Research Reports No.17 Abstract(2011)

17-01

Effect of nitrite on the development of insulin resistance in obese mice

Kazuo Ohtake

pision of Pathophysiology, Department of Clinical Dietetics and Human Nutrition, Faculty of Pharmaceutical Science, Josai University

Insulin resistance can result from high levels of circulating fatty acids and inflammatory mediators which disrupt insulin signaling pathway in target organs such as skeletal muscle. However it is not well known how nitric oxide (NO) is implicated in the impaired signaling process of insulin resistance. We therefore evaluated the effect of nitrite supplementation on insulin resistance and examined a possibility of NO involvement in insulin signaling pathway of diabetic small animal models.
KKAy mice were used as a model of obese insulin resistant diabetes and C57BL/6 as control. All mice had free access to water and chow. Body weight, amounts of food intakeand blood glucose levels were checked. KKAy mice were pided randomly into 3 groups (control without nitrite, nitrite-treated with 50 mg/L, and 150 mg/L in drinking water), and 2 groups of C57BL/6 (without nitrite and nitrite-treated with 150 mg/L in drinking water). After 10 weeks, blood and gastrocnemius muscles were collected following 16h fasting, then applied to calculation of the homeostasis model assessment of insulin resistance (HOMA-IR) using plasma concentrations of glucose and insulin, and to the measurement of tissue concentrations of nitrite and nitrate (NOx) in gastrocnemius muscles using HPLC system (ENO-20).
Although diabetic KKAy mice groups showed remarkably increasing body weights along the 10-weeks experimental period compared to C57BL/6 control groups, nitrite treatment had no impact on body weight changes among each group. There were tendencies toward decreased fasting plasma levels of insulin and glucose with nitrite treatment on 10th week, but not statistically significant. Whereas HOMA-IR of diabetic KKAy mice was significantly reduced with nitrite treatment in a dose-dependent manner, which was consistent with reduced nitrite levels in the muscle of diabetic KKAy mice followed by restoration to the same levels as C57BL/6 control groups with nitrite supplementation. Additionally our recent study showed that nitrite treatment to KKAy mice remarkably enhanced Akt phosphorylation of skeletal muscle, which is an important insulin-initiated signal transduction for glucose transport.
These results suggested a possible involvement of NO and its bioavailability in the signaling process associated with insulin resistance in diabetic KKAy mice.

17-02

Identification analysis of major allergen proteins in cochineal dye

Hiroshi Akiyama

pision of Novel Foods and Immunochemistry, National Institute of Health Sciences

Carmine is a natural red pigment obtained from the dried gravid female cochineal insects (Dactilopius coccus or Coccus cacti). We identified a full-length cDNA encoding a protein, which we named CC38K, with 335 amino acids and a molecular mass calculated as 38 kDa. This amino acid sequence included all the partial amino acid sequences obtained from the purified proteins identified by IgE from allergic patients. The CC38K sequence showed homology to phospholipases. We prepared the polyclonal anti-body to the purified cochineal allergen protein and developed the ELISA detection method for determining the allergen protein using chemiluminescence detection. The allergen protein could not be detected in cochineal color products and the ingredient.
We examined the IgE response to patient's and normal sera on the CC38K after de-glycosilation using enzymes to investigate the association of sugar chain on the allergenecity. We found that there are little differences between the IgE response to patient sera on the CC38K after digestion of glycopepetidase A and that without the digestion.

17-03

Suppressive Effect of Enzymatically Modified Isoquercitrin on Phenobarbital-Induced Liver Tumor Promotion in Rats
- Studies on Modifying Effects of Antioxidants to Hepatocellular Tumor Promotion of an Oxidative Stress-Mediatied Nongenotoxic Carcinogen -

Kunitoshi Mitsumori

Graduate School, Institute of Agriculture, Tokyo University of Agriculture and Technology

To investigate the effect of enzymatically modified isoquercitrin (EMIQ) on hepatocellular tumor promotion induced by phenobarbital (PB), male rats were administered a single intraperitoneal injection of 200mg/kg N-diethylnitrosamine (DEN) then fed a diet containing PB (500 ppm) for 8 weeks, with or without EMIQ (2000 ppm) in the drinking water. One week after PB administration, rats underwent a two-thirds partial hepatectomy. The PB-induced increase in the number and area of glutathione S-transferase placental form-positive foci and the proliferating cell nuclear antigen-positive ratio was significantly suppressed by EMIQ. Real-time reverse transcription-polymerase chain reaction analysis revealed increases in mRNA expression levels of Cyp2b2 and Mrp2 in the DEN-PB and DEN-PB-EMIQ groups compared with the DEN-alone group, while the level of Mrp2 decreased in the DEN-PB-EMIQ group compared with the DEN-PB group. There were no significant changes in microsomal reactive oxygen species (ROS) production and oxidative stress markers between the DEN-PB and DEN-PB-EMIQ groups. Immunohistochemically, the constitutive active/androstane receptor (CAR) in the DEN-PB group was clearly localized in the nuclei but its immuoreactive intensity was decreased in the DEN-PB-EMIQ group. These results indicate that EMIQ suppressed the liver tumor-promoting activity of PB by inhibiting nuclear translocation of CAR, and not by suppression of oxidative stress.

17-04

Fundamental Safety Assessment of Natural Flavoring Substances

The natural source of flavoring additives 513 items were listed in 2006. Since there are various kinds of complicated materials like animals, complexes, products, components, mushrooms, seaweeds, resins and materials related to the regulations, we removed such species from this project and relisted up 450 items. We suggested that 450 items can be covered by three years during 2010 to 2012 resulted that we pided 150 items in a year. In order to make sure the safety of flavoring additives, the following investigations have been done.
1) Since all original plant species of flavoring additives have not been determined, we confirmed and determined their scientific names inpidually.
2) The survey of eating experience as food of the natural sources is important for the confirmation of safety. Therefore, the accumulated scientific data regarding eating experiences published in various fields were incorporated in this project resulting in evaluation of safety.
3) Recently so many natural compounds have been isolated and structurally elucidated depending on the development of purification and structural elucidation technique. We therefore prepared data base of components contained in inpidual flavoring additive.
The above data base promoted the scientific safety data of flavoring additives obtained from RTECS data.

17-05

Development of quality assessment method for optically active food flavor compounds and its biological behavior
(Enantiomeric analysis of rose flavor compounds by head-space SPME-GC/MS)

Koichi Saito

Department of Analytical Chemistry, Faculty of Pharmaceutical Sciences, Hoshi University

A quality assessment method for rose flavor compounds such as linalool, nerol, citronellol, phenethyl alchohol, and geraniol, was developed for commercial food flavor analysis. The simultaneous determination of the 5 kind flavor compounds, including the optical purity test for enantiomeric separation of d,l-linalool and d,l-citronellol, was achieved with a gas chromatograph-mass spectrometer (GC/MS) equipped with a cyclodextrin column b-DEX 225. The extraction of the flavor compounds from the food samples was carried out by head-space (HS) solid-phase micro extraction (SPME) technique in order to avoid the matrix effect. Optimization of the HS-SPME technique brought about excellent recoveries from the food samples. Of the 9 kind commercial samples (4 supplements, 3 candies, 2 beverages) subjected to the optical purity test, some (or all) flavors were found to be in each sample. As for the citronellol enantiomeric analysis, l-citronellol was mainly detected in most of the samples, and racemic form was minor detected. Although the l-citronellol was suggested to be derived from natural flavor compound, the racemic form was seemed to be from chemically synthesized compound. On the other hand, linalool was found to exist as nearly equal mixture of d-form and l-form, which suggested racemic form derived from chemically synthesized compound. These results show that the developed method is effective for the quality assessment of flavor compounds in commercial foods.

17-06

Studies on regulatory effect of food additives containing flavonoids on the blood glucose level

Hitoshi Ashida

Department of Agrobioscience, Graduate School of Agricultural Science, Kobe University

Dietary intake of flavonoids is suggested to be potentially beneficial for the prevention of various diseases. To prevent hyperglycemia and diabetes mellitus, regulation of the postprandial blood glucose level is one of the important issues. In this study, we investigated whether four flavonoids, quercetin, quercetin 3-glucoside (isoquercitrin), quercetin 3-O-beta-rutinoside (rutin), and enzymatically modified isoquercitrin (EMIQ), stimulated a glucose uptake activity in L6 myotubes and mouse skeletal muscle. In L6 myotubes, glucose uptake activity was stimulated by both EMIQ and isoquercitrin in a dose-dependent manner, and reached a maximum at the concentration of 10 nM and 3 mM, respectively. Quercetin also increased glucose uptake activity without significance. Both EMIQ and quercetin increased translocation of glucose transporter 4 (GLUT4) on the plasma membrane. Translocation of GLUT4 was confirmed by in vivo experiment using skeletal muscle of ICR mice, after oral administration of quercetin and EMIQ at 10 and 100 mg/kg BW. Isoquercitrin and rutin also stimulated translocation of GLUT4 in vivo. Interestingly, EMIQ suppressed the postprandial blood glucose level at 10 and 50 mg/kg BW in oral glucose tolerance test, but quercetin did not. Moreover, EMIQ did not affect maltase and sucrase activities in vivo, although it inhibited these　疝och-glycosidases in vitro. These results indicate that EMIQ have a possibility of preventing hyperglycemia through stimulation of the glucose uptake activity accompanied by translocation of GLUT4 in skeletal muscle.

17-07

A Screening analysis of polysorbates in foods

Chie Nomura

Osaka Prefectural Institute of Public Health

In April 2008, the polysorbates emulsifiers (PSs) were approved as food additives, thereafter it has been increasingly used in Japan. The analytical method in foods was notified by the Ministry Health, Labour and Welfare. In the official method, PSs were extracted from processed foods with the mixture of acetonitrile-methanol-n-hexane and the extract was cleaned up on an alumina column and a silica gel cartridge to remove food color and other interfering substances. In such a complicated process, including salting-out and liquid-liquid partition, this method often showed poor recovery of PSs.
To simplify operations and improve purification process and recovery rates, we have examined commercially available column cartridges: a silica gel, a diol, an Ag-Ion, an alumina-B, and a graphitized carbon black column cartridge. In this paper, we propose an effective new method for analysis of PSs.
A new method uses ethyl acetate containing 10% methanol for extraction of PSs, and a diol cartridge (MEGA Bond Elut-Diol) and an Alumina-B cartridge (Sep-Pak Plus Alumina B) for removal of fat and food color. The presence and the content of PSs are determined by TLC and colorimetric assay, respectively. Using a new method, the recoveries of PS80 in foods fortified at the levels 0.50-5 g/kg ranged from 47 to 92%. The limit of quantitation for PS80 in foods was 0.05 g/kg.

17-08

Study on masking effect of pectin on the contaminants in food

Yoshiko Konishi

National Institute of Health Sciences

Mycotoxins are natural contaminants that can cause adverse effects on human and animal health. It is too difficult for achieve zero exposure to mycotoxins, however, many approachs to reduce intake of mycotoxins from final food products have been tried. The development of new food processing methods useful for reducing exposure to mycotoxins is especially needed.
Pectin is a constituent of plants that has been used in the food industry as a natural ingredient. It has the ability to form gels at low concentrations. Low methoxyl pectin forms a gel easily in the presence of Ca (2+).
In this study, we developed pectin gels containing the mycotoxin deoxynivalenol (DON) and examined the effect on the absorption of DON. A low methoxy amidate (LMA) pectin was used. Three groups were examined: pectin gel (1%) containing DON (Gel containing DON group), pectin solution (1%) mixed with DON (Pectin with DON), and DON solution only (DON in PBS). They were administered orally to mice at a level of 5 mg DON/kg body weight. Blood was collected at 15, 30, and 90 min and at 3, 6, 8, and 24 h. At 6 h, urine samples were taken.
The sera of the DON in PBS and Pectin with DON showed a sharp peak of DON at 15 min after gavage that diminished by 90 min. On the other hand, the Gel containing DON group showed a relatively broad and low peak at 15 min after gavage. AUC of DON in serum in the Gel containing DON group was 1/4 of that in the DON in PBS. DON levels in the urine, liver and other organs in the Gel containing DON group were lower than those in DON in PBS. These results demonstrate that pectin gelation suppressed absorption of DON in the gastrointestinal system. The low methoxyl pectin would be a valuable processing tool to reduce the exposure to this mycotoxin.

17-09

The development of the identification method by the morphologic technique of the gum arabic

Eiji Sakai

Gifu Pharmaceutical University

The gum arabic ia the secretions obtatined from the stems and branches of Acacia senegal Willdenow or Acacia seyal Delile. It is used by many industrial products including food and medical supplies. Uses are different by the difference of the original plant. Therefore I performed microscopy for the purpose of the discrimination method development of rough raw materials. I cannot distinguish it with the powdery form, but the possibility that I could suppose the original plant by scanning the piece of a plant included in it was suggested.

17-10

Characterization of the function of food additives by using a cell-based assay
involving human taste receptor-expressing cells

Takumi Misaka

Graduate School of Agricultural and Life Sciences, The University of Tokyo

In mammals, sweet taste perception is mediated by the heterodimeric G protein-coupled receptor T1R2/T1R3. An interesting characteristic of this sweet taste receptor is that it has multiple ligand binding sites. In this study, we constructed a cell line that stably expresses the human sweet taste receptor, i.e., hT1R2/hT1R3, and a functional chimeric G protein, i.e., hGa16gust44, by using the Flp-In system for measuring receptor activity. A cell-based assay was performed to examine the response of this cell line to various sweet tastants. In the case of cells that stably expressed hT1R2/hT1R3, intense and frequent responses were observed on treatment with compounds that activate hT1R2/hT1R3, such as aspartame, acesulfame K, and sucrose. Because our cell line provides a constant response to sweeteners for long-term, it could be a useful evaluation system for characterizing novel taste modulators, especially sweet-taste enhancers.

17-11

Functional interaction between food colorants and food constituents

Yoshimasa Nakamura

Graduate School of Natural Science and Technology, Okayama University

The fluorescein-type food colorants, including phloxine (PhB), erythrosine B and rose bengal, induced photocytotoxicity in human acute promyelocytic leukemia HL-60 cells among 12 artificial food colorants that were approved by the Food Sanitation Law of Japan. A significant DNA ladder formation and increased caspase-3 activity were observed in HL-60 cells treated with PhB and light exposure, suggesting that apoptosis induction might be involved in the inhibition of cell viability. Co-treatment of catalase completely blocked the photocytotoxicity by PhB in HL-60 cells but not in Jurkat cells. The increased level of intracellular peroxides in PhB-treated HL-60 cells was significantly inhibited by catalase. These results suggested that PhB induced a differential photodynamic action in the myeloperoxidase-containing leukemia cells through an extracellular hydrogen peroxide-dependent mechanism. The food-derived antioxidants such as a-tocopherol and cysteine inhibited PhB-induced photocytotoxicity at higher concentrations. However, it should be noted that lower concentration of cysteine potentiated PhB-induced photocytotoxicity possibly through enhancement of one-electron reduction in Type I reaction. In addition, such fluorescein-type red colorants also showed potent inhibitory activity against NADPH oxidase-dependent ROS production under dark conditions. This report attempts to provide biological evidence that artificial food colorants can functionally interact with food antioxidants.

17-12

Study on the roles of flavonoids on the intestinal barrier function

Takuya Suzuki

Hiroshima University, Graduate School of Biosphere Science

One of the most important functions of gastrointestinal epithelial cells is to provide a physical barrier to the diffusion of pathogens, toxins, and antigens from the luminal environment into the circulation. Defect of the barrier function is associated with various diseases, including inflammatory bowel disease. The intestinal barrier integrity depends on the coordinate expression and interaction of proteins in cell-cell junctional complexes, including tight junction (TJ), which regulates paracellular permeability to solutes between the epithelial cells. The present study explored the promotive effect of polyphenols used as food additives on the intestinal TJ barrier function using intestinal epithelial cells and a mouse model of colitis. In human intestinal Caco-2 cells, apical administrations of kaempferol, resveratrol, hesperetin, and naringenin promoted the TJ barrier integrity, while daidzein and genistein decreased the integrity. Resveratrol increased the assembly of TJ proteins, ZO-1, ZO-2, and occludin, to the TJ complex. In mice, administration of dextran sodium sulfate impaired the intestinal barrier integrity, as indicated by increase in FITC-dextran flux and decrease in the TJ protein expression, and induced colitis, as indicated by decreases in colon length and body weight. Supplemental feeding of resveratrol partially ameliorated the impaired intestinal barrier and colitis. Taken together, some polyphenols used as food additives have roles in the regulation of intestinal TJ barrier and resveratrol has promotive and protective effects of intestinal barrier and ameliorative effect on colitis.

17-13

Pharmacokinetis and mechanisms for intestinal absorption of flavonoid glycosides and their anti-allergic activity

Toshiaki Makino

Department of Pharmacognosy, Graduate School of Pharmaceutical Sciences, Nagoya City University

Quercetin, a flavonol contained in various vegetables and herbal medicines, has various biological activities including anti-cancer, anti-allergic and anti-oxidative activities. However, its low oral bioavailability due to insolubility in water has limited its use as a food additive or dietary supplement. In previous reports, we found that the elongation of a-linkage of glucose moiety in quercetin-3-O-glucoside (isoquercitrin, IQC) enhances the bioavailability of quercetin and that IQC, quercetin-3-O-maltoside (Q3M) and enzymatically modified isoquercitrin (a-oligoglucosyl isoquercitrin, EMIQ) were rapidly hydrolyzed to quercetin by rat intestinal epithelium epithelial enzymes such as mucosal maltase-glucoamylase (MGM) and lactase phrolizin hydrolase (LPH). In the present study, we investigate their anti-allergic effects using murine passive cutaneous anaphlaxis (PCA) reaction. EMIQ significantly suppressed PCA reaction in dose dependent manners. The suppressive effects of EMIQ and IQC at the dosage of 4 mmol/kg at 4 mmol/kg were 70% and 35% , respectively, while quercetin and a-polyglucosyl rutin did not show significant effects. The elongation of a-linkage of glucose moiety in IQC enhances not only the bioavailability of quercetin but also its anti-allergic effects.

17-14

Analysis of the effect of green odor compounds on the brain when used as food additives and search for biomarkers of response to green odor compounds

Yoko Kobayashi 1,2), Hironari Kako 2), and Hidehiko Yokogoshi 2)

Green odor is mainly composed of 8 odors emitted by 6-carbon aliphatic alcohols and aldehydes. Green odor compounds are widely used in foods as flavoring agents to reconstitute the fresh green flavor. Odor compounds in foods are ingested along with nutrients. According to our overall hypothesis, odor compounds added to foods directly stimulate and regulate brain function after they are ingested. Previously, we reported that green odor compounds stimulated dopamine release from rat striatal slices and rat pheochromocytoma (PC12) cells that were capable of dopamine synthesis and release. Among all the reagents that we analyzed, n-hexanal (hexanal), a green odor compound, showed the strongest effect on dopamine release. Further, in PC12 cells, proteins with a molecular weight of approximately 130 kDa (130-kDa proteins) were dephosphorylated after stimulation with hexanal.
The aims of this study are to examine the effect of hexanal in the brain after oral intake by using a microdialysis method and to search for biomarkers of response to hexanal using PC12 cells.
In rats, hexanal when directly administered to the brain using a microprobe stimulated dopamine release. This release was regulated by both intracellular and extracellular calcium. Therefore, an increase in extracellular dopamine was detected after oral administration of hexanal in rats that underwent microdialysis with a microprobe. The increase in extracellular dopamine was not detected after stimulation with the volatile hexanal.
When PC12 cells were stimulated with reagents that strongly stimulate dopamine release, the 130-kDa protein was dephosphorylated. This protein was examined to determine whether it could act as a biomarker in response to hexanal. Calcium-dependent secretion activator 1 (CAPS1) was identified from a database as a candidate and was subjected to western blot analysis. However, an anti-CAPS antibody did not react with this 130-kDa protein.
These results suggested that hexanal when orally ingested reaches the brain and stimulates dopamine release, and that a 130-kDa protein, other than CAPS1, is a biomarker candidate for hexanal response.

17-15

Screening of Sugar Ester Surfactants That Enable Extremely High Inclusion of O/W Micelles in Dried Powders

Koreyoshi Imamura

Department of Bioscience and Biotechnology, Faculty of Engineering, Okayama University

An amorphous matrix, comprised of sugar molecules, can stably include various sizes of matters including low MW substances, protein, clusters, and cells. Micelles of oil in water can be embedded in an amorphous sugar matrix, which is essential to transform lipid solution into dry powder in food manufacturing industries. The combinations of surfactant and sugar as well as oil phase would be determinant for the micelle-inclusion characteristics of amorphous sugar matrix during drying. In this study, various types of surfactant, especially sugar surfactants, were investigated for their impacts on the micelle-inclusion characteristics of amorphous sugar matrix during freeze-drying. Sugar esters, having longer aliphatic chain and containing di- and tri-esters, tended to show greater retention of micelles after freeze-drying. Micelle-inclusion characteristics for various types of sugar were also compared and discussed in connection with the flexibility of sugar matrix at the drying stage where the loss of micelle from sugar matrix occurred. The micelle retention during freeze-drying was significantly improved when mono- and polysaccharides were mixed and thus the matrix flexibility was optimized. Furthermore, some sugar esters were enzymatically synthesized and purified.

17-16

Synergistic effect of paired antimicrobial combinations against L. monocytogenes growth in RTE seafood

Hajime Takahashi

Department of Food Science and Technology, Faculty of Marine Science, Tokyo University of Marine Science and Technology

Listeria monocytogenes causes foodborne illnesses through consumption of ready-to-eat (RTE) foods. We previously reported that RTE minced tuna and fish roe are highly contaminated with L. monocytogenes, and antimicrobials such as nisin, lysozyme, -polylysine, and chitosan are effective in controlling its growth in these RTE seafoods. In this study, the synergistic effect of paired antimicrobial combinations against L. monocytogenes growth in RTE seafood was tested. 102 CFU/g of L. monocytogenes and a mixture of two different antimicrobials were inoculated into minced tuna and salmon roe, and incubated at 10ｰC or 25ｰC for 7 days and 12 h, respectively. Among four different combinations tested, two combinations including nisin showed strong antilisterial effect in RTE seafood. These two combinations (nisin-containing Nisaplin and -polylysine-containing SAN KEEPER No.381, Nisaplin and lysozyme-containing ART FRESH 50/50) were bactericidal as well as bacteriostatic, reducing L. monocytogenes in number first and maintaining the low cell number for a long period of time. This growth control method toward L. monocytogenes is useful in RTE seafood where heating or controlling pH or water activity is not practical.

17-17

Development of Recyclable Catalyst towards Syntheses of
Macrocyclic Perfumery Ingredients

Hisahiro Hagiwara

Graduate School of Science and Technology, Niigata University

An expedient and cost-effective synthesis of E-3-methylcyclopentadec-2-en-1-one, an important precursor for R-(-)-muscone, was developed. Vapor-phase intramolecular aldol condensation of n-hexadecan-2,15-dione was catalyzed by zinc oxide at 360 ｰC to give a mixture of five cyclopentadecen-1-one isomers. Zinc oxide was re-used up to 12 times after calcination. Addition of hydrogenchloride to the mixture proceeded in the presence of 10 mol% of conc. sulfuric acid without solvent to afford 3-chloro-3-methylcyclopentadecan-1-one exclusively. Subsequent dehydrochlorination with sodium hydroxide in ethanol furnished E-3-methylcyclopentadec-2-en-1-one preferentially.

17-18

Study on a refreshment and relaxation effects of a life space perfume component. 2.

Akiyoshi Sawabe

Department of Applied Biological Chemistry, Faculty of Agriculture, Kinki University

Environmental stress is present in casual everyday life variously. How to feel about the stress is various so as to say so many men, so many minds. In this study, we investigated the building materials used in an office and life space. We reviewed a refreshment effect and aromatherapy indication by stress marker evaluation as environmental harmony evaluation for essential oils of building materials and the isolation products. In this study, for cortisol and amylase in saliva of human, the essential oil of hiba (Thujopsis dolabrata) indicated suppressive effect on stress. And their components were cedrol, widdrol, 疝och-bisabolol, citronrllic acid, and hinokithiol.

17-19

Possible as enhancer for food additives and characterization of flavoring compounds in citrus peels from Iyo

Yoshiaki Amakura

College of Pharmaceutical Sciences, Matsuyama University

Ehime is a leading prefecture which cultivates citrus fruits in Japan, and various citrus fruits as well as Citrus unshiu (Satsuma) are grown as special products. Among them, there are rare citrus fruits which are in short supply. Therefore, scientific data on their constituents including the essential oils are limited. In this study, the volatile components of citrus peel oil samples from Ehime were analyzed in order to evaluate the characteristic aroma constituents which contribute to development of the citrus flavor. First, the volatile components of 3 citrus (Kawachi-bankan, Iyokan, and Haruka) peel oils were characterized by GC-MS. Limonene (62.2%) was the most abundant compound in the Kawachi-bankan oil, followed by g-terpinene (8.1%), myrcene and nootkatone (each 2.8%). Nootkatone was not detected in the other 3 samples including commercial orange oil, therefore, it was regarded as a characteristic component of it. The composition pattern of the Iyokan peel oil was similar to that of Haruka [Limonene (89.3%), g-terpinene (3.9%), linalool (1.5%) and myrcene (1.4%) in the Iyokan peel, and limonene (83.9%), g-terpinene (4.0%), linalool (2.3%) and myrcene (1.3%) in the Haruka peel, were detected]. Next, the contents of 11 constituents (flavanol and their glycosides, polymethoxyflavone, etc.), which are characteristic citrus components in the EtOH extracts of 3 citrus peels were analyzed by HPLC. More remarkable was the high level of auraptene in Kawachi-bankan. Furthermore, the synergetic effect of each citrus oil as a food additive (antioxidant) was evaluated, and the influence of the citrus oils from Ehime on food additives was discussed.

17-20

Study on Physical Properties of Care Foods for Dysphagic Patients

Hitoshi Kumagai

Kyoritsu Women's University

To determine the appropriate properties as foods for dysphagic patients, the relationship between the velocities through the pharynx and physical properties or texture was investigated.
The viscosity, m, and the velocity through the pharynx of solutions prepared from commercial thickening agents were compared with "fractured yogurt". The value of m for the yogurt-like thickener solution was smaller than m, for the "fractured yogurt" around the shear rate m of 10 s^-1. In addition, the value of Vmax, an index for the safety of care foods, for the yogurt-like and jam-like solutions was larger than that for the fractured yogurt.
TPA (Texture Profile Analysis) parameters, "hardness" and "cohesiveness" were measured for gels and solutions prepared from commercial thickening agents. The "hardness" correlated with Vmax better than the "cohesiveness", suggesting that the "hardness" is better for an index for foods for dysphagic patients than the "cohesiveness".
The relation between the dynamic viscoelasicity and the velocities through the pharynx was examined. The value of Vmax decreased with increase in h' and was close to that of yogurt above the concentration of sol-gel transition point.

17-21

The evaluation of food texture using biomedical measurement during gel swallowing

Kazuhiro Hori, Sumiko Yokoyama*, Hirokazu Hayashi, Takahiro Ono*, Kenichi Tamine*, Rika Yahagi, Hiroshige Taniguchi, Makoto Inoue

As the bolus conditions are known to modify the swallowing behaviors, it should be important to evaluate the effects of the food texture on tongue function. The purpose of this study is to clarify the influence of gel consistency on tongue pressure during swallowing.
At first, the influence of gel consistency and the way of ingestion on tongue pressure during swallowing was measured with tongue pressure sensor sheet. The 5 ml of water and gel agent at three kinds of concentrations were chosen as a test material. In the results, the duration and magnitude of tongue pressure during squeezing or mastication were increased with increasing the concentrations of materials as well as the number of squeezing in one trial. Though they had week tendency during swallowing after squeezing, they were not affected by the concentrations during swallowing after mastication.
Secondary, the tongue pressure and videoendoscopy were recorded synchronously. The numbers of squeezing and swallowing were increased with increasing the concentrations and volume of materials. The oral transit time and pharyngeal transit time were elongated with increasing the volume of gel agents.
In conclusion, the synchronous measurement of tongue pressure and videoendoscopy suggested evaluation of oral and pharyngeal phase which were affected by the concentrations and volume of gel agents.

17-22

Application of Nisin as a disinfectant for elimination of
Listeria monocytogens in food processing plants)

Hiromi Nakamura

Osaka City Institute of Public Health and Environmental Sciences

Listeria monocytogenes is a food-borne pathogen that can lead to potentially life-threatening listeriosis in high-risk populations. In many food processing plant environments, persistent strains (PS) appear to be present for months and years. In a previous report, we observed a few clones of L. monocytogenes (distinguished with serotype, PFGE and PCR-based typing methods) were frequently isolated for a long time (up to six years) from a cold-smoked fish processing plant. We suggested that this plant might harbor PS of L. monocytogenes. The organisms are capable of forming biofilms. Biofilm formation of PS and TS (transient strains, which were isolated once during our investigation at this plant) on microtiter plates were assayed by crystal violet (CV) staining. Biofilm production of PS was significantly greater than that observed for TS. Appropriate disinfection procedures are required to eliminate Listeria biofilms in food processing plants. Nisin was recently approved for use as a food additive in Japan. The aims of present study were the following: (i) to determine the minimal inhibitory concentration (MIC) of nisin by agar dilution methods, (ii) to assess sanitizing efficacy of nisin to L. monocytogenes biofilms in food processing plants. In this study, we assessed nisin as disinfectant to compare sanitizing efficacy with benzalkonium chloride (BAC) commonly used industrial sanitizer.
The MIC of disinfectants was determined by agar dilution methods. The MIC of nisin was two to three times as much as BAC. The MIC of each disinfectant to PS was higher than that observed to TS. CV assay and ATP bioluminescence methods were used to assess sanitizing efficacy of nisin and BC to PS and TS in the biofilms on the microtiter plate. Treatment with both nisin and BAC at concentrations higher was still not enough to remove from the surface of plastic by CV assay. On the contrary, it was shown that ATP bioluminescence is good as an indicator of live biomass. Treatment with BAC, ATP bioluminescence showed that the bacteria in biofilms were killed more by the BAC treatment as BAC concentration increased, especially in the case of TS. While, ATP bioluminescence in the biofilm of PS increased by nisin treatment as nisin concentration increased. Further studies are needed to clarify the factor that ATP bioluminescence was increased in the biofilm of PS