Antimicrobial and indicator properties of edible film containing clove bud oil-loaded chitosan capsules and red cabbage for fish preservation.

For safe preservation and consumption of fish, freshness monitoring and antimicrobial control is crucial. Edible films comprising natural antimicrobial and spoilage indicator agents represent a convenient method for such preservation. Edible chitosan-based films were prepared using red cabbage (RC) and clove bud oil (CBO)-loaded chitosan/carrageenan capsules as spoilage indicator and antimicrobial agents, respectively. CBO-loaded capsules were prepared by the ionic gelation of chitosan and carrageenan. Films containing CBO capsules exhibited significantly higher antimicrobial activity than films containing non-encapsulated free CBO, as confirmed by minimum inhibitory concentration and time-kill assays. The highest antimicrobial activity was observed in the largest capsules (1.7 µm). After incubation for 48 h, the pH of fish peptone agar containing Pseudomonas fluorescens increased from approximately 6.0 to 9.0, and a color change from purple to deep blue was clearly observed during the growth of fish-spoiling bacteria. Thus, our results suggested that edible films containing CBO-loaded capsules and RC showed the potential to inhibit microbial growth in fish and to visibly indicate fish freshness.

The Effects of Laughter Therapy for the Relief of Employment-Stress in Korean Student Nurses by Assessing Psychological Stress Salivary Cortisol and Subjective Happiness.

OBJECTIVES: Korean student nurses may be exposed to stress caused by their future employment (employment stress). The aim of this study was to investigate the effects of a Laughter Program on psychological stress, by assessing salivary cortisol and the subjective happiness of student nurses in order to relieve employment stress. METHODS: A quasi-experimental, non-equivalent, control-group, and pre-test/post-test was conducted in 4th year student nurses (n = 48) from 2 universities in Korea at a time when participants' final exams and job searches were simultaneously occurring. Physiological stress (salivary cortisol), and psychological stress measured using modified Cornell Medical Index questionnaire and the Subjective Happiness Scale were used to determine the effects of the program. RESULTS: The results of the study showed that the Laughter Program was effective in relieving employment stress and increasing the subjective well-being of student nurses. Psychological stress (p < 0.001), salivary cortisol levels (p < 0.001), and subjective happiness (p < 0.001) were statistically significantly improved after the intervention compared with before the Laughter Program. CONCLUSION: This study is an effective evidence-based intervention to reduce student nurses employment stress and improve subjective happiness.

Nanoencapsulation of synergistic antioxidant fruit and vegetable concentrates and their stability during in vitro digestion.

BACKGROUND: Natural antioxidants have received increased attention owing to their safe use without side effects; however, their application has been limited because of lower antioxidant activity and stability during digestion when compared with those of synthetic antioxidants. Although research is ongoing to overcome these problems, it is still challenging to find effective solutions. In this study, we aimed to improve the properties and stability of natural antioxidants during in vitro digestion by synergistic combination and nanoencapsulation. RESULTS: Ten selected fruit and vegetable concentrates (acai berry, aronia, blackberry, cranberry, wild berry, raspberry, blueberry, red grape, cabbage, and spinach) were evaluated for their antioxidant capacity when combined via the oxygen radical absorbance capacity (ORAC) assay. Among the 45 combinations, the highest synergistic ORAC value was noted for the blueberry and cabbage concentrates (BUCA; 0.8 and 1.2 mg mL-1 ) at an antioxidant ratio of 5:5. Chitosan/carrageenan (CSCR) nanoparticles are physically more stable than chitosan/gum arabic nanoparticles during in vitro digestion and were selected for the oral delivery of BUCA. Under simulated intestinal conditions, BUCA-loaded CSCR nanoparticles showed significantly more stable antioxidant activity and total phenolic content than non-nanoencapsulated BUCA. The highest antioxidant stability was observed in the BUCA-loaded CSCR nanoparticles prepared with 0.2 mg mL-1 carrageenan, which showed two-times higher ORAC value and ten-times higher total phenolic content than non-nanoencapsulated BUCA after 12 h of in vitro digestion. CONCLUSION: CSCR nanoencapsulation of natural antioxidants could be an effective technique for improving antioxidant stability during digestion. © 2019 Society of Chemical Industry.

T-Cell Differentiation to T Helper 9 Phenotype is Elevated by Extremely Low-Frequency Electromagnetic Fields Via Induction of IL-2 Signaling.

Owing to the development of information technology and the electronics industry, and the increase in the use of electronic products, an increasing number of people are exposed to electromagnetic fields (EMFs) in daily life. There has been concern about the effects of EMFs on the human body. Th9 cells, which are characterized by the generation of interleukin-(IL-9), are a recently defined subset of T helper (Th) cells. In this study, we investigated the effect of extremely low-frequency (60 Hz) EMFs, such as those generated by household power sources, at 0.8 mT intensity on CD4+ T cells. The exposure of CD4+ T cells to such EMFs under Th9-polarizing conditions increased IL-9 secretion and gene expression of transcription factors that are important for Th9 development. The expression of GATA3 increased in the early stage, and the phosphorylation of STAT5 and STAT6, which regulate the expression of GATA3, increased. In addition, EMFs increased the expression of IL-2 by the T cells. In conclusion, the differentiation of CD4+ T cells to the Th9 phenotype was increased by exposure to extremely low-frequency EMFs, and this appeared to be dependent on the IL-2 signaling pathway. Furthermore, co-cultures of EMF-exposed Th9 cells and mast cells showed an increased expression of mast cell proteases, FcεR1α, and mast cell-derived inflammatory cytokines compared with co-cultures of non-EMF-exposed Th9 cells and mast cells. Our results suggest that EMFs enhance the differentiation of CD4+ T cells to the Th9 phenotype, resulting in mast cell activation and inflammation. Bioelectromagnetics. 2019;40:588-601. © 2019 Bioelectromagnetics Society.

Genetic Association and Expression Analyses of the Phosphatidylinositol-4-Phosphate 5-Kinase (PIP5K1C) Gene in Alcohol Use Disorder-Relevance for Pain Signaling and Alcohol Use.

BACKGROUND: The gene encoding phosphatidylinositol-4-phosphate 5-kinase (PIP5K1C) has been recently implicated in pain regulation. Interestingly, a recent cross-tissue and cross-phenotypic epigenetic analysis identified the same gene in alcohol use disorder (AUD). Given the high comorbidity between AUD and chronic pain, we hypothesized that genetic variation in PIP5K1C might contribute to susceptibility to AUD. METHODS: We conducted a case-control association study of genetic variants in PIP5K1C. Association analyses of 16 common PIP5K1C single nucleotide polymorphisms (SNPs) were conducted in cases and controls of African (427 cases and 137 controls) and European ancestry (488 cases and 324 controls) using standard methods. In addition, given the prominent role of the opioid system in pain signaling, we investigated the effects of acute alcohol exposure on PIP5K1C expression in humanized transgenic mice for the µ-opioid receptor that included the OPRM1 A118G polymorphism, a widely used mouse model to study analgesic response to opioids in pain. PIP5K1C expression was measured in the thalamus and basolateral amygdala (BLA) in mice after short-term administration (single 2 g/kg dose) of alcohol or saline using immunohistochemistry and analyzed by 2-way analysis of variance. RESULTS: In the case-control association study using an NIAAA discovery sample, 8 SNPs in PIP5K1C were significantly associated with AUD in the African ancestry (AA) group (p < 0.05 after correction; rs4807493, rs10405681, rs2074957, rs10432303, rs8109485, rs1476592, rs10419980, and rs4432372). However, a replication analysis using an independent sample (N = 3,801) found no significant associations after correction for multiple testing. In the humanized transgenic mouse model with the OPRM1 polymorphism, PIP5K1C expression was significantly different between alcohol and saline-treated mice, regardless of genotype, in both the thalamus (p < 0.05) and BLA (p < 0.01). CONCLUSIONS: Our discovery sample shows that genetic variants in PIP5K1C are associated with AUD in the AA group, and acute alcohol exposure leads to up-regulation of PIP5K1C, potentially explaining a mechanism underlying the increased risk for chronic pain conditions in individuals with AUD.

Portulaca oleracea L. Extract Enhances Glucose Uptake by Stimulating GLUT4 Translocation to the Plasma Membrane in 3T3-L1 Adipocytes.

This study investigated the effects of Portulaca oleracea L. extract on glucose uptake in 3T3-L1 adipocytes. P. oleracea extract (POE) markedly enhanced glucose uptake, which was caused by increased GLUT4 expression at the plasma membrane (PM) in 3T3-L1 adipocytes. This increase in PM-GLUT4 expression was associated with insulin receptor substrate-1 (IRS-1) phosphorylation, phosphatidylinositol 3-kinase (PI3K) activation, and Akt phosphorylation, and finally, enhanced intracellular glucose uptake. POE was not associated with protein kinase C (PKC)λ/ζ phosphorylation in the insulin signaling pathway, but did promote 5'-AMP-activated kinase (AMPK) phosphorylation. Increased glucose uptake through POE was inhibited through treating with the PI3K inhibitor or AMPK inhibitor in 3T3-L1 adipocytes. This result suggested that POE may enhance glucose uptake by stimulating GLUT4 translocation to the PM through activating the PI3K and AMPK pathway in 3T3-L1 adipocytes.

Improving the water solubility and antimicrobial activity of silymarin by nanoencapsulation.

The aims of this study were to improve the water solubility and antimicrobial activity of milk thistle silymarin by nanoencapsulation and to assess the functions of silymarin nanoparticle-containing film as an antimicrobial food-packaging agent. Silymarin nanoparticles were prepared using water-soluble chitosan (WCS) and poly-γ-glutamic acid (γ-PGA). As the WCS and silymarin concentrations increased, particle size and polydispersity index (PDI) significantly increased. Nanoencapsulation significantly improved the water solubility of silymarin 7.7-fold. Antimicrobial activity of silymarin was effectively improved when silymarin was entrapped within the nanocapsule compared to when it was not entrapped. Films incorporating silymarin nanoparticles had better antimicrobial activity than films incorporating free silymarin. The results suggest that silymarin nanoparticles have applications in antimicrobial food additives and food packing.

Nanoencapsulation of Red Ginseng Extracts Using Chitosan with Polyglutamic Acid or Fucoidan for Improving Antithrombotic Activities.

The potential of nanoencapsulation using bioactive coating materials for improving antithrombotic activities of red ginseng extract (RG) was examined. RG-loaded chitosan (CS) nanoparticles were prepared using antithrombotic materials, polyglutamic acid (PGA) or fucoidan (Fu). Both CS-PGA (P-NPs, 360 ± 67 nm) and CS-Fu nanoparticles (F-NPs, 440 ± 44 nm) showed sustained ginsenoside release in an acidic environment and improved ginsenoside solubility by approximately 122.8%. Both in vitro rabbit and ex vivo rat platelet aggregation of RG (22.3 and 41.5%) were significantly (p < 0.05) decreased within P-NPs (14.4 and 30.0%) and F-NPs (12.3 and 30.3%), respectively. Although RG exhibited no effect on in vivo carrageenan-induced mouse tail thrombosis, P-NPs and F-NPs demonstrated significant effects, likely the anticoagulation activity of PGA and Fu. Moreover, in the in vivo rat arteriovenous shunt model, P-NPs (156 ± 6.8 mg) and F-NPs (160 ± 3.2 mg) groups showed significantly lower thrombus formation than that of RG (190 ± 5.5 mg). Therefore, nanoencapsulation using CS, PGA, and Fu is a potential for improving the antithrombotic activity of RG.

Extraction optimization and nanoencapsulation of jujube pulp and seed for enhancing antioxidant activity.

The aim of this study was to optimize extraction conditions for jujube pulp and seed in order to obtain maximum active ingredient yield and antioxidant activity, as well as to prepare chitosan nanoparticles loaded with jujube pulp and seed extracts for enhancing stability. The extraction conditions, i.e. temperature, time, and ethanol concentration, were optimized at the following respective values: 61.2 °C, 38 h, and 60.4% for pulp, and 58 °C, 34 h, and 59.2% for seed. The jujube nanoparticle size significantly increased with a higher chitosan/sodium tripolyphosphate ratio and extract concentration. Entrapment efficiency was greater than 80% regardless of preparation conditions. The stabilities of jujube pulp and seed extract in terms of total phenolic content and antioxidant activity were effectively enhanced by nanoencapsulation. In conclusion, jujube pulp and seed extracts prepared using optimal conditions could be useful as a natural functional food ingredient with antioxidant activity, and nanoencapsulation can be used to improve the stability of jujube extract. Therefore, these results could be used to promote the utilization of not only jujube pulp but also seed, by product.

A rapid antimicrobial susceptibility test based on single-cell morphological analysis.

A rapid antibiotic susceptibility test (AST) is desperately needed in clinical settings for fast and appropriate antibiotic administration. Traditional ASTs, which rely on cell culture, are not suitable for urgent cases of bacterial infection and antibiotic resistance owing to their relatively long test times. We describe a novel AST called single-cell morphological analysis (SCMA) that can determine antimicrobial susceptibility by automatically analyzing and categorizing morphological changes in single bacterial cells under various antimicrobial conditions. The SCMA was tested with four Clinical and Laboratory Standards Institute standard bacterial strains and 189 clinical samples, including extended-spectrum ß-lactamase-positive Escherichia coli and Klebsiella pneumoniae, imipenem-resistant Pseudomonas aeruginosa, methicillin-resistant Staphylococcus aureus, and vancomycin-resistant Enterococci from hospitals. The results were compared with the gold standard broth microdilution test. The SCMA results were obtained in less than 4 hours, with 91.5% categorical agreement and 6.51% minor, 2.56% major, and 1.49% very major discrepancies. Thus, SCMA provides rapid and accurate antimicrobial susceptibility data that satisfy the recommended performance of the U.S. Food and Drug Administration.

Characteristics and antioxidant activity of Elsholtzia splendens extract-loaded nanoparticles.

Elsholtzia splendens extract-loaded chitosan nanoparticles prepared by ionic gelation were characterized by particle size, zeta potential, entrapment efficiency, and loading efficiency. As the initial concentration of E. splendens extract was increased, the loading efficiency and zeta potential significantly increased, whereas the entrapment efficiency and particle size significantly decreased. The optimum concentration of E. splendens extract for maximum loading efficiency was found to be 0.8 mg/mL. Both free E. splendens extract and E. splendens extract-loaded chitosan nanoparticles showed concentration-dependent antioxidant activity. However, the lipid peroxidation inhibitory activity of E. splendens extract was effectively enhanced when it was entrapped within chitosan nanoparticles. Chitosan nanoparticle encapsulation is therefore a potentially valuable technique for improving the antioxidant activity of E. splendens extract.

Optimization, in vitro release and bioavailability of gamma-oryzanol-loaded calcium pectinate microparticles reinforced with chitosan.

Response surface methodology was used to optimize coating conditions, including chitosan concentration (X(1)) and coating time (X(2)), for sustained release of chitosan-coated Ca-pectinate (CP) microparticles containing oryzanol (OZ). The optimized values of X(1) and X(2) were found to be 1.48% and 69.92 min, respectively. These optimized values agreed favorably with the predicted results, indicating the utility of predictive models for the release of OZ in simulated intestinal fluid. In vitro release studies revealed that the chitosan-coated CP microparticles were quite stable under acidic conditions, but swell and disintegrate under alkaline conditions. In vivo release study of OZ, physically entrapped within chitosan-coated CP microcapsules, demonstrated the sustained release of OZ and could be used to improve the bioavailability of OZ following oral administration.

Characteristics and antioxidant activity of catechin-loaded calcium pectinate gel beads prepared by internal gelation.

Catechin-loaded calcium pectinate gel beads prepared by internal gelation were characterized for their catechin entrapment efficiency and release behavior. The entrapment efficiency was higher when the beads were prepared with a lower catechin-to-pectin ratio, shorter gelling time, higher pectin concentration, and lower acetic acid concentration. The entrapment efficiency was much higher under all tested conditions, when the beads were prepared by internal gelation instead of external gelation. The catechin release was slower for the beads prepared with lower catechin-to-pectin ratio, longer gelling time, and higher concentrations of pectin and acetic acid in both simulated gastric and intestinal fluids. Antioxidant power of catechin was effectively maintained in alkaline simulated intestinal fluid when catechin was entrapped within the beads, compared to cases where it was not entrapped, indicating that the beads can protect catechin molecules from the alkaline environment and release them in a sustained fashion.

Alpha-tocopherol-loaded Ca-pectinate microcapsules: optimization, in vitro release, and bioavailability.

We used response surface methodology to optimize microcapsule preparation conditions, including the ratio of pectin:alpha-tocopherol (TP) (X(1)), emulsifier concentration (X(2)), and CaCl(2) concentration (X(3)) for maximal entrapment efficiency (EE) of TP-loaded Ca-pectinate microcapsules. The values of X(1), X(2), and X(3), optimized for maximal EE were a ratio of 9.7:6.3, and 1.33% and 5.09%, respectively. The experimental results obtained from the optimum formulation agreed with the predicted results, indicating the usefulness of models for EE. TP release from the Ca-pectinate microcapsules prepared according to the optimized conditions was slow and incomplete in simulated gastric fluid, whereas it was relatively rapid and considerably sustained in simulated intestinal fluid. An in vivo release study revealed that physical entrapment of TP within Ca-pectinate microcapsules can be a good technique to demonstrate the sustained release pattern of TP and to improve the bioavailability for TP following oral administration.

Gamma-oryzanol-loaded calcium pectinate microparticles reinforced with chitosan: optimization and release characteristics.

Response surface methodology was used to optimize microparticle preparation conditions, including the ratio of pectin:gamma-oryzanol (OZ) (X(1)), agitation speed (X(2)), and the concentration of emulsifier (X(3)), for maximal entrapment efficiency (EE) of OZ-loaded Ca pectinate microparticles. The optimized values of X(1), X(2), and X(3) were found to be 2.72:5.28, 1143.5 rpm, and 2.61%, respectively. Experimental results obtained for the optimum formulation agreed favorably with the predicted results, indicating the usefulness of predicting models for EE. In order to evaluate the effect of chitosan-coating and blending on the release pattern of the entrapped OZ from microparticles, chitosan-coated and blended Ca pectinate microparticles were prepared. Release studies revealed that the chitosan treatments, especially the chitosan-coating, were effective in suppressing the release in both simulated gastric fluid (SGF) and intestinal fluid (SIF).

Optimization of calcium pectinate gel beads for sustained-release of catechin using response surface methodology.

Response surface methodology was used to optimize bead preparation conditions, including CaCl(2) concentration (X(1)), hydroxypropylmethylcellulose concentration (X(2)), and bead-hardening time (X(3)), for the sustained-release of catechin from the calcium pectinate gel beads reinforced with liposomes and hydroxypropylmethylcellulose into simulated gastric fluid (SGF) and intestinal fluid (SIF). The optimized values of X(1), X(2), and X(3) were found to be 5.82%, 0.08%, and 10.29min, respectively. The beads prepared according to the optimized conditions released only about half of the entrapped catechin into SGF while most of the entrapped catechin was released into SIF after 24h incubation.

Preparation and characterization of calcium pectinate gel beads entrapping catechin-loaded liposomes.

Calcium pectinate gel (CPG) beads entrapping catechin-loaded liposomes were prepared with or without hydroxypropylmethylcellulose (HPMC) (denoted as CPG-LH and CPG-L beads, respectively) and characterized in comparison with the CPG beads prepared without liposome and HPMC (denoted as CPG-C beads). For all types of beads, the catechin entrapment efficiency decreased by about 40-50% as the concentration of CaCl2 in gelling media increased from 2 to 6%. At a constant CaCl2 level, the entrapment efficiency was higher in the order of CPG-LH, CPG-L, and CPG-C beads. The in vitro release test showed that in simulated intestinal fluid the rate of catechin release was higher in the order of CPG-C, CPG-L, and CPG-LH beads, indicating that the catechin release was slowed by liposome and further retarded when HPMC was used simultaneously, whereas not in simulated gastric fluid. The addition of cholesterol in liposome could not retard but accelerated the catechin release. The results suggest that the CPG beads reinforced with liposome and HPMC could be employed for a sustained oral delivery of catechins, although further improvements are necessary.