Antibacterial effects of various molecular weight chitosans against Alicyclobacillus acidoterrestris in orange juice.

Alicyclobacillus acidoterrestris has been gaining attention due to its unique thermo-acidophilic properties and being associated with the deterioration of pasteurized beverages. The objective of this study was to evaluate the antibacterial activity of chitosan with various molecular weights (MWs) (164, 85, 29.2, and 7.1 kDa) and concentrations (0-100 µg/mL) against A. acidoterrestris and its effect on guaiacol production. Various chitosan MWs were co-incubated for 7 days, and the bacterial growth, guaiacol, and vanillic acid contents during storage were determined. The chitosans performed antibacterial effects against A. acidoterrestris. Further, 164 kDa chitosan showed excellent results in controlling the growth and guaiacol formation in A. acidoterrestris. These findings demonstrated the efficacy of chitosan antibacterial activity against A. acidoterrestris and mitigating the guaiacol formation. Chitosan's antibacterial properties are attributed to the elimination of cells and suppression of guaiacol production. This study introduces a new approach for reducing A. acidoterrestris contamination in fruit juices, with potential product quality and safety advantages.

Evaluation of Hypoglycemic and Antioxidant Activities of Soybean Meal Products Fermented by Lactobacillus plantarum FPS 2520 and Bacillus subtilis N1 in Rats Fed with High-Fat Diet.

The hypoglycemic and antioxidant activities of Lactobacillus plantarum FPS 2520 and/or Bacillus subtilis N1 fermented soybean meal (SBM) in rats fed a high-fat diet (HFD) were investigated by assessing plasma glucose levels, insulin resistance, and oxidative stress-induced organ damage. Supplementation with FPS 2520- and/or N1-fermented SBM (500 and 1000 mg/kg of body weight per day) to HFD-induced obese rats for 6 weeks significantly down-regulated the concentration of plasma glucose during the oral glucose tolerance test (OGTT), as well as the concentration of fasting plasma glucose, insulin, and the value of the homeostasis model assessment of insulin resistance (HOMA-IR). In addition, plasma and hepatic levels of malondialdehyde (MDA) were alleviated in rats fed fermented SBM, especially SBM fermented by mixed strains. Moreover, fermented SBM treatment reduced HFD-exacerbated increases in plasma aspartate aminotransferase (AST), alanine aminotransferase (ALT), creatinine, and uric acid levels. Based on these results, we clearly demonstrate the effect of fermented SBM on improving insulin resistance and oxidation-induced organ damage. Therefore, it is suggested that fermented SBM has the potential to be developed as functional foods for the management of obesity-induced hyperglycemia and organ damage.

Applications of Nisin and EDTA in Food Packaging for Improving Fabricated Chitosan-Polylactate Plastic Film Performance and Fish Fillet Preservation.

This study aimed to increase the antibacterial activity of chitosan-polylactic acid (PLA) composite film by adding nisin and ethylenediaminetetraacetic acid (EDTA). We evaluated the mechanical, physicochemical, and antibacterial properties of various PLA composite films, as well as the enhancement effect of PLA composite films with EDTA + nisin on the preservation of grouper fillets. Films of PLA alone, PLA plus chitosan (C5), PLA plus nisin + EDTA (EN2), and PLA plus chitosan plus nisin + EDTA (C5EN1 and C5EN2) were prepared. The addition of EDTA + nisin to the chitosan-PLA matrix significantly improved the antibacterial activity of the PLA composite film, with C5EN1 and C5EN2 films showing the highest antibacterial activity among the five films. Compared with the fish samples covered by C5, the counts of several microbial categories (i.e., mesophilic bacteria, psychrotrophic bacteria, coliforms, Aeromonas, Pseudomonas, and Vibrio) and total volatile basic nitrogen content in fish were significantly reduced in the samples covered by C5EN1. In addition, the counts of samples covered by C5EN1 or C5 were significantly lower compared to the uncovered and PLA film-covered samples.

Exploration of Hypoglycemic Activity of Saccharomyces pastorianus Extract and Evaluation of the Molecular Mechanisms.

Although the hypoglycemic potential of brewer's yeast extract has been reported, there is limited information pertaining to the hypoglycemic ingredients of Saccharomyces pastorianus extract and their mechanisms of action available. This study aimed to investigate the in vivo and in vitro hypoglycemic effect of S. pastorianus extract and to elucidate its molecular mechanisms. S. pastorianus extract was mainly composed of proteins followed by carbohydrates. In diabetic rats, oral administration of S. pastorianus extract significantly reduced the levels of plasma glucose and enhanced the activity of hepatic glucose-6-phosphatase dehydrogenase. Treatment with S. pastorianus extract increased the localization of type 4 glucose transporter (GLUT4), PTP, and insulin receptor at 3T3-L1 cell membranes and raised the levels of P38 MAPK, PI3K, and AKT in the cytosol. In agreement with these results, pretreatment of 3T3-L1 cells with inhibitors of PTP, PI3K, Akt/PKB, and p38 MAPK inhibited glucose uptake induced by application of S. pastorianus extract. Most importantly, a 54 kDa protein with hypoglycemic activity was identified and suggested as the major ingredient contributing to the hypoglycemic activity of S. pastorianus extract. In summary, these results clearly confirm the hypoglycemic activity of S. pastorianus extract and provide critical insights into the underlying molecular mechanisms.

Antibacterial Activity of Chitosan-Polylactate Fabricated Plastic Film and Its Application on the Preservation of Fish Fillet.

This research prepared chitosan-PLA plastic films by extrusion, analyzed the physical and mechanical properties and antibacterial activity of the fabricated plastic films, and used them to preserve grouper fillet. We added chitosan (220 kDa, 93% DD) in the weight ratio of 0.5-2% into the PLA to prepare the chitosan-PLA films. With the increasing chitosan dosage, both the water vapor transmission rate and moisture content of chitosan-PLA films increased. Among the three doses of chitosan (0.5%, 1%, and 2%) added to PLA, 0.5% chitosan-PLA film had the highest antibacterial activity. This plastic film had an inhibitory efficiency of over 95% against Escherichia coli, Pseudomonas fluorescens, and Staphylococcus aureus. The action of covering the fish fillet with 0.5% chitosan-PLA film significantly reduced several microbes' counting (i.e., mesophiles, psychrophiles, coliforms, Pseudomonas, Aeromonas, and Vibrio) and total volatile basic nitrogen (TVBN) value in the grouper fillets stored at 4 °C. Thus, such action prolongs the fish fillets' shelf life to up to at least nine days, and this 0.5% chitosan-PLA film shows promising potential for preserving refrigerated fish.

Chitosan Inhibits Helicobacter pylori Growth and Urease Production and Prevents Its Infection of Human Gastric Carcinoma Cells.

This study investigated the effects of shrimp chitosan with 95% degree of deacetylation (DD95) in combination with clinical antibiotics on the growth and urease production of Helicobacter pylori. The inhibitory effect of DD95 on the adherence of H. pylori to the human intestinal carcinoma cells (TSGH9201) was also investigated. Five strains of H. pylori, including three standard strains and two strains of clinical isolates were used as the test strains. The inhibitory effects of DD95 on growth and urease production of various strains of H. pylori increased with increasing DD95 concentration and decreasing pH values from pH 6.0 to pH 2.0. Urease activity of H. pylori at pH 2.0 in the presence of 4000 µg/mL of DD95 decreased by 37.86% to 46.53%. In the presence of 50 µg/mL antibiotics of amoxicillin, tetracycline, or metronidazole at pH 6.0 and pH 2.0, H. pylori counts were decreased by 1.51-3.19, and 1.47-2.82 Log CFU/mL, respectively. Following the addition of 4000 µg/mL DD95 into the 50 µg/mL antibiotic-containing culture medium with pH 6.0 and pH 2.0, overall H. pylori counts were strongly decreased by 3.67-7.61 and 6.61-6.70 Log CFU/mL, respectively. Further, DD95 could inhibit the adherence of H. pylori on TSGH 9201 cells, as evidenced by fluorescent microscopy and thus may potentially protect against H. pylori infection.

Evaluation of Antiobesity Activity of Soybean Meal Products Fermented by Lactobacillus plantarum FPS 2520 and Bacillus subtilis N1 in Rats Fed with High-Fat Diet.

Single strain or mixed strains of Lactobacillus plantarum FPS 2520 and Bacillus subtilis N1 were used to ferment soybean meal (SBM), and the antiobesity activity of the fermented SBM product was investigated in rats fed with high-fat diet (HFD). After fermentation, free amino nitrogen, total peptide, and isoflavone contents were markedly raised, and genistein and daidzein were the major isoflavones in the fermented SBM. After fed with HFD for 10 weeks, obese Sprague-Dawley rats were orally treated with various fermented products for 6 weeks. The body weight gains, as well as weights of abdominal fat and epididymis fat, of rats fed with fermented SBM products were significantly downregulated. The treatment with the mixed-strains fermented SBM product significantly decreased plasma levels of triglyceride (TG), total cholesterol (TC), and low-density lipoprotein-cholesterol, but increased the level of high-density lipoprotein-cholesterol. Moreover, the levels of TG, TC, fatty acid synthase, and acetyl-CoA carboxylase (ACC) in liver were diminished, and the activities of hormone-sensitive lipase and lipoprotein lipase in adipose tissue were augmented. Taken together, these data demonstrated the antiobesity activity of fermented SBM products, among which the product fermented by the mixed strains being the most effective one. Therefore, these fermented SBM products are potential to be developed as functional foods or additives for treatment of obesity and prevention against obesity-induced complications.

Effects of Chitosan on Clostridium perfringens and Application in the Preservation of Pork Sausage.

The effects of chitosan with 95% deacetylation degree (DD95) on the spore germination, cell proliferation, and heat resistance of Clostridium perfringens CCRC 10,648 and CCRC 13,019 were investigated, and its application on pork sausage with sodium nitrite reduction was also evaluated. DD95 chitosan can strongly reduce the heat resistance of both strains. The D80 and D100 values for strain CCRC 13,019 decreased from 40.98 and 4.64 min to 39.21 and 3.26 min, respectively, as a result of adding 250 ppm DD95; meanwhile, addition of chitosan decreased the D80 and D100 values for CCRC 10,648 from 41.15 and 6.46 min to 39.52 and 3.78 min, respectively. In pork sausage, addition of 3000 ppm DD95 chitosan considerably slowed down the bacterial proliferation and volatile basic nitrogen production. There were no significant differences in color (L* and b* values), shearing force, and hardness in the pork sausages with or without DD95 chitosan during storage at 4 and 25 °C. However, the addition of DD95 chitosan in pork sausage significantly retarded the decrease of the a* value. Therefore, DD95 chitosan could reduce the concentration of sodium nitrite required in pork sausages for color retention.

Evaluation of the hypoglycaemic and antioxidant effects of submerged Ganoderma lucidum cultures in type 2 diabetic rats.

We aim to investigate the hypoglycaemic and antioxidant effects of submerged Ganoderma lucidum cultures and elucidate the potential mechanisms behind these effects using a type 2 diabetic rat model. Diabetic rats were daily fed with a high-fat diet supplemented with 1% or 3% freeze-dried whole submerged cultures of G. lucidum or mycelia for 5 weeks. We observed significantly decreased fasting plasma glucose levels, homoeostasis model assessment equation-insulin resistance, and plasma glucose in oral glucose tolerance test. Furthermore, we observed increased levels of glycogen, hepatic hexokinase, glucose-6-phosphate dehydrogenase, and intestinal disaccharidase activities. G. lucidum supplement downregulated the plasma levels of aspartate aminotransferase, alanine aminotransferase, creatinine, and urea nitrogen as well as liver and kidney levels of thiobarbituric acid reactive substances. Based on the hypoglycaemic and antioxidant effects of G. lucidum submerged cultures, we recommend the potential application of these products as functional foods or additives for controlling type 2 diabetes. Abbreviations ALT: Alanine aminotransferase; AST: Aspartate aminotransferase; BUN: Blood urea nitrogen; BW: Body weight; CREA: Creatinine; FPG: Fasting plasma glucose; G6Pase: Glucose-6-phosphatase; G6PD: Glucose-6-phosphate dehydrogenase; HOMA-IR: Homoeostasis model assessment of insulin resistance; OGTT: Oral glucose tolerance test; PTP: Protein tyrosine phosphatase; STZ: Streptozotocin; TBARS: Thiobarbituric acid reactive substances.

Ganoderma lucidum culture supplement ameliorates dyslipidemia and reduces visceral fat accumulation in type 2 diabetic rats.

Diabetic rats were daily fed with a high-cholesterol diet containing 1% or 3% freeze-dried whole submerged G. lucidum culture or its mycelia for 5 weeks. Body weight, adipose tissue weight and plasma triglyceride levels were reduced, while high-density lipoprotein-cholesterol levels were elevated in rats fed with G. lucidum powder supplement diets. Notably, G. lucidum supplements downregulated the activities of hepatic acetyl-CoA carboxylase, fatty acid synthase and lipoprotein lipase, but upregulated the activity of hormone-sensitive lipase in the perirenal adipose tissues. Moreover, G. lucidum supplements increased the faecal triglyceride excretion. Therefore, daily supplementation of submerged G. lucidum culture, especially mycelia, can ameliorate dyslipidemia and reduce visceral fat accumulation in diabetic rats fed with a high-fat diet, which is closely related to the modulation of lipid synthesis, metabolism, and excretion.

Multi-residue analysis using liquid chromatography tandem mass spectrometry for detection of 20 coccidiostats in poultry, livestock, and aquatic tissues.

In this study, we developed a novel analysis method based on liquid chromatography/tandem mass spectrometry (LC-MS/MS) to allow the simultaneous identification of 20 coccidiostats in eight matrix categories, including the muscles of chicken, swine, cow, and fish as well as chicken eggs, bovine milk, and porcine viscera. In the pretreatment procedure, acetonitrile/methanol (95:5, v/v) containing 1% formic acid, 5 g of sodium acetate, and 6.0 g of anhydrous magnesium sulfate was used for extraction, followed by a clean-up procedure using n-hexane saturated with ACN to facilitate the elimination of analytes from high lipid samples. Chromatographic separations were achieved using a Poroshell 120SB C18 column and operated with a gradient mobile phase system consisting of methanol (with 0.1% formic acid) and 5 mM ammonium formate, and the MS detection was monitored simultaneously. The method was validated in accordance with the Guidelines for the Validation of Food Chemical Methods by the Taiwan Food and Drug Administration. The limit of quantitation among 8 matrices were 0.5-2 ng g-1. The proposed method proved highly effective in detecting the presence of targeted veterinary drugs, providing a high degree of precision and accuracy over a broad range of matrices.

Effect of chitosan molecular weight on anti-inflammatory activity in the RAW 264.7 macrophage model.

Chitosan (300 kDa) was degraded by cellulase to chitosans with molecular weights (MWs) of 156, 72, 7.1, and 3.3 kDa and a chitooligosaccharide mixture (COS). Effects of these on NO secretion, cytokine production, and mitogen-activated protein kinase pathways in lipopolysaccharide (LPS)-induced murine RAW 264.7 macrophages were investigated. Larger chitosans (300, 156, 72 kDa) significantly inhibited NO production, whereas smaller chitosans (7.1 & 3.3 kDa, COS) increased NO production. The 156 and 72 kDa chitosans significantly inhibited TNF-α and IL-6 production, whereas the 7.1 kDa chitosan and COS significantly induced their production. The 156 and 72 kDa chitosans inhibited NF-κB activation and iNOS expression by binding to the CR3 (for 156 kDa chitosan), or CR3 and TLR4 receptor (for 72 kDa chitosan). The smaller chitosans (e.g. 7.1 kDa chitosan and COS) activated NF-κB and enhanced iNOS expression by binding to CD14, TLR4, and CR3 receptors to activate JNK signaling proteins.

Oral administration with chitosan hydrolytic products modulates mitogen-induced and antigen-specific immune responses in BALB/c mice.

The aim of this study was to investigate whether oral administration in BALB/c mice with chitosan hydrolytic products including chitosan hydrolysate, LMWC and a chitooligosaccharide mixture (oligomixture), modulates mitogen-induced and antigen-specific immune responses. A water-soluble chitosan hydrolysate was obtained via cellulase degradation of chitosan, and a LMWC and the oligomixture were separated from this hydrolysate. In non-immunized mice, both the chitosan hydrolysate and oligomixture significantly increased the phagocytic activity of peritoneal macrophages. Three chitosan hydrolytic products significantly increased the mitogen-induced proliferation of splenocytes and Peyer's patch (PP) lymphocytes. LMWC and oligomixture up-regulated IFN-γ secretion, and induced predominantly Th1 cytokine secretion in splenocytes. In antigen-specific immunity, similar effects of the chitosan hydrolytic products were observed on augmenting ovalbumin (OVA)-, as well as mitogen-, induced proliferation of splenocytes harvested from OVA-immunized mice. Interestingly, oligomixture was the most potent chitosan hydrolytic product to elicit OVA-specific IgM, IgG, and IgA production, while LMWC was the most potent one to elevate splenic IFN-γ production and IFN-γ/IL-4 (Th1/Th2) ratio. These results provide the distinct immunomodulatory properties of chitosan hydrolytic products in response to mitogens and specific antigen, paving the way for further development and application of dietary chitosan hydrolytic products against immune disorders and infection.

Effects of chitosan molecular weight on its antioxidant and antimutagenic properties.

Chitosan (300.0kDa) was enzymatically degraded to obtain six degraded chitosans with molecular weights (MWs) of 156.0, 72.1, 29.2, 7.1, 3.3, and 2.2kDa. The antioxidant and antimutagenic properties of all seven chitosan samples were evaluated in vitro. The antioxidant scavenging activities of chitosans on hydrogen peroxide and 2, 2-diphenyl-1-picryl hydrazyl radical, and on chelating ferrous ion were significantly increased with chitosan's decreasing MW, with 2.2-kDa chitosan have the highest impact. All seven chitosan samples exhibited strong anitmutagenic effects against direct (4- nitroquinoline 1-oxide) and indirect (benzo[α]pyrene) mutagens in the presence of rat liver enzyme S9 mix in Salmonella typhimurium TA 98 and TA 100. The effect of MW on antimutagenicity was similar to that on antioxidant activity. The underlying mechanism for chitosan antimutagenicity proved to be its desmutagenic effect on mutagens.

pH Effects on solubility, zeta potential, and correlation between antibacterial activity and molecular weight of chitosan.

Six chitosans with molecular weights (MWs) of 300, 156, 72.1, 29.2, 7.1, and 3.3 kDa were prepared by cellulase degradation of chitosan (300 kDa) and ultrafiltration techniques. We examined the correlation between activity against Escherichia coli and Staphylococcus aureus and chitosan MW, and provided the underlying explanation. In acidic pH conditions, the chitosan activity increased with increasing MW, irrespective of the temperature and bacteria tested. However, at neutral pH, chitosan activity increased as the MW decreased, and little activity was observed for chitosans with MW >29.2 kDa. At pH 5.0 and 6.0, chitosans exhibited good water solubility and zeta potential (ZP) decreased with the MW, whereas the solubility and ZP of the chitosans decreased with increasing MW at pH 7.0. Particularly, low solubility and negative ZP values were determined for chitosans with MW >29.2 kDa, which may explain the loss of their antibacterial activity at pH 7.0.

In Vitro and In Vivo Anticancer Effects of Sterol Fraction from Red Algae Porphyra dentata.

Porphyra dentata, an edible red macroalgae, is used as a folk medicine in Asia. This study evaluated in vitro and in vivo the protective effect of a sterol fraction from P. dentata against breast cancer linked to tumor-induced myeloid derived-suppressor cells (MDSCs). A sterol fraction containing cholesterol, ß -sitosterol, and campesterol was prepared by solvent fractionation of methanol extract of P. dentata in silica gel column chromatography. This sterol fraction in vitro significantly inhibited cell growth and induced apoptosis in 4T1 cancer cells. Intraperitoneal injection of this sterol fraction at 10 and 25 mg/kg body weight into 4T1 cell-implanted tumor BALB/c mice significantly inhibited the growth of tumor nodules and increased the survival rate of mice. This sterol fraction significantly decreased the reactive oxygen species (ROS) and arginase activity of MDSCs in tumor-bearing mice. Therefore, the sterol fraction from P. dentata showed potential for protecting an organism from 4T1 cell-based tumor genesis.