Xiaoqun Zeng

Immunomodulatory activity of selenium exopolysaccharide produced by Lactococcus lactis subsp. Lactis

Exopolysaccharide (EPS) was isolated and purified from Lactococcus lactis subsp. Lactis culture and then selenium chloride oxide (SeCl2O) was added to synthesise selenium-exopolysaccharide (Se-EPS). This study sought to evaluate the immunomodulatory effects of EPS and Se-EPS on the activity of peritoneal macrophages and spleen lymphocytes. In mouse peritoneal macrophages, the EPS and Se-EPS enhanced phagocytosis and increased nitric oxide (NO), IFN-γ, IL-1, IL-6 and IL-12 secretion, but not IL-10 levels. Preliminary in vitro tests revealed that EPS and Se-EPS stimulated mouse spleen lymphocyte proliferation, and strongly improved levels of TNF-α, IL-2 and IL-6 mRNA in spleen cells. These findings indicate that EPS and Se-EPS can enhance the immune response via enhanced macrophage and spleen lymphocyte stimulation. Se-EPS showed stronger immunomodulatory activity than EPS.

Highly Sensitive Electrochemical Determination of Alfatoxin B1 Using Quantum Dots-Assembled Amplification Labels

A competitive electrochemical immunoassay for highly sensitive detection of AFB1 is demonstrated using layer-by-layer (LBL) assembled quantum dots (QDs) as labels. To investigate the effects of the higher sensitivity of square wave voltammetric stripping (SWV) and of the LBL technique on the proposed immunoassays, the proposed assay was compared to electrochemical (EC) and fluorescent immunoassays, which did not use LBL technology. Peanut samples were analyzed using the three immunoassays. The limits of detection (LODs) were 0.018, 0.046 and 0.212 ng/mL, respectively, while the sensitivities were 0.308, 1.011 and 4.594 ng/mL, respectively. The proposed electrochemical immunoassay displayed a significant improvement in sensitivity, thereby providing a simple and sensitive alternative strategy for determining AFB1 levels in peanut samples.

Transepithelial transport of milk-derived angiotensin-I converting enzyme inhibitory peptide with the RLSFNP sequence

BACKGROUND To exert an antihypertensive effect after oral administration, angiotensin I-converting enzyme (ACE)-inhibitory peptides must remain active after intestinal transport. The purpose of this article is to elucidate the transport permeability and route of ACE-inhibitory peptide Arg-Leu-Ser-Phe-Asn-Pro (RLSFNP) across the intestinal epithelium using Caco-2 cell monolayers. RESULTS Intact RLSFNP and RLSFNP breakdown fragments F, FNP, SFNP and RLSF were found in RLSFNP transport solution across Caco-2 cell monolayers using ultra-performance liquid chromatography-tandem mass spectrometry. RLSFNP fragments FNP, SFNP and RLSF also contributed to ACE inhibitory effects. Protease inhibitors (bacitracin and leupeptin) and absorption enhancers (sodium glycocholate hydrate, sodium deoxycholate and Na2 EDTA) improved the transport flux of RLSFNP. A transport inhibitor experiment showed that intact RLSFNP may be transported via the paracellular route. CONCLUSION Intact RLSFNP can be transported across the Caco-2 cell monolayers via the paracellular route. Extensive hydrolysis was the chief reason for the low permeability of RLSFNP.

Proteome analysis of Lactobacillus plantarum strain under cheese-like conditions.

UNLABELLED As a food grade fermentation starter, Lactobacillus plantarum (L. plantarum) also named as the secondary starters during cheese ripening. In this study, the concentration of NaCl was screened as the main factor in the cheese-like conditions (15°C, pH5.2, 6% NaCl) to assess the potential properties of L. plantarum. A comprehensive proteome profile of L. plantarum strain was analyzed with iTRAQ proteomics methods fractionated by SCX chromatography. Proteins involved in carbohydrate transport and metabolism, cell envelope, peptide-glycan biosynthesis and lipid transport and metabolism were found significant changes. Meanwhile, the same trends were found in mRNA expression levels analyzed by RT-PCR. Some general transportation proteins related to ion transporters were detected as more abundant, which may reveal a rescue mechanism of the microbe in sodium-dependent glucose transfer. The understanding of L. plantarum proteome in salt tolerance could be useful to get strain adapted for specific applications. BIOLOGICAL SIGNIFICANCE The bacterial biota has a primary role in affecting cheese quality. Under cheese-like conditions, L. plantarum mainly increased the levels of enzymes that responsible for the flavour development during cheese ripening. However, the mechanisms of proteomic adaptation remain largely unclear in unraveling details of the salt tolerance of L. plantarum. This study revealed a dramatic change involved in carbohydrate transport and metabolism, cell envelope, peptide-glycan biosynthesis, lipid transport and metabolism, and glycolysis. Meanwhile, these pathways provide a comprehensive proteome profile of L. plantarum survived under cheese-like conditions. Furthermore, this study shows that iTRAQ proteomics provide more reliable information in describing the molecular rescue strategy of L. plantarum in sodium-dependent glucose transfer.

Effects of Salt Stress on Carbohydrate Metabolism of Lactobacillus plantarum ATCC 14917.

Lactic acid bacteria are widely used in fermented foods, especially cheese products. In this study, we observed the salt tolerance of Lactobacillus plantarum ATCC 14917 after exposure to different concentrations of NaCl in MRS medium. Quantitative proteomic profiles using two-dimensional electrophoresis identified 384 proteins, of which 26 were upregulated and 31 downregulated. Matrix-Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry was then used to identify 11 proteins, of which three were linked to carbohydrate metabolism. The downregulation of carbamoyl phosphate synthase in carbohydrate metabolism revealed a bacterial regulation mechanism to save energy in order to survive during the salt tolerance. Other proteins were found involved in transcription–translation processes, fatty acid biosynthesis, and the primary metabolic process.

iTRAQ proteomic analysis of N-acetylmuramic acid mediated anti-inflammatory capacity in LPS-induced RAW 264.7 cells.

Lactobacillus acidophilus probiotic bacteria have lasting beneficial health effects in the gastrointestinal tract, including protecting against pathogens, improving immunomodulation, and producing beneficial bacteria‐derived molecules. In lipopolysaccharide (LPS) induced RAW 264.7 cells treated with peptidoglycan or N‐acetylmuramic acid (NAM) from L. acidophilus, 390 differentially expressed proteins (8.76%) were identified by iTRAQ analysis, 257 (5.77%) of which were upregulated and 133 (2.99%) were downregulated under LPS‐induced conditions. Most of these proteins were grouped into the following inflammation‐related cellular signaling: lysosome pathway, calcium signaling pathway, and Toll‐like receptor (TLR) signaling pathway. Among them, clathrin, SERCA, and interleukin 1 receptor antagonist were differentially expressed to a significant degree in peptidoglycan or NAM pretreated RAW 264.7 cells. Bioinformatics analysis indicated that NAM may mediate an anti‐inflammatory process via a Ca2+‐dependent NF‐κB pathway. These observations reveal new insights into the molecular mechanisms involved in the suppression of LPS‐induced macrophage inflammation by L. acidophilus.

Metabolomics analysis of Lactobacillus plantarum ATCC 14917 adhesion activity under initial acid and alkali stress

The adhesion ability of Lactobacillus plantarum affects retention time in the human gastro-intestinal tract, as well as influencing the interaction with their host. In this study, the relationship between the adhesion activity of, and metabolic changes in, L. plantarum ATCC 14917 under initial acid and alkali stress was evaluated by analyzing auto-aggregation, protein adhesion and cell adhesion in vitro. Based on scanning electron microscope (SEM) and transmission electron microscope (TEM) analysis, the morphology of the bacteria became thickset and the thickness of their cell walls decreased under initial alkali stress. The fold changes of auto-aggregation, adhere to mucin and HT-29 cell lines of L. plantarum ATCC 14917 in the acid group were increased by 1.141, 1.125 and 1.156, respectively. But decreased significantly in the alkali group (fold changes with 0.842, 0.728 and 0.667). Adhesion—related protein increased in the acid group but declined in the alkali group at the mRNA expression level according to real time polymerase chain reaction (RT-PCR) analysis. The changes in the metabolite profiles of L. plantarum ATCC 14917 were characterized using Ultra-Performance Liquid Chromatography-Electrospray ionization-Quadrupole-Time of Flight-mass spectrometry (UPLS-ESI-Q-TOF-MS). In the alkali group, the content of a lot of substances involved in the energy and amino acid metabolism decreased, but the content of some substances involved in the energy metabolism was slightly increased in the acid group. These findings demonstrate that energy metabolism is positively correlated with the adhesion ability of L. plantarum ATCC 14917. The amino-acids metabolism, especially the amino acids related to pH-homeostasis mechanisms (lysine, aspartic acid, arginine, proline and glutamic acid), showed an obvious effect on the adhesion ability of L. plantarum ATCC 14917. This investigation provides a better understanding of L. plantarum’s adhesion mechanisms under initial pH stress.

The Proliferation Mechanism of Lactobacillus plantarum RB1 Stimulated by Stachyose

The proliferation mechanism of Lactobacillus plantarum RB1 promoted by stachyose was investigated in this work. The hydrolysis of stachyose, the glycometabolism, and the cytoactivity of L. plantarum RB1 were detected after proliferation. The specific activity of α-galactosidase of L. plantarum RB1 in the stachyose group was significantly higher than the control group (without stachyose), which indicated that the stachyose induced L. plantarum RB1 and produced more α-galactosidase to hydrolyze stachyose. The glycometabolism which includes glycolysis and tricarboxylic acid (TCA) cycle was significantly enhanced in the stachyose group compared with the control group. For the glycolysis, the reducing sugar content in the fermentation broth was significantly lower, while the lactic acid content and the specific activity of lactic dehydrogenase (LDH) as the key enzyme in glycolysis were higher than in the control group. For the TCA cycle, the specific activity of pyruvate dehydrogenase (PDH) as a gatekeeping enzyme leads glycolysis to TCA cycle energy-generating pathways was significantly enhanced compared with the control group. Moreover, the cell metabolic activity of L. plantarum RB1 in stachyose was significantly higher than the control group. These results indicated that the stachyose highly promotes proliferation of lactic acid bacteria (LAB) by inducing LAB to produce more α-galactosidase to hydrolyze stachyose, increasing glycometabolism and cytoactivity of LAB, which revealed the mechanisms how the stachyose promotes the proliferation of LAB.

Effect of selenylation modification on antitumor activity of peptidoglycan from Lactobacillus acidophilus

Selenium is an essential trace element for human with the antitumor properties. In the present study, the peptidoglycan (PG) derived from Lactobacillus acidophilus was modified by selenylation with the HNO3-Na2SeO3 method. Reaction temperature, reaction duration and the selenide content were optimized according to orthogonal design of three-factors. In addition, the molecular structure of selenizing peptidoglycan (Se-PG) was determined by infrared spectroscopy analysis. Furthermore, the antitumor activity of Se-PG was also investigated in HT-29 cells. The results showed that Se-PG exerted a greater antitumor activity than non-modified PG in a dose-dependent manner. These findings indicated that selenylation modification can enhance the antitumor activity of PG, and Se-PG could achieve its potential in antitumor activity.

Purification and characterization of α‐acetolactate decarboxylase (ALDC) from newly isolated Lactococcus lactis DX

BACKGROUND Diacetyl (2,3-butanedione) is a common flavor aroma from fermented dairy products. There is a need to screen new microorganisms that can efficiently produce large amounts of diacetyl. RESULTS A new lactic acid bacterium that produced high concentrations of diacetyl was identified based on Gram staining, microscopic examination and 16S rDNA sequence analysis as Lactococcus lactis DX. Its α-acetolactate decarboxylase (ALDC) was purified using 0.45 g mL(-1) ammonium sulfate precipitation, Sephacryl S-300 and S-200 HR and native-PAGE. The purified ALDC displayed a monomer structure and had a molecular mass of about 73.1 kDa, which was estimated using SDS-PAGE. IR analysis showed that the ALDC had a typical protein structure. The optimal temperature and pH for ALDC activity were 40 °C and 6.5 respectively. The ALDC of L. lactis DX was activated by Fe(2+) , Zn(2+) , Mg(2+) , Ba(2+) and Ca(2+) , while Cu(2+) significantly inhibited ALDC activity. Leucine, valine and isoleucine activated the ALDC. CONCLUSION A strain that had high ability to produce diacetyl was identified as L. lactis DX. The difference in diacetyl production may be due to the ALDC, which is different from other ALDCs.