Exploration of Exopolysaccharide from Leuconostoc mesenteroides HDE-8: Unveiling Structure, Bioactivity, and Food Industry Applications.

A strain of Leuconostoc mesenteroides HDE-8 was isolated from homemade longan fermentation broth. The exopolysaccharide (EPS) yield of the strain was 25.1 g/L. The EPS was isolated and purified, and the structure was characterized using various techniques, including X-ray diffraction (XRD), nuclear magnetic resonance (NMR) spectroscopy, Fourier-transform infrared (FT-IR) spectroscopy, high-performance size exclusion chromatography (HPSEC), and scanning electron microscopy (SEM). The monosaccharide composition of the EPS was glucose, with a molecular weight (Mw) of 1.7 × 106 Da. NMR spectroscopy revealed that the composition of the HDE-8 EPS consisted of D-glucose pyranose linked by α-(1→4) and α-(1→6) bonds. The SEM analysis of the EPS showed an irregular sheet-like structure. Physicochemical analysis demonstrated that EPSs exhibit excellent thermal stability and high viscosity, making them suitable for fermentation in heat-processed and acidic foods. Additionally, milk coagulation tests showed that the presence of EPSs promotes milk coagulation when supplemented with sucrose. It suggests that EPSs have wide-ranging potential applications as food additives, improving the texture and taste of dairy products. This study provides practical guidance for the commercial use of HDE-8 EPSs in the food and related industries.

Biosynthesis and characterization of antibacterial bacterial cellulose composite membrane composed of montmorillonite and exopolysaccharides.

Bacterial cellulose (BC), as a natural renewable polymer material, has the advantages of porous nanonetwork structure, high degree of polymerization, high purity, high crystallinity, excellent mechanical properties and biocompatibility. However, BC lacks antibacterial properties, which leads to the limitation of BC material in food packaging and medical materials. In this study, a new antibacterial material using the combination of montmorillonite (MMT), BC and exopolysaccharides (EPS) produced by Weissella confusa H2 was synthesized. Fourier infrared spectroscopy (FT-IR) and X-ray diffraction (XRD) analysis showed that BC-EPS, BC-MMT and BC-EPS-MMT composite membranes conformed to the typical type I cellulose structure. Compared to BC membrane, scanning electron microscopy (SEM) showed that the porosity of BC-EPS, BC-MMT and BC-EPS-MMT composite membranes was low and compact. The physical properties of BC-EPS, BC-MTT and BC-EPS-MTT composite membranes showed lower water vapor transmittance. The BC-MTT and BC-EPS-MTT composite membranes exhibit a lower swelling ratio in 120 min. The thermal properties show that BC-EPS, BC-MTT and BC-EPS-MTT composite membranes have higher thermal stability (352 °C, 310 °C, 314 °C). Additionally, both BC-MMT and BC-EPS-MMT demonstrated strong inhibitory effects against various bacterial strains, including Staphylococcus aureus, Escherichia coli, Salmonella paratyphi A, and Bacillus subtilis. The exceptional properties exhibited by composite membranes establishes them as a highly promising option in the field of food packaging and medical material applications.

Purification, characterization and probiotic proliferation effect of exopolysaccharides produced by Lactiplantibacillus plantarum HDC-01 isolated from sauerkraut.

In this study, an exopolysaccharide (EPS)-producing strain of Lactiplantibacillus plantarum HDC-01 was isolated from sauerkraut, and the structure, properties and biological activity of the studied EPS were assessed. The molecular weight of the isolated EPS is 2.505 × 106 Da. Fourier transform infrared spectrometry (FT-IR) and nuclear magnetic resonance (NMR) results showed that the EPS was composed of glucose/glucopyranose subunits linked by an α-(1 → 6) glycosidic bond and contained an α-(1 → 3) branching structure. X-ray diffraction (XRD) analysis revealed the amorphous nature of the EPS. Scanning electron microscopy (SEM) and atomic force microscopy (AFM) showed that the isolated EPS had a smooth and compact surface with several protrusions of varying lengths and irregularly shaped material. Moreover, the studied EPS showed good thermal stability, water holding capacity, and milk coagulation ability and promoted the growth of probiotics. L. plantarum EPS may be used as prebiotics in the fields of food and medicine.

Purification of exopolysaccharides from Lactobacillus rhamnosus and changes in their characteristics by regulating quorum sensing genes via polyphenols.

To explore the effect of Lonicera caerulea fruit polyphenols (LCP) on caries-causing bacteria, strain RYX-01 with high production of biofilm and exopolysaccharides (EPS) was isolated from the oral cavity of caries patients and was identified as Lactobacillus rhamnosus by 16S rDNA analysis and morphology. The characteristics of EPS produced by RYX-01 (EPS-CK) and those produced by adding L. caerulea fruit polyphenols (EPS-LCP) were compared to reveal whether LCP reduced the cariogenicity of RYX-01 by influencing the structure and composition of EPS. The results showed that LCP could increase the content of galactose in EPS and destroy the original aggregation state of EPS-CK but had no significant effect on the molecular weight and functional group composition of EPS (p > 0.05). At the same time, LCP could inhibit the growth of RYX-01, reduce EPS and biofilm formation and inhibit the expression of quorum sensing (QS, luxS)- and biofilm formation (wzb)-related genes. Therefore, LCP could change the surface morphology, content and composition of RYX-01 EPS and reduce the cariogenic effect of EPS and biofilm. In conclusion, LCP can be used as a potential plaque biofilm inhibitor and QS inhibitor in drugs and functional foods.

Characterization of Dextran Biosynthesized by Glucansucrase from Leuconostoc pseudomesenteroides and Their Potential Biotechnological Applications.

Glucansucrase was purified from Leuconostoc pseudomesenteroides. The glucansucrase exhibited maximum activity at pH 5.5 and 30 °C. Ca2+ significantly promoted enzyme activity. An exopolysaccharide (EPS) was synthesized by this glucansucrase in vitro and purified. The molecular weight of the EPS was 3.083 × 106 Da. Fourier transform infrared (FT-IR) and nuclear magnetic resonance (NMR) spectroscopy showed that the main structure of glucan was 97.3% α-(1→6)-linked D-glucopyranose units, and α-(1→3) branched chain accounted for 2.7%. Scanning electron microscopy (SEM) observation of dextran showed that its surface was smooth and flaky. Atomic force microscopy (AFM) of dextran revealed a chain-like microstructure with many irregular protuberances in aqueous solution. The results showed that dextran had good thermal stability, water holding capacity, water solubility and emulsifying ability (EA), as well as good antioxidant activity; thus it has broad prospects for development in the fields of food, biomedicine, and medicine.

Characterization of exopolysaccharide produced by Levilactobacillus brevis HDE-9 and evaluation of its potential use in dairy products.

The bacterial strain HDE-9 was isolated from sauerkraut and identified as Levilactobacillus brevis. An exopolysaccharide (EPS) was isolated and purified from L. brevis HDE-9, and a preliminary investigation of its structural characteristics and biological activity was conducted. The molecular weight of the EPS was >1.0 × 106 Da. Fourier transform infrared (FT-IR) spectroscopy and nuclear magnetic resonance (NMR) spectroscopy revealed that the EPS was composed of α-(1 â†’ 6) linked d-glucopyranose units. X-ray diffraction (XRD) data on the EPS revealed its non-crystalline amorphous structure. Scanning electron microscopy (SEM) of the EPS revealed a smooth surface with sheet structures. The EPS exhibited the high value in thermal stability, water solubility, water holding capacity (WHC), and emulsification activity (EA). The water contact angle of the EPS revealed relatively high hydrophobicity in the presence of sucrose. The EPS also showed a strong milk solidification capacity in a dose-dependent manner. The EPS could significantly improve the texture of yoghurt, indicating its potential application as a functional starter in the production of fermented dairy products.

Purification, characterization and partial biological activities of exopolysaccharide produced by Saccharomyces cerevisiae Y3.

In this study, the exopolysaccharide (EPS) from Saccharomyces cerevisiae Y3 was extracted and purified, the preliminary structure characteristics and biological activities was investigated. The S. cerevisiae Y3 EPS was obtained by ethanol precipitation and gel filtration chromatography. MW of purified Y3 EPS was 93,477 Da. High-performance liquid chromatography (HPLC), fourier transform infrared spectroscopy (FT-IR), nuclear magnetic resonance spectroscopy (NMR) and methylation analyses showed that the EPS was a heteropolysaccharide, which composed of 1-α-D mannose (39.8%), 1,2-α-D mannose (19.6%), 1,6-α-D linked mannose (10.4%), 1,3,6-ß-D glucose (27.5%), and 1-ß-D linked glucose (1.9%). Scanning electron microscope (SEM) and atomic force microscope (AFM) further revealed smooth and dense sheet-like structure with reticular configuration. The Congo red test and X-ray diffraction (XRD) reflected an irregular coil conformation and non-crystalline amorphous nature. The EPS exhibited good hydrophilicity, thermal stability and antioxidation ability for DPPH radicals, hydroxyl radicals and NO2-, as well as good prebiotic properties. These results indicated that Y3 EPS could be explored as a promising functional adjunct for application in probiotics and antioxidation.

Analysis of the structure and properties of dextran produced by Weissella confusa.

An EPS produced by Weissella confusa H2 was purified through Sephadex G-100, and the preliminary structure characteristics and biological activities of H2 EPS were analyzed. Molecular mass of purified H2 EPS was 2.705 × 106 Da as measured with gel permeation chromatography (GPC). Composition of monosaccharides, nuclear magnetic resonance (NMR) spectroscopy spectroscopy and fourier transform infrared (FT-IR) showed that the EPS was a linear homopolysaccharide, mainly constituted of glucose and it is suggested that the EPS was dextran with α-(1 â†’ 6) glycosidic bonds and a few α-(1 â†’ 3) branches. Atomic force micrograph (AFM) and scanning electron microscopy (SEM) analysis of dextran further revealed sheets branched microstructure anchored with many irregular protuberances in aqueous solution. The XRD pattern reflected non-crystalline amorphous nature. In addition, the solubility, water-holding capacity, thermal property, rheological property and heavy metal chelating activity of the purified H2 dextran were determined. The dissolution percentage and water holding capacity of the dextran were 98.78 ± 1.37% and 426.03 ± 7.26%, respectively. The dextran exhibited good hydrophilicity, thermal stability and heavy metal chelating activity. Rheological studies exhibited rotational speed, pH, temperature, metal ions solutions dependent semiviscous nature. These results support its use as an additive in the food and environmental protection fields.

Optimization of cultivation strategy and medium for bacteriocin activity of Enterococcus faecium HDX-2.

A lactic acid bacteria (LAB) isolated from pickled Chinese cucumber was screened for bacteriocin production. The strain was identified to be Enterococcus faecium HDX-2. Based on the Plackett-Burman (PB) experiment, glucose, Ca2+, and initial pH were found to be the most significant parameters of bacteriocin production. Afterward, effects of the three main parameters on bacteriocin activity were further investigated by central composite design (CCD) and the optimum composition was glucose 22 g/L, Ca2+ 0.6 mM, and initial pH 7.2. Optimum results showed that bacteriocin activity was increased to 1337.60 ± 36.71 AU/mL, 2.23-fold higher than in MRS medium without parameters optimization. The bacteriocin also showed significant antimicrobial activity against Listeria monocytogenes in meat and cheese model system.

Intraspecific and interspecific quorum sensing of bacterial community affects the fate of antibiotic resistance genes during chicken manure composting under penicillin G stress.

In this study, the effects of penicillin G (PENG) on the fate of bacterial communities and ß-lactamase antibiotic resistance genes (ARGs) during chicken manure composting were assessed, to illustrate the roles of PENG in ARGs behavior. The results showed that the total absolute abundances of 9 ARGs and 4 mobile genetic elements (MGEs) was significantly increased by PENG (P < 0.05). Dozens of potential hosts for ARGs were predominantly affiliated with Firmicutes, Proteobacteria, and Actinobacteria. Meanwhile, the higher concentration of PENG significantly increased the abundance of luxI and luxS in quorum sensing (QS) (P < 0.05), which enhanced the frequency of inter/intraspecific gene "communication." Redundancy analysis and structural equation modeling further revealed that QS had a strong regulatory role in horizontal gene transfer of ARGs mediated via MGEs. These results provide new insight into the mechanism of ARGs propagation in aerobic composting modified by PENG.

The response surface optimization of exopolysaccharide produced by Saccharomyces cerevisiae Y3 and its partial characterization.

Response surface methodology (RSM) was used to optimize the conditions of exopolysaccharides (EPSs) by Saccharomyces cerevisiae Y3. The results indicated that the yield of EPS reached 4.52 ± 0.14 g/L with 10.30% (w/v) sucrose, 0.64% (w/v) yeast extract, liquid volume 141.5 mL, which was 2.40 times the original EPS yield. Y3 EPS contained 83.65 ± 0.16% of total sugars, 15.27 ± 0.26% of uronic acid, 0.78 ± 0.02% of protein and 0.30 ± 0.12% of sulfuric acid groups. Y3 EPS maintained a relatively low viscosity, with intrinsic viscosities of 306.58 mL/g (25 °C) and 200.91 mL/g (35 °C), respectively. The EPS had high water solubility index (WSI), high water holding capacity (WHC) and good emulsifying ability (EA). Meanwhile, the EPS could absorb metal ions such as Cu2+, Fe2+ and Zn2+. In addition, Y3 EPS exhibited good antioxidant properties and coagulated skim milk with a concentration-dependent manner. These results indicated that S. cerevisiae Y3 EPS had applicable prospects in medicine, food, especially the dairy industry.

Ecofriendly green biosynthesis and characterization of novel bacteriocin-loaded bacterial cellulose nanofiber from Gluconobacter cerinus HDX-1.

A new strain of bacterial cellulose (BC)-producing Gluconobacter cerinus HDX-1 was isolated and identified, and a simple, low-cost complexation method was used to biosynthesis Lactobacillus paracasei 1∙7 bacteriocin BC (BC-B) nanofiber. The structure and antibacterial properties of the nanofibers were evaluated. Solid-state nuclear magnetic resonance (NMR), Fourier transform infrared spectroscopy (FT-IR) and x-ray diffraction (XRD) analysis showed that BC and BC-B nanofibers had typical crystalline form of the cellulose I. X-ray photoelectron spectrometer (XPS), scanning electron microscope (SEM) and atomic force microscopy (AFM) revealed that the bacteriocin and BC were successfully compounded, and the structure of BC-B nanofiber was tighter than BC nanofiber, with lower porosity, swelling ratio and water vapor transmission rate (WVTR). The tensile strength and Young's modulus of BC-B nanofibers were 13.28 ± 1.26 MPa and 132.10 ± 4.92 MPa, respectively, higher than that of BC nanofiber (6.12 ± 0.87 MPa and 101.59 ± 5.87 MPa), indicating that bacteriocin enhance the mechanical properties of BC nanofiber. Furthermore, the BC-B nanofibers exhibited significant thermal stability, antioxidant capacity and antibacterial activity than BC nanofiber. Therefore, bacteriocin-loaded BC nanofiber may be used as antimicrobial agents in active food packaging and medical material.

Functional Identification of the Dextransucrase Gene of Leuconostoc mesenteroides DRP105.

Leuconostoc mesenteroides DRP105 isolated from Chinese sauerkraut juice is an intensive producer of dextran. We report the complete genome sequence of Leu. mesenteroides DRP105. This strain contains a dextransucrase gene (dsr) involved in the production of dextran, possibly composed of glucose monomers. To explore the dextran synthesis mechanism of Leu. mesenteroides DRP105, we constructed a dsr-deficient strain derived from Leu. mesenteroides DRP105 using the Cre-loxP recombination system. The secondary structure prediction results showed that Leu. mesenteroides DRP105 dextransucrase (Dsr) was coded by dsr and contained 17.07% α-helices, 29.55% ß-sheets, 10.18% ß-turns, and 43.20% random coils. We also analyzed the dextran yield, monosaccharide change, organic acid, and amino-acid content of Leu. mesenteroides DRP105 and Leu. mesenteroides DRP105-Δdsr. The result showed that the lack of dsr changed the Leu. mesenteroides DRP105 sugar metabolism pathway, which in turn affected the production of metabolites.

Purification and biochemical characterization of a novel glucansucrase from Leuconostoc citreum B-2.

OBJECTIVE: Although the extracellular polysaccharides have been analyzed in the previous period, the biochemical, enzymological characters and stimulation and inhibition effect on glucansucrase are not fully understood. RESULTS: After three steps purification, salting out, DEAE-Sepharose and Sephadex G-75, the final specific activity was 264.84 U/mg protein with 4.31-fold. The SDS-PAGE analysis of fraction gave a single band 170.35 kDa in the stained gel. The active band was analyzed with LC-MS/MS to identify glucansucrase. The highest coverage rate of dextransucrase from Leu. citreum (ACY92456.2) was 55.60%, the results were speculated that the glucansucrase secreted from Leu. citreum B-2 may be a novel glucansucrase. The purified enzyme was optimally active at 20-30 °C and pH 6.0-8.0. Metal ions K+, Na+, Ca2+, Mn2+, Mg2+, and Cr+ had an apparent stimulating effect on enzyme activity, especially in divalent ions Ca2+ and Mn2+, the residual activities were higher than 200%. In a reverse, Hg+, acetonitrile, SDS, salt, and guanidine expressed inhibition effect on enzyme residual activity. The KM and Vmax were detected to be 4.82 mM and 0.97 U/mg, respectively. CONCLUSION: All these data collectively indicate that B-2 glucansucrase is a novel one, which have good properties and may applied to new food areas.

Optimization and characterization of exopolysaccharides with a highly branched structure extracted from Leuconostoc citreum B-2.

The fermentation conditions for exopolysaccharides (EPS) with a highly branched structure extracted from Leuconostoc citreum B-2 were optimized by response surface methodology (RSM). The results showed that under the optimal fermentation conditions of sucrose, yeast extract and pH of 99.80 g/L, 5.94 g/L and 6.12, respectively, the B-2 EPS yield was 59.33 ±â€¯1.34 g/L, which was 5.93 times greater than that from the initial nonoptimized conditions. Chemical composition analysis showed that the carbon, hydrogen, nitrogen and sulfur contents of the B-2 pure EPS (P-EPS) were 39.05 ±â€¯0.25%, 7.19 ±â€¯0.08%, 0.72 ±â€¯0.03% and 0%, respectively. The content of uronic acid was high, reaching 20.94 ±â€¯6.30%. B-2 EPS had certain 1,1-diphenyl-2-picrylhydrazyl (DPPH), hydroxyl (·OH), superoxide anion (O2-), hydrogen peroxide (H2O2), nitroso radical (NO2-) scavenging abilities and reducing power. The degradation temperature of the B-2 EPS was 313 °C, indicating that B-2 EPS have high thermal stability. The emulsification effect on vegetable oils was better than that of hydrocarbons. The EPS produced by Leu. citreum B-2 was activated in different concentrations of sucrose-supplemented skimmed milk, which caused the skimmed milk to have different degrees of solidification. Moreover, B-2 EPS promoted the growth of probiotic bacteria, especially Lactobacillus delbrueckii.

Purification, characterization and mode of action of enterocin, a novel bacteriocin produced by Enterococcus faecium TJUQ1.

Enterococcus faecium TJUQ1 with high bacteriocin-producing ability was isolated from pickled Chinese celery. In this study, enterocin TJUQ1 was purified by ammonium sulfate precipitation, reversed-phase chromatography (Sep-Pak C8) and cation-exchange chromatography. The activity of the purified bacteriocin was 44,566.41 ± 874.69 AU/mg, which corresponds to a purification fold of 35.89 ± 2.34. The molecular mass was 5520 Da by MALDI-TOF MS and Tris-Tricine SDS-PAGE. The result of LC-MS/MS showed that the bacteriocin shared 59.15% identity with enterocin produced by E. faecium GN (accession no. O34071). PCR amplification revealed that E. faecium TJUQ1 possesses a gene encoding enterocin B with 60% identity to enterocin B. Circular dichroism (CD) spectroscopy showed that the molecular conformation was 32.6% helix, 19.5% beta, 12.9% turn and 35.0% random. The stability of enterocin TJUQ1 was measured. After exposure at 121 °C for 15 min, the residual antimicrobial activity of enterocin TJUQ1 was 85.95 ± 1.32%. The antimicrobial activity of enterocin TJUQ1 was still active over a pH range of 3-11. Enterocin TJUQ1 was inactivated after exposure to proteolytic enzymes but was not inactivated by lipase or amylase. These results showed that enterocin TJUQ1 was a novel class II bacteriocin. Enterocin TJUQ1 showed wide antibacterial activity against food-borne gram-negative and gram-positive pathogens, such as Staphylococcus aureus, Listeria monocytogenes, Escherichia coli and Salmonella enterica. The MIC was 5.26 ± 0.24 µg/mL against L. monocytogenes CMCC 1595. SEM and TEM were used to observe the changes in the morphological and intracellular organization of L. monocytogenes CMCC 1595 cells treated with enterocin TJUQ1. The results demonstrated that enterocin TJUQ1 increased extracellular electrical conductivity, facilitated pore formation, triggered the release of UV-absorbing materials, ATP and LDH, and even caused cell lysis in L. monocytogenes CMCC 1595 cells. Based on the characterization, the wide inhibitory spectrum and mode of action determined so far, enterocin TJUQ1 is a potential preservative for the food industry.

Characterization of antibacterial bacterial cellulose composite membranes modified with chitosan or chitooligosaccharide.

To synthesize biomaterials for antibacterial applications, chitosan (CS) and chitooligosaccharide (COS) were successfully incorporated into bacterial cellulose (BC) matrix respectively. The representative Gram-positive bacteria (Staphylococcus aureus) and Gram-negative bacteria (Escherichia coli) were selected to evaluate the antibacterial effects of the composite membranes. The results manifested BC-CS and BC-COS membranes exhibited remarkable antibacterial properties against both bacteria with the inhibition ratio of 99.99 ±â€¯0.01% (S. aureus) and 99.99 ±â€¯0.01% (E. coli) for BC-CS, 99.64 ±â€¯0.18% (S. aureus) and 90.56 ±â€¯0.06% (E. coli) for BC-COS. The morphology obtained by scanning electron microscopy (SEM) showed BC-CS and BC-COS composite membranes possessed lower porosity and denser structure in comparison to pure BC. Moreover, in contrast with BC-CS, BC-COS performed better mechanical properties and also displayed stronger ABTS radical scavenging capacity of 92.62 ±â€¯2.85%. These results suggested that the development of BC-COS composite membrane would open up the possibility for the antibacterial applications in food and medical fields.

Purification and characterization of an exopolysaccharide from Leuconostoc lactis L2.

An exopolysaccharide (EPS)-producing bacterium was isolated from the homemade fermentation product of hami melon (Cucumis melo var. saccharinus) and identified as Leuconostoc lactis L2 mainly based on 16S rDNA sequence analysis and the API 50 CHL test. The monosaccharide composition of Leu. lactis L2 EPS was determined to be glucose by gas chromatography (GC), and its molecular weight was 3.7 × 103 kDa by high-performance size exclusion chromatography (HPSEC). The EPS microscopic characteristics were compact, branched and tubular under scanning electron microscopy (SEM). Atomic force microscopy (AFM) showed a smooth and consistent polymeric matrix. The EPS was a linear polymer composed of α-(1 → 6)-linked D-pyranose residues, based on the Fourier transform infrared (FT-IR), 1H, 13C nuclear magnetic resonance (NMR) and heteronuclear single quantum coherence (HSQC) spectra. This EPS exhibited a high water solubility index (WSI) and water holding capacity (WHC) (91.90 ±â€¯2.45% and 509.45 ±â€¯28.59%, respectively), as well as good emulsifying properties with many organic solvents. These results collectively suggest the applicable potential of this EPS in the food, feed and pharmaceutical fields.

Determination of glucansucrase encoding gene in Leuconostoc mesenteroides.

A glucansucrase encoding gene was cloned into pET-28a(+) vector and expression in Escherichia coli BL21(DE3). An about 160 kDa recombinant glucansucrase was purified with a yield of 50.73% and a 4.02-fold increase in activity. The 1464 amino acid residue enzyme belongs to the GH70 subfamily and shares 90% similarity with Leuconostoc sp. glucansucrase. The optimal temperature and pH were 30 °C and pH 5.5, and 80% of activity was retained after incubation at 10-30 °C and pH 5-7. Enzyme activity was strongly activated by Ca2+ and Mn2+ and inhibited by various metal ions and chemical agents, and a high affinity for sucrose (Km = 11.6 mM, Vmax = 8.1 mmol/(mL·min)). Circular dichroism (CD) and Raman spectra collectively indicated a high proportion of random coil structure.