyses. The ethyl acetate extract of BLR24 (EA-BLR24) showed good anti-MRSA activity with the MIC and MBC values of 25 μg/mL and 50 μg/mL, separately. The inhibition of biofilm formation was up to 34.67% under MIC concentration treatment. Meanwhile, EA-BLR24 could significantly reduce the expression of biofilm-related genes (icaA, sarA, and agrA) of MRSA. Based on LC-MS/MS analysis, twenty compounds in EA-BLR24 could be annotated using the GNPS platform, mainly diketopiperazines. The anti-MRSA compound (Fr.1.1) was obtained from EA-BLR24 by bioassay-guided fractionation and determined as gliotoxin. The results indicated that endophytic Trichoderma virens BLR24 isolated from the medical plant A. japonica roots could be a promising source of natural anti-MRSA agents. Endophytic fungal secondary metabolites are abundant in biologically active compounds. Endophytic fungi from medicinal plants could be a source yielding bioactive metabolites of pharmaceutical importance.

Molecular level interaction, HOMO-LUMO, MEP, UV–Vis, Hirshfeld, topological analysis, and in-vitro of isoflavones from Eremostachys Vicaryi Benth. Ex Hook. f.

Eremostachys vicaryi Benth. Ex Hook. f. (Labiatae), a Pakistani (Baluchistan) herbal medicinal plant commonly known as “Stagh,” which has a sharp taste and smell. It acts as an antiseptic and antimicrobial agent and is used for fish poisoning, treating skin diseases, and holding a cooling effect. Following the assessment of the plant extract's cytotoxic and phytotoxic assays using phytochemical screening, the antioxidant and anti-inflammatory properties of the pure ingredients that were separated from the ethyl acetate soluble fractions have been evaluated. The bioassay-guided fractionation and purification of E. vicaryi B. crude methanol extract yielded two isoflavones (1–2) that showed promising antioxidative activities with DPPH and cytochrome-c-reduction assays. Compounds 1 and 2 have shown considerable anti-inflammatory efficacy, 1 being a good inhibitor of the superoxide within the living system. Initial assumptions on the one-electron transfer process indicated that Compound 1 has a smaller ionization potential value compared to Compound 2 and so has greater antioxidant potential than its predecessor, isoflavone. Moreover, the anti-inflammatory and antioxidant activity of isoflavones was also probed by first-principles calculations. Based on the findings, it has been concluded that the crude extract of E. vicaryi B. and its organic components exhibit noteworthy phytotoxic and cytotoxic properties. The isolated pure isoflavones 1 and 2 showed significant antioxidant and anti-inflammatory activities. Solute-solvent interactions were studied and reported for both compounds. An analysis was conducted on the UV–Vis electronic transition for different solvents. Using a Multiwave function analyzer the topological findings such as ELF, LOL, and RDG were done to comprehend the surface electron density, delocalization, localized orbital, and weak interaction in the compounds. The antioxidant activity is evaluated by radical methods using DPPH, and cytochrome-c-reduction assays using ascorbic acid as a standard. Through cell-based respiratory burst testing, the anti-inflammatory activity of standard indomethacin was examined. The ground state geometries of isoflavones and reference compounds were optimized by Density Functional Theory- (DFT) by computing various electronic properties. In-vitro bioassay results were validated by first-principles calculations.