Garlic Bioconverted by Bacillus subtilis Stimulates the Intestinal Immune System and Modulates Gut Microbiota Composition.

SCOPE: This study evaluates the potential of bioconverted garlic ferments (BGFs) to stimulate the intestinal immune system and modulate cecal microbiota composition. METHODS AND RESULTS: In vitro, BGF significantly enhances Peyer's patch (PP)-mediated bone marrow cell proliferation and increases the production of interferon-gamma (IFN-γ), granulocyte macrophage-colony stimulating factor (GM-CSF), interleukin (IL)-6, and immunoglobulin A (IgA) but not IL-4, IL-5, and immunoglobulin E (IgE). Oral administration of BGF to C3H/HeN mice for 4 weeks significantly increases the GM-CSF (42.1-45.8 pg mL-1) and IFN-γ (6.5-12.1 pg mL-1) levels in PP cells. BGF also significantly elevates the levels of tumor necrosis factor-alpha (TNF-α, 165.0-236.3 pg mg-1), GM-CSF (2.4-3.0 ng mg-1), and IFN-γ (1.5-3.2 ng mg-1) in the small intestinal fluid, and TNF-α (2.2-3.1 pg mL-1) and IFN-γ (10.3-0.21.5 pg mL-1) in the mouse serum. Cecal microbial analysis reveals that BGF increases Bacteroidota and Verrucomicrobiota and decreases Actinobacteria and Bacillota at the phylum level in mice. At the genus level, BGF significantly increases the abundance of Fusimonas (250 mg kg-1 BW-1 day-1), Bacteroides (125 and 250 mg kg-1 BW-1 day-1), and Akkermansia (125 mg kg-1 BW-1 day-1) and decreases that of Bifidobacterium (62.5 and 250 mg kg-1 BW-1 day-1) and Limosilactobacillus (125 and 250 mg kg-1 BW-1 day-1). CONCLUSION: This study provides the first evidence of BGF's ability to modulate the intestinal immune system and gut microbiota, supporting its potential as a novel functional material to enhance gut immunity.

Immunostimulatory effects of Bacillus subtilis-fermented garlic (Aglio): an in-depth in vitro and in vivo analysis.

This study evaluated the immunostimulatory potential of garlic fermented with Bacillus subtilis (Aglio) and identified the underlying mechanisms using in vitro and in vivo models. Aglio significantly enhanced macrophage activity, with increased TNF-α (9.3-46.6 fold), MCP-1 (5.3-41.4 fold), IL-6 (2.1-32.1 fold), and IL-12 (1.1-5.5 fold) secretion compared to those of the standard garlic extract. This macrophage-stimulatory activity was associated with MAPK (ERK, JNK, and p38) and NF-κB (IκBα and p65) signaling pathway activation. Aglio significantly increased splenocyte proliferation (1.8-2.9 fold) and TNF-α (32.5-96.6 fold), IFN-γ (26.6-362.3 fold), GM-CSF (2.1-3.9 fold), and IL-6 (10.3-11.6 fold) secretion. Gene expression analysis revealed Th1-related T-Bet upregulation and Th2- and Th17-related GATA3 and FOXP3 downregulation, indicating a Th1-mediated splenocyte activation mechanism. Oral administration of Aglio (125 and 250 mg kg-1) to BALB/c mice increased splenocyte proliferation (2.1-3.3 fold) and elevated splenic cytokine (TNF-α, 1.9-2.7 fold; GM-CSF, 2.2-2.3 fold; IL-6, 1.9 fold) and antibody (IgA, 1.4-1.8 fold; IgG, 1.0-1.7 fold) levels. Aglio administration also increased serum TNF-α (2.1-3.3 fold), IL-6 (1.0-1.1 fold), and IgG (1.6-1.9 fold) levels. Nutrient analysis indicated that Aglio lacked detectable carbohydrates and had negligible protein and polyphenol contents compared to standard garlic extract, suggesting complete biotransformation during fermentation. These findings demonstrate Aglio-mediated immune activation, highlighting its potential as a functional food or nutraceutical agent for immune enhancement.

Phytochemical Profile, Antioxidant, Anti-Atopic, and Anti-Inflammatory Activities of Filipendula glaberrima Nakai at Different Growth Stages.

Since atopic dermatitis is an inflammatory skin disease, natural remedies, such as Filipendula glaberrima Nakai (FG), with anti-inflammatory properties are possible promising therapeutic options. This study aimed to investigate the therapeutic potential of FG extracts at different growth stages. Seven compounds were isolated from the FG leaf extracts using open-column chromatography, and they were analyzed using HPLC. The extracts were further evaluated for their total polyphenol and flavonoid content (TPC and TFC). The in vitro antioxidant properties of the FG extracts were evaluated using radical scavenging assays, whereas their anti-inflammatory activities were assessed by evaluating their ability to inhibit the production of inflammation-associated biomarkers using the Griess assay and ELISA, respectively. The MTT assay was used to evaluate the viability and cytotoxicity of the FG extracts in keratinocyte cell lines. The results showed that the full-flowering stage exhibited the highest TPC, TFC, and antioxidant activities, thus suggesting a positive correlation between these properties. All FG extracts showed significant anti-inflammatory activity by inhibiting the production of pro-inflammatory biomarkers in lipopolysaccharide-stimulated macrophages. Additionally, the FG extracts suppressed the production of cytokines and chemokines in keratinocytes, indicating their anti-atopic potential. HPLC analysis revealed that the full-flowering stage had the highest content of all the analyzed phytochemicals (gallic acid, (+)-catechin, hyperin, miquelianin, astragalin, afzelin, and quercetin). These results suggest that the full-flowering stage of FG is the most promising source for therapeutic applications owing to its superior phytochemical profile and biological activities. This study highlights the potential of FG extracts, particularly in its full-flowering stage, as a natural therapeutic agent for the management of inflammation-related diseases, and it can also serve as a reference for further research on FG.

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.