Predicting the bioactive compounds of Lentinula edodes and elucidating its interaction with genes associated to obesity through network pharmacology and in-vitro cell-based assay.

Mushrooms are traditionally used for various medicinal purposes in traditional oriental medicine. The Japanese and Chinese are familiar with the medicinal macro fungus Lentinula edodes (Shiitake mushroom). This study aims to evaluate the role of chemical compounds from L. edodes using network pharmacology and in-vitro studies for management of Obesity. Bioactive compounds in extracts of L. edodes were identified by GC-MS analysis. Compounds were later screened for their drug-like property by Lipinski's rule. In addition, public databases (SEA, STP, Omim and DisGenet) were searched to identify genes associated with selected molecules and obesity, as well as genes that overlap obesity target genes with genes related to L. edodes. Additionally, analysis was performed using Enrichr KG to predict the disease targets of L. edodes. Finally, network was constructed between the overlapping genes and bioactive molecules using Rstudio. Further in-vitro studies were carried out using 3T3-L1 cell line. The genes related to the selected compounds and obesity were identified and overlapped. The disease targets of L. edodes was predicted by enrichment analysis and was found to be linked to obesity. Furthermore, the hub gene was found to be fatty acid amide hydrolase, and the key bioactive compound was hexadecanoic acid methyl ester. The in-vitro cell culture studies confirmed the inhibition of adipogenesis in mushroom extract-treated 3T3-L1 cells and the augmentation of adiponectin. The study suggests that the hub gene fatty acid amide hydrolase might alleviate obesity by inhibiting arachidonoyl ethanolamide signaling, which would enhance the action of fatty acid amide hydrolase and limit appetite in L. edodes extract.

Soil treatment using a biosurfactant producing bacterial consortium in rice fields contaminated with oily sludge- a sustainable approach.

A consortium of two biosurfactant-producing bacteria (Bacillus pumilus KS2 and Bacillus cereus R2) was developed to remediate petroleum hydrocarbon-contaminated paddy soil. Soil samples from a heavily contaminated rice field near Assam's Lakwa oilfield were collected and placed in earthen pots for treatment. After each month of incubation, 50 g of soil from each earthen pot was collected, and the soil TPH (ppm) in each sample was determined. The extracted TPH samples were analysed by Gas chromatography-mass spectrometry (GC-MS) to confirm microbial degradation. The soil samples were examined for changes in pH, conductivity, total organic content (TOC), water holding capacity, and total nitrogen content in addition to TPH degradation. An increasing trend in TPH degradation was observed with each passing month. After six months of treatment, the sample with the lowest initial TPH concentration (1735 ppm) had the highest degradation (91.24%), while the soil with the highest amount of TPH (5780 ppm) had the lowest degradation (74.35%). A wide range of aliphatic hydrocarbons found in soil samples was degraded by the bacterial consortium. The soil samples contained eight different low- and high-molecular-weight PAHs. Some were fully mineralized, while others were significantly reduced. With the decrease in the TPH level in the polluted soil, a significant improvement in the soil's physicochemical qualities (such as pH, electrical conductivity, total organic content, and water-holding capacity) was observed.

Report on Vincristine-Producing Endophytic Fungus Nigrospora zimmermanii from Leaves of Catharanthus roseus.

Vincristine is an anti-cancer compound and one of the most crucial vinca alkaloids produced by the medicinal plant Catharanthus roseus (L.) G. Don. (Apocynaceae). This plant is home to hundreds of endophytic microbes, which produce a variety of bioactive secondary metabolites that are known for their medicinal properties. In this study, we focused on isolating an endophytic fungus that could increase the yield of vincristine under laboratory conditions as an alternative to plant-mediated extraction of vincristine. The endophytic fungus Nigrospora zimmermanii (Apiosporaceae) was isolated from Catharanthus roseus and it was found to be producing the anticancer compound vincristine. It was identified using high-performance thin-layer chromatography by matching the Rf value and spectral data with the vincristine standard and mass spectrometry data and the reference molecule from the PubChem database. The generation study of this microbe showed that the production of vincristine in the parent fungus was at its maximum, i.e., 5.344 µg/mL, while it was slightly reduced in subsequent generations. A colonization study was also performed and it showed that the fungus N. zimmermanii was able to re-infect the plant Catharanthus roseus after 20 days of inoculation. The colonization study showed that N. zimmernanii could infect the plant after isolation. This method is an efficient and easy way to obtain a high yield of vincristine, as compared to plant-mediated production.

Effect of Vitamin-D-Enriched Edible Mushrooms on Vitamin D Status, Bone Health and Expression of CYP2R1, CYP27B1 and VDR Gene in Wistar Rats.

Vitamin D deficiency is highly prevalent in India and worldwide. Mushrooms are important nutritional foods, and in this context shiitake (Lentinula edodes), button (Agaricus bisporus) and oyster (Pleurotus ostreatus) mushrooms are known for their bioactive properties. The application of ultraviolet (UV) irradiation for the production of substantial amounts of vitamin D2 is well established. Levels of serum 25-hydroxy vitamin D (25-OHD), parathyroid hormone (PTH), calcium, phosphorus and alkaline phosphatase (ALP) were significantly (p < 0.05) improved in vitamin-D-deficient rats after feeding with UVB irradiated mushrooms for 4 weeks. Further, microscopic observations indicate an improvement in the osteoid area and the reduction in trabecular separation of the femur bone. In addition, the level of expression of the vitamin D receptor (VDR) gene and genes metabolizing vitamin D were explored. It was observed that in mushroom-fed and vitamin-D-supplemented groups, there was upregulation of CYP2R1 and VDR, while there was downregulation of CYP27B1 in the liver. Further, CYP2R1 was downregulated, while CYP27B1 and VDR were upregulated in kidney tissue.

ZnO Nanoparticles of Rubia cordifolia Extract Formulation Developed and Optimized with QbD Application, Considering Ex Vivo Skin Permeation, Antimicrobial and Antioxidant Properties.

The objective of the current research is to develop ZnO-Manjistha extract (ZnO-MJE) nanoparticles (NPs) and to investigate their transdermal delivery as well as antimicrobial and antioxidant activity. The optimized formulation was further evaluated based on different parameters. The ZnO-MJE-NPs were prepared by mixing 10 mM ZnSO4·7H2O and 0.8% w/v NaOH in distilled water. To the above, a solution of 10 mL MJE (10 mg) in 50 mL of zinc sulfate was added. Box-Behnken design (Design-Expert software 12.0.1.0) was used for the optimization of ZnO-MJE-NP formulations. The ZnO-MJE-NPs were evaluated for their physicochemical characterization, in vitro release activity, ex vivo permeation across rat skin, antimicrobial activity using sterilized agar media, and antioxidant activity by the DPPH free radical method. The optimized ZnO-MJE-NP formulation (F13) showed a particle size of 257.1 ± 0.76 nm, PDI value of 0.289 ± 0.003, and entrapment efficiency of 79 ± 0.33%. Drug release kinetic models showed that the formulation followed the Korsmeyer-Peppas model with a drug release of 34.50 ± 2.56 at pH 7.4 in 24 h. In ex vivo studies ZnO-MJE-NPs-opt permeation was 63.26%. The antibacterial activity was found to be enhanced in ZnO-MJE-NPs-opt and antioxidant activity was found to be highest (93.14 ± 4.05%) at 100 µg/mL concentrations. The ZnO-MJE-NPs-opt formulation showed prolonged release of the MJE and intensified permeation. Moreover, the formulation was found to show significantly (p < 0.05) better antimicrobial and antioxidant activity as compared to conventional suspension formulations.

Enhancement of vincristine under in vitro culture of Catharanthus roseus supplemented with Alternaria sesami endophytic fungal extract as a biotic elicitor.

Vincristine, one of the major vinca alkaloid of Catharanthus roseus (L.) G. Don. (Apocynaceae), was enhanced under in vitro callus culture of C. roseus using fungal extract of an endophyte Alternaria sesami isolated from the surface-sterilized root cuttings of C. roseus. Vindoline, a precursor molecule for vincristine production, was detected for the first time in the fungal endophyte A. sesami which was used as a biotic elicitor in this study to enhance vincristine content in the C. roseus callus. It was identified using high-performance liquid chromatography and mass spectroscopy techniques by matching retention time and mass data with reference molecule. Supplementing the heat sterilized A. sesami endophytic fungal culture extract into the callus culture medium of C. roseus resulted in the enhancement of vincristine content in C. roseus callus by 21.717% after 105-day culture.

Okra (Abelmoschus Esculentus) as a Potential Dietary Medicine with Nutraceutical Importance for Sustainable Health Applications.

Recently, there has been a paradigm shift from conventional therapies to relatively safer phytotherapies. This divergence is crucial for the management of various chronic diseases. Okra (Abelmoschus esculentus L.) is a popular vegetable crop with good nutritional significance, along with certain therapeutic values, which makes it a potential candidate in the use of a variety of nutraceuticals. Different parts of the okra fruit (mucilage, seed, and pods) contain certain important bioactive components, which confer its medicinal properties. The phytochemicals of okra have been studied for their potential therapeutic activities on various chronic diseases, such as type-2 diabetes, cardiovascular, and digestive diseases, as well as the antifatigue effect, liver detoxification, antibacterial, and chemo-preventive activities. Moreover, okra mucilage has been widely used in medicinal applications such as a plasma replacement or blood volume expanders. Overall, okra is considered to be an easily available, low-cost vegetable crop with various nutritional values and potential health benefits. Despite several reports about its therapeutic benefits and potential nutraceutical significance, there is a dearth of research on the pharmacokinetics and bioavailability of okra, which has hampered its widespread use in the nutraceutical industry. This review summarizes the available literature on the bioactive composition of okra and its potential nutraceutical significance. It will also provide a platform for further research on the pharmacokinetics and bioavailability of okra for its possible commercial production as a therapeutic agent against various chronic diseases.

Metabolic differentiation and quantification of gymnemic acid in Gymnema sylvestre (Retz.) R.Br. ex Sm. leaf extract and its fermented products.

INTRODUCTION: Gymnemagenin is the bioactive metabolite found in Gymnema sylvestre leaves and possesses different therapeutic potential. Due to its lower abundance and higher market potential, gymnemagenin was obtained from chemical conversion and bacterial biotransformation. OBJECTIVE: To obtain the probiotic-based fermentative conversion of gymnemic acid-enriched G. sylvestre leaf extract to gymnemagenin-containing nutraceuticals and its metabolites based chromatographic comparison. MATERIAL AND METHODS: Gymnema sylvestre leaves were extracted through soxhalation, and the extract was prepared and characterised. Gymnemic acid was fermented, separately, by Lactobacillus casei, Lactobacillus rhamnosus, Bifidobacterium bifidum, and by their mix co-culture. The fermented materials were analysed for their gymnemagenin content, antioxidant potential, antidiabetic potential, and metabolomics analysis. RESULTS: Extraction yielded about 35% w/w of raw plant material, and 8.5% was found to be as total saponin content. Extract at higher concentration (≥ 5%, w/v) significantly altered the growth behaviour of probiotics. High-performance thin-layer chromatography (HPTLC) based quantification of gymnemagenin revealed that a maximum increase of 95.5% gymnemagenin was found in extract incubated with B. bifidum followed by mix co-culture containing (B. bifidum, L. casei, and L. rhamnosus), L. casei, and L. rhamnosus. However, liquid chromatography mass spectrometry (LC-MS) analysis resulted in the identification of a total of 56 metabolites. CONCLUSION: Chromatographically profiled, and probiotic-based fermented G. sylvestre leaves can be used as a potent nutraceutical for diabetes and other metabolic disorders.

Trigonelline inhibits intestinal microbial metabolism of choline and its associated cardiovascular risk.

Gut microbiota based metabolism of choline produces trimethylamine (TMA) which is further converted to a pro-atherosclerotic metabolite, trimethylamine-N-oxide (TMAO) by flavin monooxygenase (FMO3). Trigonelline from the plant Trigonella foenum-graecum has been reported for the treatment of CVD. Aim of the present study was to check the effect of trigonelline on the gut microbiota based conversion of TMA to TMAO. Trigonelline was isolated from hydroalcoholic extract of seeds of Trigonella foenum-graecum. The isolated trigonelline was characterized through TLC and UPLC-MS. Anaerobic microbe responsible for the metabolism of choline to TMA was isolated by culturing the human gut microbiota in choline enriched medium. The isolated bacteria was identified at molecular level based on PCR amplification of 1500bp of 16S rRNA gene sequence. Isolated FMO3 was used for ex vivo conversion of TMA to TMAO. Further, we investigated the effect of trigonelline in isolated gut microbe based metabolism of choline, lipid profile and TMAO levels in mice with or without suppression of gut microbiota with antibiotics. Liquid-liquid purification and chromatographic analysis confirmed the trigonelline purity (87.26%) and which was also confirmed by mass spectroscopy with m/z 137.4 in positive ionization mode. A total of 30 anaerobic microbes responsible for TMA production were isolated and Citrobacter freundii was the superior among others for the production of TMA. In vitro culture of C. freundii in choline enriched medium supplemented with trigonelline resulted in significantly reduction TMA and followed by TMAO production. In ex vivo, a maximum of 85.3% TMAO production was reduced by trigonelline at concentration of about 300 µg/mL. Serum level of lipids and TMAO were significantly altered in choline fed animals with or without suppression of gut microbiota and this phenomenon was reversed upon the oral administration of trigonelline in a dose-dependent manner. This study demonstrates the effect of trigonelline on gut microbiota responsible for choline metabolism and this can be used as a model for evaluation of herbal drugs and its effect in gut microbiota prompted cardiovascular disorders.

Attenuation of neurobehavioral and neurochemical abnormalities in animal model of cognitive deficits of Alzheimer's disease by fermented soybean nanonutraceutical.

The present study was performed to evaluate the efficacy of nanonutraceuticals (NN) for attenuation of neurobehavioral and neurochemical abnormalities in Alzheimer's disease. Solid-state fermentation of soybean with Bacillus subtilis was performed to produce different metabolites (nattokinase, daidzin, genistin and glycitin and menaquinone-7). Intoxication of rats with colchicine caused impairment in learning and memory which was demonstrated in neurobehavioral paradigms (Morris water maze and passive avoidance) linked with decreased activity of acetylcholinesterase (AChE). NN treatment led to a significant increase in TLT in the retention trials as compared to acquisition trial TLT suggesting an improved learning and memory in rats. Further, treatment of NN caused an increase in the activity of AChE (42%), accompanied with a reduced activity of glutathione (42%), superoxide dismutase (43%) and catalase (41%). It also decreased the level of lipid peroxidation (28%) and protein carbonyl contents (30%) in hippocampus as compared to those treated with colchicine alone, suggesting a possible neuroprotective efficacy of NN. Interestingly, in silico studies also demonstrated an effective amyloid-ß and BACE-1 inhibition activity. These findings clearly indicated that NN reversed colchicine-induced behavioral and neurochemical alterations through potent antioxidant activity and could possibly impart beneficial effects in cognitive defects associated with Alzheimer's disease.

Ultrasound-assisted extraction of gymnemic acids from Gymnema sylvestre leaves and its effect on insulin-producing RINm-5 F ß cell lines.

INTRODUCTION: Gymnema sylvestre is an important anti-diabetic medicinal plant, hence it is necessary to study the effective extraction of its active medicinal components. OBJECTIVE: To develop an efficient ultrasound-assisted extraction method for anti-diabetic gymnemic acids from Gymnema sylvestre leaves and measure their effect on insulin-producing RINm-5 F ß cells. METHODS: Box-Behnken's design and response surface methodology was applied to the ultrasound-assisted extraction of gymnemic acids from Gymnema sylvestre leaves. Analysis of gymnemic acids was carried out by high-performance thin-layer chromatography by converting total gymnemic acids into gymnemagenin by alkali hydrolysis. Effects of extracts on insulin production were tested on cultured, insulin-producing RINm-5 F ß cell lines. RESULTS: The point prediction tool of the design expert software predicted 397.9 mg gymnemic acids per gram of the defatted G. sylvestre leaves using ultrasound-assisted extraction, with ethanol at 60 °C for 30 min. The predicted condition shows 93.34% validity under experimental conditions. The ultrasound-assisted extract caused up to about four times more insulin production from RINm-5 F ß cells than extracts obtained from Soxhlet extraction. CONCLUSIONS: Response surface methodology was successfully used to improve the extraction of gymnemic acids from G. sylvestre leaves. The ultrasound-assisted extraction process may be a better alternative to prepare such herbal extracts because it saves time and may prevent excess degradation of the target analytes.

Antibacterial activity of resin rich plant extracts.

BACKGROUND: The in vitro antibacterial activity of resin rich methanolic extracts (RRMEs) of Commiphora myrrha, Operculina turpethum, and Pinus roxburghii. MATERIALS AND METHODS: Different concentration were studied by agar-well diffusion method against Gram-positive (Staphylococcus aureus, Bacillus subtilis, Micrococcus luteus, Enterococcus faecalis) and Gram-negative bacterial strains (Escherichia coli, Pseudomonas aeruginosa, Salmonella typhi, Shigella dysenteriae). RESULTS: Among all the bacterial strains tested, E. faecalis was most sensitive and S. typhi was resistant to C. myrrha and P. roxburghii. The extracts of O. turpethum were active against all tested strains in which B. subtilis and S. aureus were the most sensitive. CONCLUSION: This suggested that the antibacterial activity of RRMEs of O. turpethum was more than C. myrrha and P. roxburghii. This probably explains the potential of these plants against a number of infections caused by bacterial strains tested.

Five new secondary metabolites from Monascus purpureus-fermented Hordeum vulgare and Sorghum bicolor.

Long grains of Hordeum vulgare and Sorghum bicolor were individually fermented with Monascus purpureus MTCC 369 under solid state fermentation. The aqueous extract of Monascus which fermented H. vulgare and S. bicolor was found to contain five different new metabolites. Silica gel column chromatography of the aqueous extract with a linear gradient of ethyl acetate, acetonitrile and carbon tetrachloride (v/v) yielded five new metabolites named benzopranyl capriate (9H-1-isoprenyl-benzopyran-5-isopropanoic acid-6-ol-6-n-decanoate), shorghumoic acid (n-octadec-8,11-dien-7α-ol-1-oic acid) and sorghumflavin A (2-n-butyloxo-6-ß-hydroxy-7-ß-isoprenyl ankaflavin) from Monascus-fermented S. bicolor, while hordeumflavin B (2-n-undecanyloxo-7-ß-isoprenyl ankaflavin) and vulgaredilone (2-dodecanyl-7-ß isopranyl monoscodilone) from Monascus-fermented H. vulgare.

Production of angkak through co-culture of Monascus purpureus and Monascus ruber

Angkak (red mold rice, red yeast rice, Chinese red rice) is a traditional Chinese medicine produced by solid-state fermentation of cooked non-glutinous rice with Monascus species. The secondary metabolite of Monascus species, monacolin K /lovastatin, has been proven to lower blood lipid levels. In this study, a co-culture of Monascus purpureus MTCC 369 and Monascus ruber MTCC 1880 was used for angkak production. Four medium parameters screened by Plackett-Burman design were optimized by response surface methodology for highest lovastatin production in angkak during solid-state fermentation by the co-culture. Maximum lovastatin production of 2.84 mg g-1 was predicted in solid medium containing 20 g rice and 40 ml liquid nutrients medium (malt extract 9.68 g l-1, dextrose 38.90 g l-1, MnSO4.H2O 1.96 g l-1, and MgSO4.7H2O 0.730 g l-1) by point prediction tool of Design Expert 7.1 software (Statease Inc. USA).

Production of Angkak Through Co-Culture of Monascus Purpureus and MONASCUS RUBER.

Angkak (red mold rice, red yeast rice, Chinese red rice) is a traditional Chinese medicine produced by solid-state fermentation of cooked non-glutinous rice with Monascus species. The secondary metabolite of Monascus species, monacolin K /lovastatin, has been proven to lower blood lipid levels. In this study, a co-culture of Monascus purpureus MTCC 369 and Monascus ruber MTCC 1880 was used for angkak production. Four medium parameters screened by Plackett-Burman design were optimized by response surface methodology for highest lovastatin production in angkak during solid-state fermentation by the co-culture. Maximum lovastatin production of 2.84 mg g(-1) was predicted in solid medium containing 20 g rice and 40 ml liquid nutrients medium (malt extract 9.68 g l(-1), dextrose 38.90 g l(-1), MnSO4.H2O 1.96 g l(-1), and MgSO4.7H2O 0.730 g l(-1)) by point prediction tool of Design Expert 7.1 software (Statease Inc. USA).