Qiang He

To alleviate the structural and nutritional deterioration of oxidized rice bran protein (RBP), this study proposes an oxidation degree-guided polyphenol matching strategy for improving the RBP structure and digestion behavior. Endogenous rice bran (RB) polyphenols were conjugated to RBP with different oxidation degrees under ultrasonic treatment to obtain RBP-polyphenol (RBP-PP) complexes. Caffeic acid (CA), gallic acid (GA), and chlorogenic acid (CGA) were identified as the optimal polyphenols for native, moderately oxidized, and excessively oxidized RBP, respectively. Optimal RBP-PP complexes exhibit enhanced structural stability, reduced protein oxidation markers, stronger antioxidant capacity, and higher digestibility. Molecular simulations reveal oxidation degree-dependent binding modes and conformational rearrangements. Peptidomic and metabolomic analyses show that polyphenol binding changes digestion behavior, increasing low-molecular-weight peptides and generating oxidation degree-dependent metabolites. This study establishes an oxidation degree-guided precision polyphenol matching strategy for the targeted regulation of oxidized RBP, providing a theoretical basis for the high-value utilization of oxidized RBP.

Fluorescence in situ hybridization (FISH) is important for bacterial genotypic analysis; however, the limited number of labels per probe results in insufficient signal gain, compromising the detection of low-abundance RNAs. Here, we present high-density protein FISH (HD-FISH) for high-gain bacterial RNA imaging. Instead of fluorophore-modified probes, this method employs a fluorescently tagged, catalytically inactive ribonuclease H1 for multi-site labeling of target RNAs. Compared to traditional FISH and hybridization chain reaction-based FISH, HD-FISH suppresses background noise while enhancing positive signals, achieving 7.43-fold and 1.89-fold increases in signal gain, respectively, thereby enabling high-gain imaging. We showed that HD-FISH can precisely quantify quinolone-resistant Salmonella enterica strains within complex microbial communities, with a recovery rate of 97.78−131.40%, and allows us to detect and identify quinolone-resistant bacteria in fecal samples. The method provides a robust tool for profiling bacterial genotypes at the single-cell level, facilitating the elucidation of microbial heterogeneity and its roles in complex populations.

Sichuan-style black soybean soy sauce is a traditional fermented condiment renowned for its complex and regionally distinctive flavor profile. This study systematically investigated the physicochemical properties, flavor compounds, and microbial succession during six months of natural fermentation to elucidate the mechanisms underlying its unique flavor formation. Results showed that the amino acid nitrogen level increased to a peak of 1.37 g/100 mL before stabilizing at 1.01 g/100 mL, accompanied by a continuous rise in total acidity (0.69–2.75 g/100 mL). A total of 132 volatile compounds were identified, with esters (e.g., hexanoic acid, methyl ester, hexadecanoic acid, and methyl ester), alcohols (e.g., (E)-2-hepten-1-ol and trans-2-undecen-1-ol), and aldehydes (e.g., benzaldehyde and benzeneacetaldehyde) serving as key differentiating components. Nine taste-active (TAV ≥ 1) and 22 odor-active (ROAV ≥ 1) compounds were recognized as major flavor determinants, among which methional (ROAV = 4.77–119.05), 1-octen-3-ol (ROAV = 40.68–149.35), and 4-ethyl-2-methoxyphenol (ROAV = 4.70–36.26) were dominant contributors imparting sauce-like, mushroom-like, and smoky-clove notes, respectively. Microbial succession revealed a transition from Weissella and Aspergillus dominance in the early stage to salt-tolerant Tetragenococcus and aroma-producing yeasts (Kodamaea and Zygosaccharomyces) in later phases. Beyond organic acids and fermentation parameters (e.g., pH and salinity), microbial interactions were identified as critical drivers shaping community assembly and succession. Metabolic pathway analysis revealed a stage-dependent mechanism of flavor formation. During the initial stage (0–2 months), Aspergillus-mediated proteolysis released free amino acids as key taste precursors. In the later stages (3–6 months), Tetragenococcus and aroma-producing yeasts dominated, synthesizing characteristic esters (e.g., benzoic acid and methyl ester, correlated with Tetragenococcus; r = 0.71, p < 0.05), phenolics (e.g., 4-ethyl-2-methoxyphenol, correlated with Wickerhamomyces; r = 0.89, p < 0.05), and sulfur-containing compounds (e.g., methional, correlated with Wickerhamomyces; r = 0.83, p < 0.05).

To comprehensively elucidate the structure-activity relationship of oxidation on rice bran protein (RBP) nutrition, a hepatic metabolism model was established to evaluate the cellular impacts of RBP digestion products (RBPDP). Chromatography and multivariate analyses revealed oxidation-dependent changes in RBPDP profiles, marked by the loss of native protective peptides and formation of enzyme-resistant aggregates, identified as nutritional deterioration and oxidation markers, respectively. Compared to native RBP, digestion products from oxidized RBP elevated oxidation products and suppressed antioxidant capacity despite Nrf2 activation while increasing pro-inflammatory cytokines via the MyD88/NF-κB pathway. The NF-κB inhibitor pyrrolidine dithiocarbamate partially attenuated damage, though its efficacy diminished with increasing RBP oxidation. Results demonstrated that oxidation-induced protective peptide depletion and aggregate formation synergistically drove oxidative damage and an inflammatory response via activated Keap1-Nrf2/ARE and MyD88/NF-κB pathways. Critically, Nrf2/NF-κB crosstalk served as a key regulatory node, providing molecular targets to optimize the nutrition of RBP-based functional foods under oxidative stress.