A consortium of Hordeum vulgare and gut microbiota against non-alcoholic fatty liver disease via data-driven analysis.

Despite many recent studies on non-alcoholic fatty liver disease (NAFLD) therapeutics, the optimal treatment has yet to be determined. In this unfinished project, we combined secondary metabolites (SMs) from the gut microbiota (GM) and Hordeum vulgare (HV) to investigate their combinatorial effects via network pharmacology (NP). Additionally, we analyzed GM or barley - signalling pathways - targets - metabolites (GBSTMs) in combinatorial perspectives (HV, and GM). A total of 31 key targets were analysed via a protein-protein interaction (PPI) network, and JUN was identified as the uppermost target in NAFLD. On a bubble plot, we revealed that apelin signalling pathway, which had the lowest enrichment factor antagonize NAFLD. Holistically, we scrutinized GBSTM to identify key components (GM, signalling pathways, targets, and metabolites) associated with the Apelin signalling pathway. Consequently, we found that the primary GMs (Eubacterium limosum, Eggerthella sp. SDG-2, Alistipes indistinctus YIT 12060, Odoribacter laneus YIT 12061, Paraprevotella clara YIT 11840, Paraprevotella xylaniphila YIT 11841) to ameliorate NAFLD. The molecular docking test (MDT) suggested that tryptanthrin-JUN is an agonist, conversely, dihydroglycitein-HDAC5, 1,3-diphenylpropan-2-ol-NOS1, and (10[(Acetyloxy)methyl]-9-anthryl)methyl acetate-NOS2, which are antagonistic conformers in the apelin signalling pathway. Overall, these results suggest that combination therapy could be an effective strategy for treating NAFLD.

Enzymatic Activity and Amino Acids Production of Predominant Fungi from Traditional Meju during Soybean Fermentation.

To investigate the effect of the predominant fungal species from Korean traditional meju and doenjang on soybean fermentation, the enzymatic activity and amino acid production of twenty-two fungal strains were assessed through solid- and liquid-state soybean fermentation. Enzymatic activity analyses of solid-state fermented soybeans revealed different enzyme activities involving protease, leucine aminopeptidase (LAP), carboxypeptidase (CaP), glutaminase, γ-glutamyl transferase (GGT), and amylase, depending on the fungal species. These enzymatic activities significantly affected the amino acid profile throughout liquid-state fermentation. Strains belonging to Mucoromycota, including Lichtheimia, Mucor, Rhizomucor, and Rhizopus, produced smaller amounts of total amino acids and umami-producing amino acids, such as glutamic acid and aspartic acid, than strains belonging to Aspergillus subgenus circumdati. The genera Penicillium and Scopulariopsis produced large amounts of total amino acids and glutamic acid, suggesting that these genera play an essential role in producing umami and kokumi tastes in fermented soybean products. Strains belonging to Aspergillus subgenus circumdati, including A. oryzae, showed the highest amino acid content, including glutamic acid, suggesting the potential benefits of A. oryzae as a starter for soybean fermentation. This study showed the potential of traditional meju strains as starters for soybean fermentation. However, further analysis of processes such as the production of G-peptide for kokumi taste and volatile compounds for flavor and safety is needed.

The seamless integration of dietary plant-derived natural flavonoids and gut microbiota may ameliorate non-alcoholic fatty liver disease: a network pharmacology analysis.

We comprised metabolites of gut microbiota (GM; endogenous species) and dietary plant-derived natural flavonoids (DPDNFs; exogenous species) were known as potent effectors against non-alcoholic fatty liver disease (NAFLD) via network pharmacology (NP). The crucial targets against NAFLD were identified via GM and DPDNFs. The protein interaction (PPI), bubble chart and networks of GM or natural products- metabolites-targets-key signalling (GNMTK) pathway were described via R Package. Furthermore, the molecular docking test (MDT) to verify the affinity was performed between metabolite(s) and target(s) on a key signalling pathway. On the networks of GNMTK, Enterococcus sp. 45, Escherichia sp.12, Escherichia sp.33 and Bacterium MRG-PMF-1 as key microbiota; flavonoid-rich products as key natural resources; luteolin and myricetin as key metabolites (or dietary flavonoids); AKT Serine/Threonine Kinase 1 (AKT1), CF Transmembrane conductance Regulator (CFTR) and PhosphoInositide-3-Kinase, Regulatory subunit 1 (PIK3R1) as key targets are promising components to treat NAFLD, by suppressing cyclic Adenosine MonoPhosphate (cAMP) signalling pathway. This study shows that components (microbiota, metabolites, targets and a key signalling pathway) and DPDNFs can exert combinatorial pharmacological effects against NAFLD. Overall, the integrated pharmacological approach sheds light on the relationships between GM and DPDNFs.

New insight into gut microbiota-derived metabolites to enhance liver regeneration via network pharmacology study.

We intended to identify favourable metabolite(s) and pharmacological mechanism(s) of gut microbiota (GM) for liver regeneration (LR) through network pharmacology. We utilized the gutMGene database to obtain metabolites of GM, and targets associated with metabolites as well as LR-related targets were identified using public databases. Furthermore, we performed a molecular docking assay on the active metabolite(s) and target(s) to verify the network pharmacological concept. We mined a total of 208 metabolites in the gutMGene database and selected 668 targets from the SEA (1,256 targets) and STP (947 targets) databases. Finally, 13 targets were identified between 61 targets and the gutMGene database (243 targets). Protein-protein interaction network analysis showed that AKT1 is a hub target correlated with 12 additional targets. In this study, we describe the potential microbe from the microbiota (E. coli), chemokine signalling pathway, AKT1 and myricetin that accelerate LR, providing scientific evidence for further clinical trials.

Elucidation of Prebiotics, Probiotics, Postbiotics, and Target from Gut Microbiota to Alleviate Obesity via Network Pharmacology Study.

The metabolites produced by the gut microbiota have been reported as crucial agents against obesity; however, their key targets have not been revealed completely in complex microbiome systems. Hence, the aim of this study was to decipher promising prebiotics, probiotics, postbiotics, and more importantly, key target(s) via a network pharmacology approach. First, we retrieved the metabolites related to gut microbes from the gutMGene database. Then, we performed a meta-analysis to identify metabolite-related targets via the similarity ensemble approach (SEA) and SwissTargetPrediction (STP), and obesity-related targets were identified by DisGeNET and OMIM databases. After selecting the overlapping targets, we adopted topological analysis to identify core targets against obesity. Furthermore, we employed the integrated networks to microbiota-substrate-metabolite-target (MSMT) via R Package. Finally, we performed a molecular docking test (MDT) to verify the binding affinity between metabolite(s) and target(s) with the Autodock 1.5.6 tool. Based on holistic viewpoints, we performed a filtering step to discover the core targets through topological analysis. Then, we implemented protein-protein interaction (PPI) networks with 342 overlapping target, another subnetwork was constructed with the top 30% degree centrality (DC), and the final core networks were obtained after screening the top 30% betweenness centrality (BC). The final core targets were IL6, AKT1, and ALB. We showed that the three core targets interacted with three other components via the MSMT network in alleviating obesity, i.e., four microbiota, two substrates, and six metabolites. The MDT confirmed that equol (postbiotics) converted from isoflavone (prebiotics) via Lactobacillus paracasei JS1 (probiotics) can bind the most stably on IL6 (target) compared with the other four metabolites (3-indolepropionic acid, trimethylamine oxide, butyrate, and acetate). In this study, we demonstrated that the promising substate (prebiotics), microbe (probiotics), metabolite (postbiotics), and target are suitable for obsesity treatment, providing a microbiome basis for further research.

Gut Microbiome in Non-Alcoholic Fatty Liver Disease: From Mechanisms to Therapeutic Role.

Non-alcoholic fatty liver disease (NAFLD) is considered to be a significant health threat globally, and has attracted growing concern in the research field of liver diseases. NAFLD comprises multifarious fatty degenerative disorders in the liver, including simple steatosis, steatohepatitis and fibrosis. The fundamental pathophysiology of NAFLD is complex and multifactor-driven. In addition to viruses, metabolic syndrome and alcohol, evidence has recently indicated that the microbiome is related to the development and progression of NAFLD. In this review, we summarize the possible microbiota-based therapeutic approaches and highlight the importance of establishing the diagnosis of NAFLD through the different spectra of the disease via the gut-liver axis.

Development of Safe and Flavor-Rich Doenjang (Korean Fermented Soybean Paste) Using Autochthonous Mixed Starters at the Pilot Plant Scale.

Doenjang (Korean fermented soybean paste) with an improved flavor and safety was prepared by the simultaneous fermentation of autochthonous mixed starters at the pilot plan scale. First, whole soybean meju was fermented by coculturing safety-verified starters Aspergillus oryzae MJS14 and Bacillus amyloliquefaciens zip6 or Bacillus subtilis D119C. These fermented whole soybean meju were aged in a brine solution after the additional inoculation of Tetragenococcus halophilus 7BDE22 and Zygosaccharomyces rouxii SMY045 to yield doenjang. Four doenjang batches prepared using a combination of mold, bacilli, lactic acid bacteria, and yeast starters were free of safety issues and had the general properties of traditional doenjang, a rich flavor and taste. All doenjang batches received a high consumer acceptability score, especially the ABsT and ABsTZ batches. This study suggests that flavor-rich doenjang similar to traditional doenjang can be manufactured safely and reproducibly in industry by mimicking the simultaneous fermentation of autochthonous mixed starters as in traditional doenjang fermentation. PRACTICAL APPLICATION: The development of a pilot plant process for doenjang fermentation using safety-verified autochthonous mixed starter will facilitate the manufacture of flavor-rich doenjang similar to traditional doenjang safely and reproducibly in industry.

Polyphenols bearing cinnamaldehyde scaffold showing cell growth inhibitory effects on the cisplatin-resistant A2780/Cis ovarian cancer cells.

Ovarian carcinoma remains the most lethal among gynecological cancers. Chemoresistance is a clinical problem that severely limits treatment success. To identify potent anticancer agents against the cisplatin-resistant human ovarian cancer cell line A2780/Cis, 26 polyphenols bearing a cinnamaldehyde scaffold were synthesized. Structural differences in their inhibitory effect on clonogenicity of A2780/Cis cells were elucidated using comparative molecular field analysis and comparative molecular similarity indices analysis. Structural conditions required for increased inhibitory activity can be derived based on the analysis of their contour maps. The two most active compounds (16 and 19) were selected and further characterized their biological activities. We found that compounds 16 and 19 trigger cell cycle arrest at the G2/M phase and apoptotic cell death in cisplatin-resistant A2780/Cis human ovarian cancer cells. The molecular mechanism of compound 16 was elucidated using in vitro aurora A kinase assay, and the binding mode between the compound 16 and aurora A kinase was interpreted using in silico docking experiments. The findings obtained here may help us develop novel plant-derived polyphenols used for potent chemotherapeutic agents. In conclusion, compounds 16 and 19 could be used as promising lead compounds for the development of novel anticancer therapies in the treatment of cisplatin-resistant ovarian cancers.

Isoflavones inhibit the clonogenicity of human colon cancer cells.

Isoflavones are a class of polyphenols that contain various substituents such as hydroxy, methoxy, and glycosyl groups. Methoxy groups are known to increase cell permeability and stability, but small structural changes can result in large differences in biological activity. In this study, the anticancer activities of several methoxy isoflavones were tested using a clonogenic survival assay. The relationship between structural properties of methoxy isoflavones and their anticancer activities on HCT116 colon cancer cell lines were studied quantitatively using comparative molecular field analysis and comparative molecular similarity indices analysis. The purpose of this study was to identify structural changes in isoflavones that increase the inhibitory effect on HCT116 colon cancer cell clonogenicity.

Synthesis of methoxybenzoflavones and assignments of their NMR data.

A phytotoxic root exudate from Acroptilon repens was identified as 7,8-benzoflavone, an inhibitor of cytochrome P450 1A2 and activator of cytochrome P450 3A4. The synthetic 5,6-benzoflavone also is a potent phytotoxin. Six 7,8-benzoflavones and eight 5,6-benzoflavones were synthesized in this study. The NMR data for a few of these compounds have been previously reported; however, the NMR data for most of them have not been reported. For reference purposes, the complete NMR data for the 14 benzoflavones are described.

Flavanones inhibit the clonogenicity of HCT116 cololectal cancer cells.

Naringenin has been shown to display various biological effects such as antioxidant, anticancer, anti-inflammatory, and antiviral activities. Taxifolin inhibits the production of lipopolysaccharide-induced prostaglandin E, and fustin suppresses the activity of acetylcholinesterase. They all belong to flavanone which is a class of flavonoids with a C6-C3-C6 skeleton. Since the anticancer activities of flavanone derivatives have rarely been reported, we examined the effects of 26 flavanone derivatives on HCT116 colorectal cancer cells. Our results suggest that flavanone derivatives control the expression of cell cycle regulatory proteins, which blocks G1 cell cycle progression and inhibits the clonogenicity of HCT116 cells. In addition, in order to design flavanone derivatives that show better anticancer activity, structure-activity relationships were examined.

In silico study of the ion channel formed by tolaasin I produced by Pseudomonas tolaasii.

A toxin produced by Pseudomonas tolaasii, tolaasin, causes brown blotch disease in mushrooms. Tolaasin forms pores on the cellular membrane and destroys cell structure. Inhibiting the ability of tolaasin to form ion channels may be an effective method to protect against attack by tolaasin. However, it is first necessary to elucidate the three-dimensional structure of the ion channels formed by tolaasin. In this study, the structure of the tolaasin ion channel was determined in silico based on data obtained from nuclear magnetic resonance experiments.

Identification and structure elucidation of a p-phenoxybenzaldehyde in bamboo shoots by HPLC-ESI/MS/MS and NMR.

In this study, a derivative of p-phenoxybenzaldehyde in bamboo shoots was investigated. Bamboo shoots were ground and extracted with water, and an aqueous suspension was purified by SPE using Oasis HLB cartridges. After the SPE procedure, the analytes were analyzed by HPLC with refractive index detection (HPLC-RI). In the HPLC-RI analysis for sucralose, a putative sucralose was detected. In the subsequent HPLC-PDA analysis, the suspicious peak showed a unique UV absorption spectrum with the maximum wavelength at 285 nm indicating the existence of an aromatic ring. The contents of the unknown compound in bamboo shoot products ranged from 0.01 to 0.15 mg/g. The identity of the unknown compound was further confirmed by HPLC-ESI/MS/MS. The molecular weight of the unknown compound was determined to be 244. The chemical structure of the unknown compound was elucidated on the basis of NMR spectroscopic analyses ((1)H, (13)C, DEPT, COSY, HMQC, and HMBC). Finally, the structure of the unknown compound was characterized as 4-(4-dihydroxymethylphenoxy)benzaldehyde.

Relationship between the structures of flavonoids and their NF-κB-dependent transcriptional activities.

It has been previously shown that some flavonoids inhibit NF-κB; however, the structure-activity relationships between chalcone, flavanone, flavone, and isoflavone derivatives and their TNFα induced NF-κB inhibitory effects on HCT116 human colon cancer cells have not yet been reported. Therefore, in this study, the effects of flavonoid structure on inhibition of NF-κB were investigated. Based on the combined results of this study, the structure of the flavonoids was shown to affect NF-κB activation.

A compound isolated from Schisandra chinensis induces apoptosis.

Schizandra chinensis has been known to have five predominant tastes: salty, sweet, sour, astringent, and bitter. It has also been shown to have various effects on the cardiovascular system, gastrointestinal system, anti-inflammatory, central nervous system, endocrine system, and stress protect. However, its anti-cancer activity on colon carcinoma HCT-116 cells has not been yet been examined. Thus, in this study, we attempted to isolate a compound from Schisandra chinensis that induced apoptosis in HCT-116 cells. An active compound was found and identified to be Gomisin A. It displayed apoptotic activity through caspase-7 cleavage in colon carcinoma HCT-116 cells. In addition, we further assessed the effects of this compound using long-term survival clonogenic assay with HCT116 cells.

3'-Chloro-5,7-dimethoxyisoflavone inhibits TNFα-induced CXCL10 gene transcription by suppressing the NF-κB pathway in HCT116 human colon cancer cells.

Tumor necrosis factor α (TNFα) is a major inflammatory cytokine that plays important roles in progression of tumorigenesis in the tumor microenvironment. CXC chemokine ligand 10 (CXCL10), expression of which is stimulated by TNFα, is involved in tumor migration, invasion, and metastasis. 3'-Chloro-5,7-dimethoxyisoflavone (CDMF) is a synthetic isoflavone derivative. Here, we found that CDMF inhibits TNFα-induced invasive motility of human colon cancer cells. We tested whether CDMF would inhibit TNFα-induced CXCL10 expression using reverse transcription-PCR, quantitative real-time PCR, and enzyme-linked immunosorbent assay in HCT116 cells. CXCL10 expression, stimulated by TNFα, was suppressed by CDMF. The transcription factor nuclear factor-κB (NF-κB) is involved in TNFα-induced transcriptional activation of the CXCL10 gene promoter. Point mutation of the NF-κB binding site abolished TNFα-induced CXCL10 promoter activity. We next examined the effect of CDMF on TNFα-induced NF-κB activity. CDMF strongly inhibited both TNFα-induced IκB phosphorylation on Ser-32 and p65/RelA phosphorylation on Ser-536. Additionally, CDMF almost blocked TNFα-induced NF-κB-dependent transcriptional activity, as demonstrated by a NF-κB cis-acting reporter assay. Overall, our results indicate that CDMF suppresses production of CXCL10, by TNFα, through inhibition of NF-κB in HCT116 cells. We propose that CDMF may have beneficial effects in reducing TNFα-induced inflammatory responses, which are essential for tumor development in the colorectal tumor microenvironment.

5-Methoxyflavanone induces cell cycle arrest at the G2/M phase, apoptosis and autophagy in HCT116 human colon cancer cells.

Natural flavonoids have diverse pharmacological activities, including anti-oxidative, anti-inflammatory, and anti-cancer activities. In this study, we investigated the molecular mechanism underlying the action of 5-methoxyflavanone (5-MF) which has a strong bioavailability and metabolic stability. Our results show that 5-MF inhibited the growth and clonogenicity of HCT116 human colon cancer cells, and that it activated DNA damage responses, as revealed by the accumulation of p53 and the phosphorylation of DNA damage-sensitive proteins, including ataxia-telangiectasia mutated (ATM) at Ser1981, checkpoint kinase 2 (Chk2) at Thr68, and histone H2AX at Ser139. 5-MF-induced DNA damage was confirmed in a comet tail assay. We also found that 5-MF increased the cleavage of caspase-2 and -7, leading to the induction of apoptosis. Pretreatment with the ATM inhibitor KU55933 enhanced 5-MF-induced γ-H2AX formation and caspase-7 cleavage. HCT116 cells lacking p53 (p53(-/-)) or p21 (p21(-/-)) exhibited increased sensitivity to 5-MF compared to wild-type cells. 5-MF further induced autophagy via an ERK signaling pathway. Blockage of autophagy with the MEK inhibitor U0126 potentiated 5-MF-induced γ-H2AX formation and caspase-2 activation. These results suggest that a caspase-2 cascade mediates 5-MF-induced anti-tumor activity, while an ATM/Chk2/p53/p21 checkpoint pathway and ERK-mediated autophagy act as a survival program to block caspase-2-mediated apoptosis induced by 5-MF.

Complete NMR data of methoxylated cis- and trans-stilbenes as well as 1,2-diphenylethanes.

Resveratrol is a polyphenol isolated from many natural sources including grapes, mulberries, eucalyptus, spruce, lilies, and peanuts. The hydroxyl groups in polyphenols can be substituted with various functional groups, allowing production of multiple derivatives. NMR spectroscopy is used to identify new derivatives. Since the complete NMR data of the known derivatives can be useful for identification of the newly isolated derivatives, here, we report the synthesis of 14 methoxylated stilbenes and four 1,2-diphenylethanes and their NMR data.

Synthesis and complete assignment of NMR data of 20 chalcones.

Chalcones, intermediates in flavonoid biosynthesis, can exhibit antibacterial, antiproliferative, and anti-inflammatory properties. Chalcones contain two benzene rings and both hydroxylated and methoxylated analogs are frequently produced by hydroxylases and O-methyltransferases in plant biosynthetic pathways. Assignments of NMR peaks in the spectra of hydroxylated and/or methoxylated chalcones can help in identifying novel chalcone derivatives isolated from natural sources by referencing these data against NMR spectra obtained from known chalcones. We report here the syntheses of 20 chalcones and complete assignments of (1)H and (13)C NMR spectra.

Relationships between structures of hydroxyflavones and their antioxidative effects.

Even hydroxyflavones show diverse biological functions, they have two common features such as showing antioxidative effects and containing hydroxyl groups. The authors tested the antioxidative effects of thirty hydroxyflavones using 1,1-diphenyl-2-picrylhydrazyl radical scavenging assay. While the scavenging activity of galangin, 3,5,7-trihydroxyflavone was 52.5%, fisetin, 3,7,3',4'-tetrahydroxyflavone showed 85.2%. To investigate the relationships between the structures of hydroxyflavones and their antioxidative effects, the three-dimensional quantitative structure-activity relationships were examined.