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Freeze-drying is the most commonly used method for preserving probiotics. The freeze tolerance of probiotics has a significant impact on both their survival rate and the expression of their functional properties. To enhance the freeze tolerance of probiotics, this study established an adaptive evolution protocol combining cold stress with repeated freeze-thaw cycles to screen for freeze-thaw-tolerant evolved strains of MP108. The safety, metabolic, and functional characteristics of these strains were then evaluated. The results showed that the combination of the 8 h cold stress treatment at 4 °C and nine cycles of freezing and thawing at -20 °C effectively enhanced the strain's freeze tolerance, and the evolved strain L134 was successfully screened through adaptive evolution. Its freeze-dried survival rate and storage survival rate after 6 months of storage were both significantly higher than those of the parental strain ( < 0.05). Furthermore, it exhibited good passage stability. At the same time, the safety and acid-producing characteristics of L134 did not show significant changes compared to the parental strain. Furthermore, its tolerance to simulated gastric fluid, antibacterial activity, and antioxidant capacity were significantly enhanced ( < 0.05). In particular, compared to MP108, L134 exhibited significantly increased hydroxyl radical scavenging capacity as well as higher activities of the antioxidant enzymes SOD and CAT ( < 0.05); the improvement in its freeze tolerance may be related to this enhanced antioxidant capacity.

Organic acids are natural antimicrobial compounds commonly used in the food industry. In this study, acetic, lactic, butyric, citric, and malic acid at minimum inhibitory concentrations and their combinations at optimal inhibition concentrations were used to treat E. coli, and the effects on the cell barrier and biofilm of E. coli were evaluated. Acetic acid showed the highest membrane-damaging effect, while citric acid and malic acid could specifically damage the cell wall of E. coli, leading to alkaline phosphatase leakage. The RT-qPCR results showed that organic acids upregulated the membrane-protein-related genes of E. coli, and the combination of organic acids had a wider range of effects than single organic acid treatment. Moreover, organic acids inhibited the formation of E. coli biofilm and cellular activity within the biofilm. This study showed that the combination of organic acids plays a synergistic inhibitory role mainly through multiple destructive effects on the cell barrier and exhibited synergistic anti-biofilm effects. The three–three combination of acetic, lactic acid, and a third organic acid (butyric, citric, or malic) can play a better synergistic antibacterial effect than the two-pair combination of acetic and lactic acid. These findings have implications for the usage, development, and optimization of organic acid combinations.

Background Oro-gastrointestinal stress in the digestive tract is the main stress to which orally administered probiotics are exposed. The regulation of oro-gastrointestinal transit (OGT) stress on the adhesion and survival of probiotics under continuous exposure to simulated salivary-gastric juice-intestinal juice was researched in this study. Results Lactobacillus plantarum S7 had a higher survival rate after exposure to simulated OGT1 (containing 0.15% bile salt) stress and OGT2 (containing 0.30% bile salt) stress. The adhesion ability of L . plantarum S7 was significantly increased by OGT1 stress ( P  < 0.05) but was not changed significantly by OGT2 stress ( P  > 0.05), and this trend was also observed in terms of the thickness of the surface material of L . plantarum S7 cells. The expression of surface proteins of L . plantarum S7, such as the 30 S ribosomal proteins, mucus-binding protein and S-layer protein, was significantly downregulated by OGT stress ( P  < 0.05); meanwhile, the expression of moonlight proteins, such as glyceraldehyde-3-phosphate dehydrogenase (GAPDH), phosphoglycorate kinase (PGK), beta-phosphoglucomutase (PGM1), GroEL and glucose-6-phosphate isomerase (PGI), was significantly upregulated ( P  < 0.05). However, the upregulation of GAPDH, PGK, PGM1 and PGI mediated by OGT1 stress was greater than those mediated by OGT2 stress. The quorum sensing pathway of L . plantarum S7 was changed significantly by OGT stress compared with no OGT stress cells ( P  < 0.05), and the expression of Luxs in the pathway was significantly upregulated by OGT1 stress ( P  < 0.05). The ABC transportation pathway was significantly altered by OGT1 stress ( P  < 0.05), of which the expression of the peptide ABC transporter substrate-binding protein and energy-coupling factor transporter ATP-binding protein EcfA was significantly upregulated by OGT stress ( P  < 0.05). The glycolide metabolism pathway was significantly altered by OGT1 stress compared with that in response to OGT2 stress ( P  < 0.05). Conclusion L . plantarum S7 had a strong ability to resist OGT stress, which was regulated by the proteins and pathways related to OGT stress. The adhesion ability of L . plantarum S7 was enhanced after continuous exposure to OGT1 stress, making it a potential probiotic with a promising future for application.

This study explored the effects of two mixed fermentation methods: one was fermenting a soymilk and milk mixture by a lactic acid bacteria fermenting agent at 0.1 g/kg and 42 °C until the acidity was 70 °T, which was set as the MFSM method, and the other was fermenting milk alone by lactic acid bacteria at 42 °C for 12 h, placing it in a 4 °C refrigerator after acidification for 24 h and then mixing it with soymilk at a 1.5:1 ratio and storing the mixture at 4 °C, which was set as the SMFSM method. The quality and flavor of the soymilk and milk mixture products were investigated on the 0th, 15th and 30th days during storage. The changes in acidity, pH, number of viable bacteria, viscosity, water-holding capacity, texture, rheological properties, sensory quality and volatile flavors were determined. The results showed that compared with the fermented soymilk and milk mixtures under the MFSM method, the samples of fermented soymilk and milk mixtures under the SMFSM method showed a significant slowdown of acidification during storage, so that the sensory quality of the products was almost unaffected by acidity on the 30th day of storage. Furthermore, the number of viable bacteria was greater than 7 log cfu/mL. The water holding capacity did not change significantly until the 30th day. There was also no whey precipitation, indicating good stability. The samples in SMFSM mode had higher aromatic contents and beans during storage than the fermented soymilk and milk mixtures in MFSM mode. The rich variety of volatile flavors and the presence of acetoin, 2-heptanone, and (E,E)-3,5-octadien-2-one throughout the storage period allowed the samples to maintain a good sensory flavor during storage.

A high degree of hydrolysis can reduce the allergenicity of milk, while lactic acid bacteria (LAB) fermentation can further enhance the antioxidant ability of enzymatic milk. LAB with a strong antioxidant ability was screened, and the effects of LAB on the bitterness, taste and flavor of enzymatic skim milk (ESM) with a high degree of hydrolysis were investigated in this paper, in addition to the response surface methodology optimized the conditions of the LAB fermentation of ESM. The results indicate that the skim milk hydrolyzed by Protamex has a higher degree of hydrolysis and lower bitterness. The scavenging rate of 2,2-Diphenyl-1-picrylhydrazyl (DPPH) free radical, the inhibition rate of hydroxyl radical (·OH) and the superoxide dismutase (SOD) activity of Lactiplantibacillus plantarum 16 and Lactococcus lactis subsp. lactis m16 are significantly higher than those of other strains (p < 0.05), while the improvement effect of L. plantarum 16 on the bitterness and flavor of ESM is better than that of L. lactis subsp. lactis m16. The fermented ESM has a strong antioxidant ability and low bitterness when the inoculum quantity of L. plantarum 16 is 5%, fermentation at 37 °C for 18 h and the pH of the ESM is 6.5, for which the DPPH free radical scavenging rate is 61.32%, the ·OH inhibition rate is 83.35%, the SOD activity rate is 14.58 and the sensory evaluation is 4.25. The contents of amino acids related to bitterness and antioxidants were reduced and increased, respectively. The ESM fermented by L. plantarum 16 has a good flavor, antioxidant ability and low bitterness.

Alcoholic liver injury is a serious risk to human health. Probiotics have become a popular form of treatment. Lacticaseibacillus casei Grx12 and Limosilactobacillus fermentum Grx07 isolated from the gut of long-lived people in Rugao, Jiangsu, were studied to determine their protective effects and possible mechanisms of action on alcoholic liver injury. The results showed that rat serum ALT and AST were restored, and liver injury was reduced after the probiotics intervention. The level of antioxidant enzymes and antioxidants such as SOD, GSH and GSH-Px in the rat liver was significantly increased (p < 0.05), which reduces the level of MDA, a peroxidation product in the liver, and thus alleviates liver oxidative stress. L. casei Grx12 and L. fermentum Grx07 also could significantly enhance the expression of Nrf2 protein in the rat liver to regulate the anti-oxidative stress response in the body and cells (p < 0.05). The levels of ADH, Na+-K+-ATPase and Ca2+-ATPase in the rat liver were significantly increased (p < 0.05), which enhanced the body’s metabolism of alcohol. The rat serum LPS and liver TNF-α, IL-6, VEGF, TGF-β1 and NF-κB levels were significantly reduced (p < 0.05), indicating that the probiotics could relieve liver inflammation. The results of this study indicate that L. casei Grx12 and L. fermentum Grx07 have certain protective effects on alcoholic liver injury in rats, likely because of their antioxidant properties and ability to prevent oxidative stress and relieve inflammation.

Obesity is a common global problem. There are many fat-reducing herbal prescriptions in traditional Chinese medicine that have been proven to be safe and functional during long-term application. Microbial fermentation can improve the efficacy of herbal medicine and improve the unsavory flavor. In this study, Shenheling extract (SHLE) composed of six medicine food homology materials was used as the research object. The purpose of this study was to evaluate the effects of Lactobacillusfermentum grx08 fermentation on the antiobesity efficacy and flavor of SHLE. We found that L. fermentum grx08 grew well in SHLE. After 72 h of fermentation, the total polysaccharides, total flavonoids, total polyphenols and total saponins of SHLE decreased, but the lipase inhibitory activity and total antioxidant capacity (FRAP) were significantly increased (p < 0.01). There were no significant differences in the α-glucosidase inhibition rate and DPPH· clearance rate before or after fermentation (p > 0.05). In addition, the fermentation reduces the unpleasant flavors of SHLE such as bitterness and grassy and cassia flavors. This study demonstrates that SHLE fermented by L. fermentum grx08 improved some anti-obesity functions and improved the unpleasant flavor.

Adhesion to the intestinal tract provides the foundation for Lactobacillus to exert its benefits. Vacuum freeze-drying (VFD) is currently one of the main processing methods for Lactobacillus products. Therefore, the effects of VFD on the adhesion and survival of Lactiplantibacillus plantarum 67 were investigated in this study. The results show that L. plantarum 67 exhibits remarkable tolerance following successive exposure to simulated saliva, gastric juice and intestinal juice, and also has a strong adhesion ability to Caco-2 cells. The adhesion and survival rates of L. plantarum 67 significantly decreased after VFD in phosphate-buffered saline (PBS), whereas they significantly increased in protective agents (PAs) (p < 0.05). Scanning electron microscope observations show that L. plantarum 67 aggregated more to Caco-2 cells in PAs than in PBS, and its shape and size were protected. Proteomics detection findings indicated that differentially expressed proteins (DEPs) related to adhesins and vitality and their pathways in L. plantarum 67 were significantly affected by VFD (p < 0.05). However, the expression of DEPs (such as cold shock protein, cell surface protein, adherence protein, chitin-binding domain and extracellular transglycosylase, membrane-bound protein) was improved by PAs. Compared with PBS, the PAs significantly adjusted the phosphotransferase system and amino sugar and nucleotide sugar metabolism pathways (p < 0.05). VFD decreased the adhesion and vitality of L. plantarum 67, while the PAs could exert protective effects by regulating proteins and pathways related to adhesion and vitality.

Intestinal colonization is beneficial to the role of probiotics, and prebiotics can promote the adhesion and colonization of probiotics in the intestine. This study optimized the combination of complex prebiotics that could improve the growth ability and adhesion ability of Limosilactobacillus fermentum (L. fermentum) DALI02 to Caco-2 cells in vitro and determined the effect of its colonization quantity and colonization time in the immunocompromised rat model. The results showed that all five prebiotics (fructo-oligosaccharides (FOS), galacto-oligosaccharides (GOS), inulin, stachyose, and xylo-oligosaccharides (XOS)) significantly promoted the growth and adhesion of L. fermentum DALI02. It was found that 0.5% (w/w) inulin had the best growth promotion effect, and 0.5% FOS had the strongest adhesion promotion (the adhesion rate was increase by 1.75 times). In addition, 0.05% FOS, 0.20% GOS, 0.30% inulin, 0.20% stachyose, and 0.30% XOS could significantly improve the adhesion rate of L. fermentum DALI02 from 1.72% to 3.98%. After 1 w of intervention, the quantity of colonization in the fermented broth with prebiotics group was significantly higher than that in the fermented broth group. The intervention time was extended from 1 d to 4 w, and the amount of colonization of L. fermentum DALI02 in the fermented broth with prebiotics group increased significantly from 4.32 lgcopies/g to 5.12 lgcopies/g. After the intervention, the serum levels of lipopolysaccharide (LPS) and D-lactic acid in rats were significantly reduced, and the most significant was in the fermented broth with prebiotics group, with LPS and D-lactic acid levels of 74.11 pg/mL and 40.33 μmol/L, respectively. Complex prebiotics can promote the growth and adhesion of L. fermentum DALI02 and significantly increase the quantity of colonization and residence time of the strain in the intestine, which helps the restoration of intestinal barrier function and other probiotic effects.

A rat model of disordered lipid metabolism was established to study the regulation of lipid metabolism and inflammatory response by Lactobacillus rhamnosus hsryfm 1301 fermented milk. The results showed that the high-fat diet caused the disorder of lipid metabolism in rats, accompanied by the occurrence of an inflammatory response. After Lactobacillus rhamnosus hsryfm 1301 fermented milk intervention, the blood lipid level was reduced along with the activity of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) as well as triglyceride (TG) and total cholesterol (TC) contents in the liver of rats (p < 0.05), the fat vacuoles of rat hepatocytes were reduced, and the lipid accumulation in the rat liver was decreased. Liver injury was restored. Meanwhile, the levels of free fatty acid (FFA) and fatty acid synthase (FAS), acetyl coenzyme A carboxylase (ACC), lipoprotein esterase (LPL) and hepatic lipase (HL) in serum and liver of rats were significantly lower than those in the model group (p < 0.05), which indicated that fatty acid synthesis was inhibited, fatty acid production was reduced and lipid metabolism was restored to balance. In addition, the levels of reactive oxygen species (ROS) and inflammatory factors in the serum of rats were also significantly reduced (p < 0.05), and the inflammatory response of rats was restored. Lactobacillus rhamnosus hsryfm 1301 fermented milk could not only inhibit fatty acid synthase but reduce the production of excessive fatty acids, thus reducing fat accumulation, restoring the balance of lipid metabolism and alleviating the inflammatory response in rats. At the same time, it can also reduce the level of ROS through the antioxidant effect, alleviate the inflammatory response, and thus alleviate the disorder of lipid metabolism.