A nondigestible saccharide,fructooligosaccharide, increases the promotive effect of a flavonoid, α‐glucosyl‐isoquercitrin,on glucagon‐like peptide 1 (GLP‐1) secretion in rat intestine and enteroendocrine cells

This study conducted in vivo and in situ experiments with rats to investigate the glucagon‐like peptide‐1 (GLP‐1) secretion in response to oral or ileal administration of α‐glucosyl‐isoquercitrin (20–40 mmol in 2 mL; Q3G), fructooligosaccharides (200 mmol in 2 mL; FOS) and Q3G + FOS. Direct effects on GLP‐1‐producing l ‐cells were also examined by an in vitro study using a murine enteroendocrine cell line, GLUTag. To evaluate the plasma GLP‐1 level, blood samples from jugular cannula for in vivo and portal cannula for in situ experiments were obtained before and after administration of Q3G, FOS, or Q3G + FOS. We found tendencies for increases but transient stimulation of GLP‐1 secretion by Q3G in in vivo and in situ experiments. Although FOS alone did not have any effects, Q3G + FOS enhanced and prolonged high plasma GLP‐1 level in both experiments. In addition, application of Q3G on GLUTag cells stimulated GLP‐1 secretion while FOS enhanced the effect of Q3G. Our results suggest that Q3G + FOS possess the potential for the management or prevention of Type 2 diabetes mellitus (T2DM) by enhancing and prolonging the GLP‐1 secretion via direct stimulation of GLP‐1 producing l ‐cell.     

Levels of phthalates and adipates in processed foods and migration of di-isononyl adipate from polyvinyl chloride film into foods

The levels of dibutyl phthalate (DBP), butylbenzyl phthalate (BBP), di-(2-ethylhexyl) phthalate (DEPHP), di-isononyl phthalate (DINP), di-(2-ethylhexyl) adipate (DEHA) and di-isononyl adipate (DINA) were determined in 50 processed foods (ham and sausage, fried dumpling and shao-mai, fish paste products, croquette and fried fish, bread, noodle, pickles, etc.). DBP, BBP, DEHP, DINP, DEHA, and DINA were contained at nd approximately 47.7, nd approximately 16.6, nd approximately 749, nd approximately 358, nd approximately 57.2 and nd approximately 20,200 ppb, respectively. High-level contamination of DINA was found in fish paste products, croquette and shao-mai, presumably because of migration from plasticized wrapping film using for food packaging. We studied the relationship between DINA migration from wrapped PVC film into fried croquette and its standing time after frying. When the croquette was wrapped immediately after frying, the migration from wrapping film into the croquette was highest (36,400 ng/g). On wrapping after standing for 5 min and 30 min, the migration level was reduced to 1/3.5 and 1/14 of the highest level, respectively.     

Evaluation of selective plating media for the detection of heat- or freeze-injured Staphylococcus aureus

The efficacy of Baird-Parker (BP) agar, mannitol-salt-egg yolk (MSEY) agar and mannitol salt (MS) agar in detecting Staphylococcus aureus FRI-100 heated at 52 degrees C for 20 min in 100 mmol/L potassium phosphate buffer was determined. Brain heart infusion agar with 1% pyruvate (BHIP agar) supported the highest recovery of injured cells and was used as the control medium. Of the three selective media, significantly higher recovery of heat-injured cells was observed on BP agar than MSEY agar, and the poorest recovery was observed on MS agar (p < 0.05). Low recovery of unheated cells was obtained for MS compared with other media (p < 0.05). A reduction in populations occurred gradually in reagent-grade water stored for 14 days at -20 degrees C. There was no significant difference between BHIP agar and MS agar in the number of freeze-injured cells recovered from 1 to 14 days.     

Comparison of soyasaponin and isoflavone contents between genetically modified (GM) and non-GM soybeans

Soyasaponins and isoflavones are main secondary metabolites in soybeans. In this report we compared the content of secondary metabolites between genetically modified (GM) and non-GM soybeans. Six cultivars/lines of GM and six cultivars/lines of non-GM soybeans were extracted with methanol. Each extract was partitioned with aqueous methanol and hexane and the aqueous methanol fraction was partially purified by HP-20 and LH-20 column chromatography to afford crude soyasaponin and isoflavone fractions. The main A-type soyasaponin, acetylsoyasaponin A1 (AcA1), and the main B-type soyasaponins, soyasaponins I and II (I and II), in the crude fractions were identified by LC/MS analyses with authentic samples. The main isoflavones, daidzin, genistin, daidzein and genistein (DI, GI, DE and GE), in the crude fractions were identified by LC photo-diode array analyses with authentic samples. The contents of AcA1, I and II in the crude soyasaponin fractions and those of DI, GI, DE and GE in the crude isoflavone fractions were analyzed by reversed-phase HPLC. The average contents (mg/100 g) of AcA1, I, II and total of B-type soyasaponins in GM soybeans were 36.4 +/- 24.2, 51.2 +/- 11.8, 26.4 +/- 7.6 and 77.7 +/- 18.5, respectively, and those in non-GM ones were 22.3 +/- 14.7, 46.3 +/- 17.8, 19.8 +/- 9.1 and 65.9 +/- 26.9, respectively. The average contents (mg/100 g) of DI, GI, DE, GE and total isoflavones in GM soybeans were 93.1 +/- 15.5, 121.8 +/- 19.4, 0.073 +/- 0.178, 0.320 +/- 0.082 and 215.3 +/- 33.3, respectively, and those in non-GM ones were 78.8 +/- 34.6, 106.7 +/- 28.3, 2.206 +/- 4.468, 0.822 +/- 0.754 and 188.5 +/- 26.7, respectively. There were no statistically significant differences in soyasaponin and isoflavone contents between GM and non-GM soybeans. Therefore, it was estimated that the GM soybeans are equivalent to the non-GM ones in terms of the composition of the main secondary metabolites.     

Biochemical characterization of horA-independent hop resistance mechanism in Lactobacillus brevis

We isolated a strain from hop-resistant Lactobacillus hrevis ABBC45, which had lost a plasmid (pRH45) harboring a putative hop resistance gene, horA. The hop resistance level of this horA-deficient strain, named ABBC45(C), was initially low but gradually induced by repeated growth in media containing progressively increasing levels of hop compounds. Although the hop resistance level was substantially lower than that of the hop-adapted wild type strain, hop-adapted ABBC45(C) (ABBC45(CR)) was still capable of growing in beer, suggesting ABBC45 possesses at least two hop resistance mechanisms. Hop resistance acquired by ABBC45(CR) gradually diminished to the pre-adapted level, when the strain was grown repeatedly in the absence of hop compounds. ABBC45(CR) was found to be cross-resistant to several structurally unrelated drugs, including ethidium bromide, daunomycin and nisin. In addition, ABBC45(CR) was shown to extrude ethidium in an energy-dependent manner, while ABBC45(C) did not show such activity. This indicates that the efflux pump was induced by adaptation to hop compounds. The efflux activity of ethidium was reduced by the addition of hop compounds, suggesting hop compounds are also the substrate of the efflux pump. It was also shown that the efflux activity was completely dissipated with the abolition of proton motive force (PMF). These results, taken together, suggest the hop resistance mechanism of ABBC45(C) is mediated by PMF-dependent multidrug efflux pump.     

Simple and easy method to evaluate uptake potential of nanoparticles in mammalian cells using a flow cytometric light scatter analysis

Many classes of nanoparticles have been synthesized and widely applied, however, there is a serious lack of information concerning their effects on human health and the environment. Considering that their use will increase, accurate and cost-effective measurement techniques for characterizing "nanotoxicity" are required. One major toxicological concern is that nanoparticles are easily taken up in the human body. In this study, we developed a method of evaluating the uptake potential of nanosized particles using flow cytometric light scatter. Suspended titanium dioxide (TiO2) particles (5, 23, or 5000 nm) were added to Chinese hamster ovary cells. Observation by confocal laser scanning microscopy showed that the TiO2 particles easily moved to the cytoplasm of the cultured mammalian cells, not to the nucleus. The intensity of the side-scattered light revealed that the particles were taken up in the cells dose-, time-, and size-dependently. In addition, surface-coating of TiO2 particles changed the uptake into the cells, which was accurately reflected in the intensity of the side-scattered light. The uptake of other nanoparticles such as silver (Ag) and iron oxide (Fe3O4) also could be detected. This method could be used for the initial screening of the uptake potential of nanoparticles as an index of "nanotoxicity".     

Safety and structural analysis of polymers produced in manufacturing process of alpha-lipoic acid

Alpha-Lipoic acid has recently been permitted for use in foodstuffs and is contained in tablets and capsules. Although alpha-lipoic acid is synthesized from adipic acid, the safety of polymers produced during the purification and drying processes has been an issue of concern. Hence, we examined the safety profiles of thermally denatured polymer (LAP-A) and ethanol-denatured polymer (LAP-B) produced in the manufacturing process of alpha-lipoic acid. Furthermore, we conducted structural analysis of these polymers by 1H-NMR and FAB-MS spectroscopy. In a consecutive ingestion test, male and female mice ingested diet containing 0.1 and 0.2% LAP-A and -B for 4 weeks. Blood uric acid, potassium and lactate dehydrogenase (LDH) tended to increase without dose-dependency. Relative liver weights were also increased. However, male dogs that were orally administered LAP-B (500 mg/kg) once did not show any abnormalities in blood parameters or general condition. These findings indicate that alpha-lipoic acid polymers are not acutely toxic; however, chronic ingestion of these polymers may affect liver and kidney functions.     

Functional analysis of genes encoding putative oxidoreductases in Aspergillus oryzae, which are similar to fungal fructosyl-amino acid oxidase

We found 11 genes (FAO1-11) encoding putative oxidoreductases in the Aspergillus oryzae genome, which are similar to fungal fructosyl-amino acid oxidases. The cDNAs corresponding to the genes were cloned and expressed in Escherichia coli. rFao2 had fructosyl-amino acid oxidase activity, whereas rFao1 did not show any enzyme activity, even though the deduced amino acid sequence of Fao1 is identical to that of one of the fructosyl-amino acid oxidase isozymes from Aspergillus oryzae. rFao7 and rFao8 showed oxidase activity toward sarcosine, L-pipecolate, and L-proline. rFao10 was active toward only sarcosine, of the substrates tested. The functions of the other proteins were also predicted from a phylogenetic analysis.     

Role of protein kinase Cbeta in rhythmic contractions of human pregnant myometrium

To assess the role of protein kinase Cbeta (PKCbeta) in human myometrial contractions during pregnancy, we evaluated the effect of a PKCbeta inhibitor (LY333531) on the pregnant and nonpregnant myometrial contractions and compared the level of PKCbeta in the pregnant myometrium with that in the nonpregnant myometrium. The effects of LY333531 on the myometrial contractions were examined by measuring contractile activity (frequency and amplitude). PKCbeta in human myometrium was assessed at mRNA level using real-time PCR method. The characteristics of contractile activity were different between the pregnant and the nonpregnant myometrium. The amplitude of rhythmic contractions in the preterm and term myometrium was increased 2- to 2.5-fold when compared with that in the nonpregnant myometrium, but the frequency of rhythmic contractions was decreased by about half. LY333531 (10(-6) M) reduced the increased amplitude in the preterm and term myometrium by about 50%, and the inhibitory effects of LY333531 in the pregnant myometrium were significantly greater than that in the nonpregnant myometrium (about 50 vs 25%). However, the frequency in the pregnant and nonpregnant myometrium was not influenced by LY333531. Real-time PCR revealed a significant, five- to sevenfold increase in the expression of PKCbeta mRNA in the preterm and term myometrium when compared with the nonpregnant myometrium. These findings suggest that the increased amplitude of human myometrial contractions during pregnancy is related to the increased level of PKCbeta. A PKCbeta inhibitor may reduce preterm uterine contractions and prevent preterm delivery.     

Effects of dissolved organic matter surrogates on the partitioning of 17beta-estradiol and p-nonylphenol between synthetic membrane vesicles and water

Partition coefficients of two estrogenic compounds, 17beta-estradiol (E2) and p-nonylphenol (NP), between synthetic membrane vesicles (K'lipw values) and water were determined using equilibrium dialysis to evaluate the potential biological uptake of these compounds with and without the presence of dissolved natural organic matter (DOM) surrogates, Suwannee River fulvic acid, dialyzed Aldrich humic acid, and polyphenolic tannic acid. Overall, K'lipw values for E2 and NP reduced by 20-30% with the increase of DOM concentration from 0 to approximately 4 mg of C/L, a typical DOM concentration in the aquatic environment. This trend for E2 and NP is similar to that obtained by other researchers for moderately hydrophobic compounds including polycyclic aromatic hydrocarbons with three or four rings. A three-phase compartment model with three independent interactions, the sorption of E2 or NP by DOM surrogate (Koc), the sorption of DOM surrogates by membrane vesicles (KlipDOM), and the partition of E2 or NP by membrane vesicles (Klipw), was proposed, and K'lipw values for E2 or NP in the presence of DOM were calculated. The model predicted the decrease in Klipw values with the increase of DOM concentration, and the predictions using the three linear interactions agreed satisfactorily with the experimental results at relatively lower concentration of DOM.