Phenylalanine Butyramide: A Butyrate Derivative as a Novel Inhibitor of Tyrosinase.

Metabolites resulting from the bacterial fermentation of dietary fibers, such as short-chain fatty acids, especially butyrate, play important roles in maintaining gut health and regulating various biological effects in the skin. However, butyrate is underutilized due to its unpleasant odor. To circumvent this organoleptic unfavorable property, phenylalanine butyramide (PBA), a butyrate precursor, has been synthesized and is currently available on the market. We evaluated the inhibition of mushroom tyrosinase by butyrate and PBA through in vitro assays, finding IC50 values of 34.7 mM and 120.3 mM, respectively. Docking calculations using a homology model of human tyrosinase identified a putative binding mode of PBA into the catalytic site. The anti-aging and anti-spot efficacy of topical PBA was evaluated in a randomized, double-blind, parallel-arm, placebo-controlled clinical trial involving 43 women affected by photo-damage. The results of this study showed that PBA significantly improved skin conditions compared to the placebo and was well tolerated. Specifically, PBA demonstrated strong skin depigmenting activity on both UV and brown spots (UV: -12.7% and -9.9%, Bs: -20.8% and -17.7% after 15 and 30 days, respectively, p < 0.001). Moreover, PBA brightened and lightened the skin (ITA°: +12% and 13% after 15 and 30 days, respectively, p < 0.001). Finally, PBA significantly improved skin elasticity (Ua/Uf: +12.4% and +32.3% after 15 and 30 days, respectively, p < 0.001) and firmness (Uf: -3.2% and -14.9% after 15 and 30 days, respectively, p < 0.01).

Synthesis, Antiproliferative Activity and Molecular Docking Analysis of Both Enantiomerically Pure Decursin Derivatives as Anticancer Agents.

Using (S)-decursinol isolated from root of Angelica gigas Nakai (AGN), we semi-synthesized and evaluated a series of both enantiomerically pure decursin derivatives for their antiproliferative activities against A549 human lung cancer cells. All synthesized compounds showed a broad spectrum of inhibitory activities against the growth of A549 cells. Especially, compound (S)-2d with (E)-(furan-3-yl)acryloyl group showed the most potent activity (IC50: 14.03 µM) against A549 cancer cells as compared with the reference compound, decursin (IC50: 43.55 µM) and its enantiomer, (R)-2d (IC50: 151.59 µM). Western blotting assays indicated that (S)-2d more strongly inhibited Janus kinase 1 (JAK1) and signal transducer and activator of transcription activation 3 (STAT3) phosphorylation than decursin in a dose-dependent manner, while having no effect on CXCR7 overexpression and total STAT3 level. In addition, (S)-2d induced cell cycle arrest at G1 phase and subsequent apoptotic cell death in A549 cancer cells. Our combined analysis of molecular docking studies and biological data suggests that the inhibition of JAK1 with (S)-2d resulted in loss of STAT3 phosphorylation and inhibition of cell growth in A549 cancer cells. These overall results strongly suggest that (S)-2d (MRC-D-004) as a novel JAK1 inhibitor may have therapeutic potential in the treatment of A549 human lung cancers by targeting the JAK1/STAT3 signaling pathway.

A serine-conjugated butyrate prodrug with high oral bioavailability suppresses autoimmune arthritis and neuroinflammation in mice.

Butyrate-a metabolite produced by commensal bacteria-has been extensively studied for its immunomodulatory effects on immune cells, including regulatory T cells, macrophages and dendritic cells. However, the development of butyrate as a drug has been hindered by butyrate's poor oral bioavailability, owing to its rapid metabolism in the gut, its low potency (hence, necessitating high dosing), and its foul smell and taste. Here we report that the oral bioavailability of butyrate can be increased by esterifying it to serine, an amino acid transporter that aids the escape of the resulting odourless and tasteless prodrug (O-butyryl-L-serine, which we named SerBut) from the gut, enhancing its systemic uptake. In mice with collagen-antibody-induced arthritis (a model of rheumatoid arthritis) and with experimental autoimmune encephalomyelitis (a model of multiple sclerosis), we show that SerBut substantially ameliorated disease severity, modulated key immune cell populations systemically and in disease-associated tissues, and reduced inflammatory responses without compromising the global immune response to vaccination. SerBut may become a promising therapeutic for autoimmune and inflammatory diseases.

Combining visual cues and pheromone blends for monitoring and management of the tarnished plant bug Lygus lineolaris (Hemiptera: Miridae).

BACKGROUND: The tarnished plant bug Lygus lineolaris (Palisot de Beauvois) is considered the most damaging pest of cotton (Gossypium hirsutum L.) in the mid-southern United States. Previous studies have reported the role of different ratios of volatile metathoracic gland components such as hexyl butyrate, (E)-2-hexenyl butyrate and (E)-4-oxo-2-hexenal in eliciting low-level attraction of L. lineolaris. In this study, we tested different visual cues (colored sticky cards) in combination with olfactory cues (pheromone blends) to optimize the attraction and capture of L. lineolaris in the field. RESULTS: Red-colored sticky cards were more attractive to L. lineolaris adults than white, blue or yellow cards. Red sticky cards combined with blends of three potential pheromone components attracted significantly more L. lineolaris adults than sticky cards without a blend added. Traps baited with a blend of hexyl butyrate, (E)-2-hexenyl butyrate and (E)-4-oxo-2-hexenal in 4:10:7 ratio, respectively, caught a significantly higher number of L. lineolaris than those baited with 10:4:2 or 7:10:4 blends or an unbaited control in the first week of the experiment. CONCLUSIONS: Combining visual cues (red color) with olfactory cues (pheromone blends) significantly increased the capture of L. lineolaris in the field. This device or a future iteration could contribute towards sustainable and environmentally appropriate early-season monitoring and management of L. lineolaris in the field. © 2023 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry. This article has been contributed to by U.S. Government employees and their work is in the public domain in the USA.

Synthesis of Short-Chain Alkyl Butyrate through Esterification Reaction Using Immobilized Rhodococcus Cutinase and Analysis of Substrate Specificity through Molecular Docking.

Alkyl butyrate with fruity flavor is known as an important additive in the food industry. We synthesized various alkyl butyrates from various fatty alcohol and butyric acid using immobilized Rhodococcus cutinase (Rcut). Esterification reaction was performed in a non-aqueous system including heptane, isooctane, hexane, and cyclohexane. As a result of performing the alkyl butyrate synthesis reaction using alcohols of various chain lengths, it was found that the preference for the alcohol substrate had the following order: C6 > C4 > C8 > C10 > C2. Through molecular docking analysis, it was found that the greater the hydrophobicity of alcohol, the higher the accessibility to the active site of the enzyme. However, since the number of torsions increased as the chain length increased, it became difficult for the hydroxyl oxygen of the alcohol to access the γO of serine at the enzyme active site. These molecular docking results were consistent with substrate preference results of the Rcut enzyme. The Rcut maintained the synthesis efficiency at least for 5 days in isooctane solvent. We synthesized as much as 452 mM butyl butyrate by adding 100 mM substrate daily for 5 days and performing the reaction. These results show that Rcut is an efficient enzyme for producing alkyl butyrate used in the food industry.

Fungal mash enzymatic pretreatment combined with pH adjusting approach facilitates volatile fatty acids yield via a short-term anaerobic fermentation of food waste.

As an alternative for commercial enzyme, crude enzyme of fungal mash could promote food waste (FW) hydrolysis, but its specific effects coupled pH adjusting on the production of volatile fatty acids (VFAs) remains unknown. The crude enzyme produced from an Aspergillus awamori, named complex-amylase (CA), was added to short-term anaerobic system of FW fermentation. Results showed that adding CA significantly improved the solubility and degradability of biodegradable and non-biodegradable organics in FW, where the SCOD concentration with adding CA increased by 116.9% relative to the control but a marginal enhancement on VFAs yield. In contrast, adding CA combined with adjusting pH 8 markedly increased the VFAs production to 32.0 g COD/L, almost 10 times as much as the control. Besides, pH adjusting altered the metabolic pathway from lactate-type to butyrate-type. Adding CA coupled pH adjusting significant increase the component of butyrate compared with pH adjusting alone. Moreover, microbial community analysis indicated that adding CA reinforced proportion of the butyrate-producing bacteria (e.g., Dialister) under basic conditions, thus enhancing the butyrate metabolic pathways. This study demonstrated that fungal mash pretreatment coupled pH conditioning could be an economical way to enhance VFAs yield for FW valorization during anaerobic fermentation.

Roles of Fe-C amendment on sulfate-containing pharmaceutical wastewater anaerobic treatment: Microbial community and sulfur metabolism.

The effects of multiple two-phase anaerobic treatment involving acidification coupling Fe-C on sulfate-containing chemical synthesis-based pharmaceutical wastewater treatment were investigated. Fe-C was added as a filler with 25% vol. to acidogenic reactors for semi-continuous operation. The results suggested that Fe-C amendment promoted sulfate removal efficiency by 47.5% and shortened the reaction time by 50% in the acidogenic phase. With mitigation of sulfate inhibition, SCOD removal efficiency and methane production were further increased by 24.6% and 398% compared to direct raw wastewater anaerobic digestion, respectively, in methanogenic phase. The results of sulfate removal kinetics confirmed a 150% increase of removal rate in acidogenic phase. However, the apparent kinetic microbial sulfate removal constant without Fe-C amendment was maintained at approximately 0.06 h-1. The Fe-C amendment not only increased the relative abundance of Methanothrix and Desulfovibrio for sulfate reduction but also enriched unclassified_p__Chloroflexi and unclassified_c__Deltaproteobacteria for acidification. Metagenomic results indicated that Fe-C enhanced dissimilatory sulfate reduction and PAPS synthesis of assimilatory step. The hydrogen sulfide production through the 3-mercaptopyruvate to pyruvate pathways was also enhanced. Butyrate-oxidizing genes were increased synchronously to convert butyrate to acetate.

Strain-level fitness in the gut microbiome is an emergent property of glycans and a single metabolite.

The human gut microbiota resides within a diverse chemical environment challenging our ability to understand the forces shaping this ecosystem. Here, we reveal that fitness of the Bacteroidales, the dominant order of bacteria in the human gut, is an emergent property of glycans and one specific metabolite, butyrate. Distinct sugars serve as strain-variable fitness switches activating context-dependent inhibitory functions of butyrate. Differential fitness effects of butyrate within the Bacteroides are mediated by species-level variation in Acyl-CoA thioesterase activity and nucleotide polymorphisms regulating an Acyl-CoA transferase. Using in vivo multi-omic profiles, we demonstrate Bacteroides fitness in the human gut is associated together, but not independently, with Acyl-CoA transferase expression and butyrate. Our data reveal that each strain of the Bacteroides exists within a unique fitness landscape based on the interaction of chemical components unpredictable by the effect of each part alone mediated by flexibility in the core genome.

Tributyrin, a Butyrate Pro-Drug, Primes Satellite Cells for Differentiation by Altering the Epigenetic Landscape.

Satellite cells (SC) are a population of muscle resident stem cells that are responsible for postnatal muscle growth and repair. With investigation into the genomic regulation of SC fate, the role of the epigenome in governing SC myogenesis is becoming clearer. Histone deacetylase (HDAC) inhibitors have been demonstrated to be effective at enhancing the myogenic program of SC, but their role in altering the epigenetic landscape of SC remains undetermined. Our objective was to determine how an HDAC inhibitor, butyrate, promotes myogenic differentiation. SC from tributyrin treated neonatal piglets showed a decrease relative to SC from control animals in the expression of enhance of zeste homologue-2 (EZH2), a chromatin modifier, ex vivo. Chromatin Immunoprecipitation-Sequencing (ChIP-Seq) analysis of SC isolated from tributyrin treated pigs showed a global reduction of the tri-methylation of lysine 27 of histone H3 (H3K27me3) repressive chromatin mark. To determine if reductions in EZH2 was the primary mechanism through which butyrate affects SC behavior, SC were transfected with siRNA targeting EZH2, treated with 0.5 mM butyrate, or both. Treatment with butyrate reduced paired-box-7 (Pax7) and myogenic differentiation-1 (MyoD) gene expression, while siRNA caused reductions in EZH2 had no effect on their expression. EZH2 depletion did result in an increase in differentiating SC, but not in myotube hypertrophy. These results indicate that while EZH2 reduction may force myogenic differentiation, butyrate may operate through a parallel mechanism to enhance the myogenic program.

Butyrylated starch protects mice from DSS-induced colitis: combined effects of butyrate release and prebiotic supply.

Butyrate has recently emerged as a promising substance for the therapy of colitis. To overcome the shortcomings implicated in the existing delivery systems of butyrate, we utilized butyrylated starch to specifically deliver butyrate to the colon. Herein, we describe the stable loading of butyrate via chemical bonds with a heterogeneous distribution throughout the particle. Butyrylated starch supply increased butyrate as well as total short-chain fatty acid contents at the end of the intervention period. Moreover, butyrylated starch showed multiple effects on the suppression of DSS-induced colitis. From the observation of the gut-liver axis, reduced hepatic inflammation and hepatocyte damage further confirmed alleviated colonic inflammation. Given that butyrylated starch has the combined effects of specific release of butyrate in the colon and extra supply of fermentable substrates for gut microbiota, this work provides an effective strategy for the assistant therapy of colitis.

Decursinol Angelate Mitigates Sepsis Induced by Methicillin-Resistant Staphylococcus aureus Infection by Modulating the Inflammatory Responses of Macrophages.

The herbal plant Angelica gigas (A. gigas) has been used in traditional medicine in East Asian countries, and its chemical components are reported to have many pharmacological effects. In this study, we showed that a bioactive ingredient of A. gigas modulates the functional activity of macrophages and investigated its effect on inflammation using a sepsis model. Among 12 different compounds derived from A. gigas, decursinol angelate (DA) was identified as the most effective in suppressing the induction of TNF-α and IL-6 in murine macrophages. When mice were infected with a lethal dose of methicillin-resistant Staphylococcus aureus (MRSA), DA treatment improved the mortality and bacteremia, and attenuated the cytokine storm, which was associated with decreased CD38+ macrophage populations in the blood and liver. In vitro studies revealed that DA inhibited the functional activation of macrophages in the expression of pro-inflammatory mediators in response to microbial infection, while promoting the bacterial killing ability with an increased production of reactive oxygen species. Mechanistically, DA treatment attenuated the NF-κB and Akt signaling pathways. Intriguingly, ectopic expression of an active mutant of IKK2 released the inhibition of TNF-α production by the DA treatment, whereas the inhibition of Akt resulted in enhanced ROS production. Taken together, our experimental evidence demonstrated that DA modulates the functional activities of pro-inflammatory macrophages and that DA could be a potential therapeutic agent in the management of sepsis.

Structural Biology-Based Exploration of Subtype-Selective Agonists for Peroxisome Proliferator-Activated Receptors.

Progress in understanding peroxisome proliferator-activated receptor (PPAR) subtypes as nuclear receptors that have pleiotropic effects on biological responses has enabled the exploration of new subtype-selective PPAR ligands. Such ligands are useful chemical biology/pharmacological tools to investigate the functions of PPARs and are also candidate drugs for the treatment of PPAR-mediated diseases, such as metabolic syndrome, inflammation and cancer. This review summarizes our medicinal chemistry research of more than 20 years on the design, synthesis, and pharmacological evaluation of subtype-selective PPAR agonists, which has been based on two working hypotheses, the ligand superfamily concept and the helix 12 (H12) holding induction concept. X-ray crystallographic analyses of our agonists complexed with each PPAR subtype validate our working hypotheses.

On Demand Attachment and Detachment of rac-2-Br-DMNPA Tailoring to Facilitate Chemical Protein Synthesis.

Herein, we developed a bifunctional reagent rac-2-Br-DMNPA 2 for the late-stage protection of peptide cysteine. Through the identification of its t-Bu ester 1 as a more competent form under ligation conditions, facile N-terminal and side-chain caging for the model peptide and protein were accomplished. Building upon this, a one-pot ligation and photolysis strategy was applied in the synthesis of the mini-protein chlorotoxin. More importantly, we extended the utility of 2 as a bifunctional linker for traceless solid-phase chemical ligation.

Effects of Phytosterol Butyrate Ester on the Characteristics of Soybean Phosphatidylcholine Liposomes.

The nutritional and structural properties of phytosterols (PS)/phytosterol esters (PEs) facilitate their use as substitutes for cholesterol in liposome encapsulation systems designed for oral drugs and health products. The purpose of this study was to determine the effect of phytosterol butyrate ester (PBE) on the properties of liposomes. PBE was encapsulated within liposomes (approximately 60 nm) prepared using soybean phosphatidylcholine using the thin-film hydration method. There was no significant change in the average particle diameter and zeta potential of these liposomal vesicles corresponding to the increasing amounts of encapsulated PBE. The incorporation of PBE increased the polydispersity index (PDI) independent of concentration. Additionally, we observed that the storage stability of PBE liposomes with uniform particle size and approximately spherical shape vesicle was better at low concentration. The results of Fourier-transform infrared (FTIR) spectroscopy and Raman spectroscopy showed that PBE was positioned at the water interface, which increased the order of hydrophobic alkyl chains in the lipid membranes. The incorporation of PBE led to an increase in the trans conformation of hydrophobic alkyl chain and consequently, the thermal stability of liposomes, which was confirmed by differential scanning calorimetry (DSC). The results of powder X-ray diffraction (XRD) analysis confirmed that PBE was present in an amorphous form in the liposomes. Additionally, the incorporation of PBE reduced the micropolarity of the lipid membrane. Thus, when preparing liposomes using thin-film hydration, the presence of PBE affected the characteristics of liposomes.

Renewable fatty acid ester production in Clostridium.

Bioproduction of renewable chemicals is considered as an urgent solution for fossil energy crisis. However, despite tremendous efforts, it is still challenging to generate microbial strains that can produce target biochemical to high levels. Here, we report an example of biosynthesis of high-value and easy-recoverable derivatives built upon natural microbial pathways, leading to improvement in bioproduction efficiency. By leveraging pathways in solventogenic clostridia for co-producing acyl-CoAs, acids and alcohols as precursors, through rational screening for host strains and enzymes, systematic metabolic engineering-including elimination of putative prophages, we develop strains that can produce 20.3 g/L butyl acetate and 1.6 g/L butyl butyrate. Techno-economic analysis results suggest the economic competitiveness of our developed bioprocess. Our principles of selecting the most appropriate host for specific bioproduction and engineering microbial chassis to produce high-value and easy-separable end products may be applicable to other bioprocesses.

Structural Insights into Notum Covalent Inhibition.

The carboxylesterase Notum hydrolyzes a palmitoleate moiety from Wingless/Integrated(Wnt) ligands and deactivates Wnt signaling. Notum inhibitors can restore Wnt signaling which may be of therapeutic benefit for pathologies such as osteoporosis and Alzheimer's disease. We report the identification of a novel class of covalent Notum inhibitors, 4-(indolin-1-yl)-4-oxobutanoate esters. High-resolution crystal structures of the Notum inhibitor complexes reveal a common covalent adduct formed between the nucleophile serine-232 and hydrolyzed butyric esters. The covalent interaction in solution was confirmed by mass spectrometry analysis. Inhibitory potencies vary depending on the warheads used. Mechanistically, the resulting acyl-enzyme intermediate carbonyl atom is positioned at an unfavorable angle for the approach of the active site water, which, combined with strong hydrophobic interactions with the enzyme pocket residues, hinders the intermediate from being further processed and results in covalent inhibition. These insights into Notum catalytic inhibition may guide development of more potent Notum inhibitors.