Improvement of sleep duration and quality through GABAA receptor by whey protein hydrolysate containing DIQK as the active main compound.

This study characterized the sleep activity, sleep mechanism, and active peptides of whey protein hydrolysates selected through behavioral analysis of fruit-flies (Drosophila melanogaster). Sleep-inducing whey protein (WP) hydrolysate was selected through fruit fly behavior analysis, and sleep activity was measured using a pentobarbital model and electroencephalographic analysis. The mechanism of action was confirmed using a γ-aminobutyric acid (GABA) receptor antagonist, and the active peptide was identified using liquid chromatography-mass spectroscopy. Whey protein hydrolysate, prepared using Alcalase and Prozyme (WP-AP), increased sleep time in a dose-dependent manner. WP-AP significantly increased not only sleep time but also slow-wave sleep and showed an insomnia-alleviating effect in a caffeine-induced insomnia mouse model. In addition, the gene and protein expression levels of GABA sub-type A (GABAA) receptors increased in the brains of mice orally administered with WP-AP. Through peptide analysis, the mixture of DIQK, VPPF peptide, and GABA contained in WP-AP was estimated to exhibit sleep activity, and due to its high content, DIQK was speculated to be the main sleep -inducing ingredient. These results indicate that WP-AP has the potential to be used as a new ingredient to improve sleep quality.

Comprehensive workflow encompassing discovery, verification, and quantification of indicator peptide in snail mucin using LC-quadrupole Orbitrap high-resolution tandem mass spectrometry.

Peptidomics analysis was conducted using high-resolution tandem mass spectrometry (MS2) to determine the peptide profile of snail-derived mucin extract (SM). The study was also aimed to identify an indicator peptide and validate a quantification method for this peptide. The peptide profiling and identification were conducted using discovery-based peptidomics analysis employing data-dependent acquisition, whereas the selected peptides were verified and quantified using parallel reaction monitoring acquisition. Among the 16 identified peptides, the selected octapeptide (TEAPLNPK) was quantified via precursor ion ionization (m/z 435.2400), followed by quantification of the corresponding quantifier ion fragment (m/z 639.3824) using MS2. The quantification method was optimized and validated in terms of specificity, linearity, accuracy, precision, and limit of detection/quantification. The validated method accurately quantified the TEAPLNPK content in the SM as 7.5 ± 0.2 µg/g. Our study not only identifies an indicator peptide from SM but also introduces a novel validation method, involving precursor ion ionization and quantification of specific fragments. Our findings may serve as a comprehensive workflow for the monitoring, selection, and quantification of indicator peptides from diverse food resources.

Yeast-Hydrolysate-Derived 1-Methyl-1,2,3,4-tetrahydro-ß-carboline-3-carboxylic Acid Inhibits Fat Accumulation during Adipocyte Differentiation.

This study aimed to investigate the impact of yeast hydrolysate (YH) on lipogenesis, elucidate its mechanistic action, and identify the active compounds responsible for its anti-adipogenic effects. YH (2 mg/mL) significantly reduced Oil Red O-stained lipids. YH (2 mg/mL) also downregulated C/EBPß and upregulated KLF2, both of which are early adipogenic factors. Moreover, YH (2 mg/mL) decreased C/EBPα, PPARγ, FABP4, FAS, ACC, and HMGCR mRNA expression. Additionally, YH significantly downregulated SEBP1c and SREBP2 and their target genes, which govern fatty acid and cholesterol metabolism; however, 2 mg/mL YH had a greater suppressive effect on SREBP1c than on SREBP2. YH (2 mg/mL) also significantly reduced the mRNA level of G6PD and malic enzyme, which are enzymes that synthesize NADPH for lipid synthesis, compared with the control. Furthermore, 1-methyl-1,2,3,4-tetrahydro-ß-carboline-3-carboxylic acid (MTCA) was identified as the active compound with anti-adipogenic effects using solvent fractionation and chromatographic analysis of YH, and 1.1 µg/mL MTCA significantly downregulated SREBP1c/SREBP2 mRNAs by 47.8% and 69.2%, respectively, along with the target genes FAS, ACC, and HMGCR by 79.0%, 77.0%, and 40.9%, respectively. Collectively, YH effectively suppressed adipogenic lipid storage by downregulating SREBP- and NADPH-synthesizing genes. These findings suggest that YH containing MTCA has the potential to act as an anti-obesity agent.

Inhibition of P2X7R-NLRP3 Inflammasome Activation by Pleurotus citrinopileatus: A Possible Protective Role in Alcoholic Hepatosteatosis.

Pleurotus citrinopileatus (golden oyster mushroom) is a widely used edible mushroom. We investigated the inhibitory effect of P. citrinopileatus aqueous extract against alcoholic steatohepatitis and its underlying mechanism. Acute and chronic ethanol-feeding murine models were established by intragastrically administering ethanol or feeding an ethanol-containing Lieber-DeCarli liquid diet to male C57BL/6 mice. In both models, P. citrinopileatus decreased serum alanine aminotransferase (ALT), aspartate transaminase (AST), triglyceride (TG), and hepatic TG levels. Hematoxylin and eosin (HE) and Oil Red O staining confirmed that P. citrinopileatus ameliorated both acute and chronic alcoholic hepatosteatosis, characterized by regulation of lipid-metabolism-related proteins, including sirtuin 1 (SIRT1), AMP-activated kinase (AMPK), and sterol regulatory element-binding protein (SREBP1). P. citrinopileatus reversed inflammatory response via modulating purinergic receptor P2X ligand-gated ion channel 7 (P2X7R)-NOD-like receptor pyrin domain 3 (NLRP3) inflammasome. P. citrinopileatus restored the expressions of those proteins to a normal level. In addition, HepG2 cells were incubated with P. citrinopileatus prior to ethanol stimulation. P. citrinopileatus reduced ethanol exposure-induced lipid deposition. Concomitantly, P. citrinopileatus increased AMPK and SIRT1 expressions, which were reduced by ethanol treatment. P. citrinopileatus ameliorated alcoholic hepatic steatosis and accompanied inflammatory response via regulating SIRT1-AMPK and P2X7R-NLRP3 inflammasome activation, highlighting a promising strategy and utility of P. citrinopileatus for alcoholic steatohepatitis as dietary health supplements.