加藤 陽二

ABSTRACT The PtO2-catalyzed hydrogenation of curcumin produced slightly predominant meso-octahydrocurcumin than raceme octahydrocurcumin. Similar result was found in the product obtained from tetrahydrocurcumin and NaBH4, whereas using palladium carbon as a catalyst increased the meso-octahydrocurcumin ratio. Compared with chemical methods, baker's yeast produced 3S,5S-octahydrocurcumin and meso-octahydrocurcumin from tetrahydrocurcumin. The different activity between raceme and meso-octahydrocurcumin was not found in our experiments.

Benzyl isothiocyanate (BITC), derived from cruciferous vegetables, is an organosulfur compound exerting antiproliferative effects in several human cancer cells. In this study, we assessed BITC as a potential osteoclastogenesis inhibitor and investigated its underlying mechanism. BITC at 5 μM significantly decreased the viability of the osteoclast-like differentiating RAW264.7 cells, coinciding with the downregulation of the primary biomarkers for osteoclast differentiation, such as the tartrate-resistant acid phosphatase activity and nuclear factor of activated T-cells gene expression. Not only BITC but also its metabolites, inhibited cell proliferation in the normal RAW264.7 cells, suggesting that BITC shows an anti-osteoclastogenesis effect in vivo after its ingestion and metabolism, possibly through an antiproliferative action. Both BITC and its metabolites also enhanced the DNA fragmentation and the caspase-3 activity, whereas their higher concentrations tended to suppress these effects. BITC was intracellularly accumulated when the cells were treated with its metabolites via their degradation into the free form. A quantitative experiment using the proteolysis/high performance liquid chromatography technique showed that the amount of BITC-lysine thiourea in the cells was also increased in a time-dependent manner, suggesting that lysine modification of the cellular proteins actually took place in the cells treated by BITC. Among the cellular proteins, the cleaved caspase-3 was identified as a potential target for lysine modification by BITC. Taken together, BITC dissociated from its metabolites as well as its free form might modulate osteoclastogenesis, possibly through inhibition of cell proliferation by protein modification.

マヌカハニーを用いて，高温加熱時における特有成分の熱安定性および，抗菌成分であるメチルグリオキサール（MGO）の減少を抑える調理法について検討した。飴の加工を想定した150℃加熱では，わずか10分間でMGOが約12％まで減少した。また，ニュージーランド第一次産業省が定める指標の一つである2’-メトキシアセトフェノンも10分間の加熱で有意に減少した。温度別に10分間加熱した試験では，90℃まではMGOの減少が認められなかったものの，120℃および150℃では顕著な減少が認められた。本研究より，120℃以上の高温で加熱されたマヌカハニーを含む加工品は，加えたもとのMGO量に比べて，最終製品では大きく損失していること予想されたことから，有効成分の減少を抑制するには，加工温度を下げ，加熱時間を短縮することが有効であることが示唆された。

Increased 5-hydroxytryptamine may be associated with the development and progression of inflammatory bowel disease. In this study, we examined the suppressive effect of flavonoids on the increased intra- and extracellular 5-hydroxytryptamine levels in rat mast RBL-2H3 cells, known to produce 5-hydroxytryptamine by the phorbol 12-myristate 13-acetate stimulation. Among the flavonoids examined, luteolin and quercetin significantly reduced the cellular 5-hydroxytryptamine concentration. Gene and protein expression analyses revealed that luteolin significantly suppressed cellular tryptophan hydroxylase 1 expression induced by phorbol 12-myristate 13-acetate stimulation. Mitogen-activated protein kinase/extracellular signal-regulated kinase signaling was also suppressed by luteolin, suggesting that this pathway is one of targets of 5-hydroxytryptamine modulation by luteolin. An in vivo experimental colitis model was prepared by administering 2.5% dextran sodium sulfate in drinking water to C57BL/6 mice for seven days. The ingestion of 0.1% dietary luteolin suppressed the increasing 5-hydroxytryptamine in the colorectal mucosa. In conclusion, luteolin possesses a suppressive effect on extensive 5-hydroxytryptamine formation in both experimental RBL-2H3 cells and colitis models.

Inflammatory bowel diseases, including Crohn’s disease and ulcerative colitis, are chronic inflammatory disorders associated with oxidative stress. The intestines produce 5-hydroxytryptamine that may negatively affect disease state under inflammatory conditions when overproduced. 5-Hydroxytryptamine is a substrate for myeloperoxidase and is converted into reactive tryptamine-4,5-dione. Here, an experimental colitis model was established through oral administration of 5% dextran sulfate sodium to ICR mice for 7 days. Furthermore, the formation of tryptamine-4,5-dione in the colorectal mucosa/submucosa and colorectal tissue was analyzed by chemical and immunochemical methodologies. First, free tryptamine-4,5-dione in the homogenate was chemically trapped by o-phenylenediamine and analyzed as the stable phenazine derivative. Tryptamine-4,5-dione localization as adducted proteins in the colorectal tissue was immunohistochemically confirmed, and as demonstrated by both methods, this resulted in the significant increase of tryptamine-4,5-dione in dextran sulfate sodium-challenged mice compared with control mice. Immunohistochemical staining confirmed tryptamine-4,5-dione-positive staining at the myeloperoxidase accumulation site in dextran sulfate sodium-challenged mice colorectal tissue. The tryptamine-4,5-dione locus in the mice was partly matched with that of a specific marker for myeloperoxidase, halogenated tyrosine. Overall, the results possibly indicate that tryptamine-4,5-dione is generated by neutrophil myeloperoxidase in inflammatory tissue and may contribute to the development of inflammatory bowel disease.

Reports on the thermal stability of manuka honey in terms of food processing have been few. This study investigated changes in nine characteristic chemicals of manuka honey during heating. Among these, methylglyoxal (MGO) and 2′-methoxyacetophenone (MAP) were significantly decreased by heating at 90 °C. To elucidate the mechanism for this decrease, artificial honey was prepared from sugars and water with MAP or MGO and then heated. The decrease of MGO was enhanced with L-proline, lysine, or arginine derivatives, accompanied by formation of 2-acetyl-1-pyrroline, MGO-derived lysine dimer, or argpyrimidine, respectively, suggesting that an amino–carbonyl reaction is one pathway for the loss of MGO. The decrease of MAP in the artificial honey depended on the volume of headspace in a vessel. MAP from heated manuka honey was also detected in the gas phase, indicating that MAP was vaporized. Heating could thus reduce the beneficial and/or signature molecules in honey.

Vascular calcification progresses under hyperphosphatemia, and represents a risk factor for cardiovascular disease in chronic kidney disease (CKD) patients. We recently indicated that phosphorus (P) fluctuations also exacerbated vascular calcification in early-stage CKD rats. Dietary fiber intake is reportedly associated with cardiovascular risk. This study investigated the effects of dietary fiber on vascular calcification by repeated P fluctuations in early-stage CKD rats. Unilateral nephrectomy rats were used as an early-stage CKD model. For 36 days, a P fluctuation (LH) group was fed low-P (0.02% P) and high-P (1.2% P) diets alternating every 2 days, and a P fluctuation with dietary fiber intake (LH + F) group was fed low-P and high-P diets containing dietary fiber alternating every 2 days. The effect on vascular calcification was measured calcium content. Effects on uremic toxin were measured levels of indoxyl sulfate (IS) and investigated gut microbiota. The LH + F group showed significantly reduced vessel calcium content compared to the LH group. Further, dietary fiber inhibited increases in blood levels of IS after intake of high-P diet, and decreased uremic toxin-producing intestinal bacteria. Dietary fiber may help suppress progression of vascular calcification due to repeated P fluctuations in early-stage CKD rats by decreasing uremic toxin-producing intestinal bacteria.

真空フライリンゴチップスと常圧フライリンゴチップスの外観，テクスチャー，嗜好性，栄養成分の比較を行い，真空フライリンゴチップスの性状に及ぼす加熱温度（75℃，85℃，95℃）の影響について検討することを目的とした。 真空フライリンゴチップスは常圧フライリンゴチップスと比較し，外観が明るく，官能評価では，色が薄く，香りが良く，油臭さが少なく，食感が好ましく，総合評価で好ましいと評価された。真空フライリンゴチップスはビタミンC量が多く，脂質酸化物が少ないことから酸化が抑制されていることが推察された。 加熱温度の比較では，真空フライリンゴチップス75℃加熱の方が，95℃加熱より色が薄く，油臭さが少なく，総合的に好ましいと有意に評価された。真空フライリンゴチップス75℃加熱の方が，95℃加熱よりビタミンC量が多いことから，アミノカルボニル反応が抑えられ，酸化が抑制されることが推察された。

Leptosperin (methyl syringate β-d-gentiobioside) is abundantly found in manuka honey, which is widely used because of its antibacterial and possible anti-inflammatory activities. The aim of this study was to examine the molecular mechanism underlying the metabolism of leptosperin. Five phytochemicals (leptosperin, methyl syringate (MSYR), glucuronate conjugate of MSYR (MSYR-GA), sulfonate conjugate of MSYR (MSYR-S), and syringic acid (SYR)) were separately incubated with HepG2 and Caco-2 cells. After incubation, we found that the concentration of MSYR decreased, whereas the concentrations of SYR, MSYR-GA, and MSYR-S increased. By profiling with inhibitors and carboxylesterases (CES1, 2), we found that the conversion from MSYR to SYR was mediated by CES1. Lipopolysaccharide-stimulated RAW264.7 cells restored MSYR-GA to MSYR possibly by the secreted β-glucuronidase. All of the mice administered with leptosperin, MSYR, or manuka honey showed higher MSYR (13.84 ± 11.51, 14.29 ± 9.19, or 6.66 ± 2.30 nM) and SYR (1.85 ± 0.66, 6.01 ± 1.20, or 8.16 ± 3.10 nM) levels in the plasma compared with that of the vehicle controls (3.33 ± 1.45 (MSYR) and 1.85 ± 0.66 (SYR) nM). The findings of our study indicate that the unique metabolic pathways of these compounds may account for possible functionalities of manuka honey.

Quercetin is a major flavonoid, present as its glycosidic forms in plant foods. In this study, quercetin-3-glucoside (Q3G) was administered intraduodenally to thoracic lymph-cannulated rats, and its lymphatic transport was investigated. The resulting lymphatic and plasma metabolites were identified with LC-MS/MS and compared with those after administration of quercetin aglycone.The total concentration of quercetin metabolites in the lymph was about four times lower than that in the plasma, and quercetin and its methylated form isorhamnetin were detected as their glucuronides, sulfates and diglucuronides both in the lymph and the plasma after Q3G and quercetin administrations. The lymph levels of the glucuronides after Q3G administration were lower than those after quercetin administration, whereas those in the plasma showed the opposite pattern. Both the lymph and plasma levels of the sulfates after Q3G administration were lower than those after quercetin administration. Some of the intestinal metabolites like quercetin monoglucuronides were transported directly into the lymph and the hepatic metabolites like the diglucuronides were eventually transferred from the plasma into the lymph.These results indicate that the absorbed Q3G is partly transported into the intestinal lymph as quercetin metabolites. Deglycosylation in the enterocyte is also suggested to affect the subsequent metabolic pathways.

There are many chemically reactive compounds, including quinone, in living systems and also food. Even after the ingestion of food polyphenols, quinones derived from catechol moieties could form endogenously in the body. Dopaquinone, dopamine quinone, estrogen-derived quinones, tryptamine-4,5-dione, and ubiquinone are examples of an endogenous quinone. These indicate that quinone is ubiquitously formed or present in living systems and food. Quinones can induce a variety of hazardous effects and also could have beneficial physiological effects. This review focuses on the chemical reactivity of quinone toward a biomolecule and its biological action.

Scope: Manuka honey, which shows strong nonperoxide-dependent antibacterial activity, contains unique components, such as methyl syringate 4-O-beta-D-gentiobioside (leptosperin) and its aglycone, methyl syringate (MSYR). To determine the potential for biological activity evoked by the ingestion of leptosperin and MSYR, we investigated the absorption and metabolism of these components in manuka honey. Methods and results: The incubation of M SYR with liver microsomes or S9 fractions in vitro resulted in the formation of MSYR-glucuronide (MSYR-GA), MSYR-sulfate (MSYR-S), and syringic acid as metabolites. Then, manuka honey (15 g) was fed to healthy human volunteers. MSYR-GA, MSYR-S, and MSYR were detected in both plasma and urine. Within plasma, their levels were highest within 0.5 h to 1 h post-ingestion, and most metabolites disappeared within 3 h. In conjunction with the disappearances, a significant amount of metabolites along with trace leptosperin was excreted in urine within 4 h. To elucidate the detailed metabolisms of leptosperin and MSYR, each compound was separately administered to mice. In each case, MSYR-GA, MSYR-S, and MSYR were detected in both plasma and urine. Conclusion: This study shows the major molecular pathway for leptosperin and MSYR metabolism and could facilitate an understanding of biological functions of manuka honey post ingestion.

真空フライと常圧フライで調理した野菜チップスの外観，テクスチャー，嗜好性，栄養成分を比較検討することを目的とした。走査型電子顕微鏡により，真空フライ品は脱水された細胞壁が観察され，常圧フライ品では油の膜でコーティングされている様子が観察された。真空フライ品が常圧フライ品より明るく，鮮やかな色を示した。官能評価により，真空フライ品が常圧フライ品に比べ，香りが良く，油臭さが少なく，食感が好ましく，総合評価で好きという評価が得られた。破断測定により，真空フライ品と常圧フライ品で一定の傾向はみられなかった。真空フライ品は常圧フライ品よりビタミンC量が有意に高値を示し，脂質酸化物が少なく，酸化抑制されていることが推察された。また，真空フライ品は常圧フライ品より抗酸化力が低値を示すことから，調理中のアミノカルボニル反応が抑制されることが推察された。

Elimination of celiac-toxic prolamin peptides and proteins is essential for Triticeae products to be gluten-free. Instead of enzymatic hydrolysis, in this study we investigated metal-catalyzed oxidation of two model peptides, QQPFP, and PQPQLPY, together with a hordein isolate from barley (Hordeum vulgare L.). We established a multiple reaction monitoring (MRM) LC-MS method to detect and quantify proline oxidation fragments. In addition to fragmentation, aggregation and side chain modifications were identified, including free thiol loss, carbonyl formation, and dityrosine formation. The immunoreactivity of the oxidized hordein isolate was considerably decreased in all metal-catalyzed oxidation systems. Cleavage of peptides or protein fragments at the numerous proline residues partially accounts for the decrease. Metal-catalyzed oxidation can thus be used in the modification and elimination of celiac-toxic peptides and proteins. (C) 2016 Elsevier Ltd. All rights reserved.

Serotonin (5-hydroxytryptamine) is a putative substrate for myeloperoxidase, which may convert it into the reactive quinone tryptamine-4,5-dione (TD). In this study, we found that the viability of human SH-SY5Y neuroblastoma cells treated with 25 mu M TD was increased to approximately 117%. On the other hand, the cell viability was significantly decreased by exposure to TD (150-200 mu M), with an increase in intracellular reactive oxygen species (ROS). Interestingly, pretreatment of SH-SY5Y cells with 100 mu M TD prevented cell death and suppressed intracellular ROS generation evoked by the addition of hydrogen peroxide (H2O2). Expression of the phase-II antioxidant enzyme NAD(P)H: quinone oxidoreductase 1 and haem oxygenase 1 were upregulated by TD at a concentration of 50-100 mu M. Nuclear factor erythroid 2-related factor 2 (Nrf2), the regulator of these enzyme, was translocated from the cytosol to the nucleus by 100 mu M TD. In summary, moderate concentrations of TD may increase the self-defence capacity of neuronal cells against oxidative stress.

The modification of 5-hydroxyindoleacetic acid (5HIAA) by myeloperoxidase with a xanthine oxidase system was investigated by chromatographic analyses. Two major products were identified as a dimer and quinone (indoleacetate dione) of 5HIAA. The formation of a quinone moiety was also confirmed by chemical trapping with o-phenylenediamine. In the presence of N-acetyl-cysteine (NAC), a quinone-NAC adduct was formed. When glyceraldehyde 3-phosphate dehydrogenase was exposed to the myeloperoxidase system with 5HIAA, quinone adducts were formed on the protein molecule. A monoclonal antibody was prepared using a quinone-modified protein as an immunogen to immunochemically detect the quinone on a protein. The established antibody recognized the quinone-NAC adduct, quinone-modified poly-L-lysine, and quinone-modified low-density lipoprotein. Quinone-modified proteins in human atherosclerotic lesions were immunohistochemically observed using the established antibody to the quinone and also a monoclonal antibody to tryptamine dione-modified protein, suggesting an occurrence of in vivo oxidation of serotonin and 5HIAA, accompanied by covalent adduction to biomolecules.

Lipid peroxidation products react with cellular molecules, such as DNA bases, to form covalent adducts, which are associated with aging and disease processes. Since lipid peroxidation is a complex process and occurs in multiple stages, there might be yet unknown reaction pathways. Here, we analyzed comprehensively 2'-deoxyguanosine (dG) adducts with oxidized arachidonic acid using liquid chromatography-tandem mass spectrometry and found the formation of 7-(2-oxo-hexyl)-etheno-dG as one of the major unidentified adducts. The formation of this adduct was reproduced in the reaction of dG with 2-octenal and predominantly with 4-oxo-2-octenal (OOE). We also found that other 2-alkenals (with five or more carbons) generate corresponding 4-oxo-2-alkenal-type adducts. Importantly, it was found that transition metals enhanced the oxidation of C4-position of 2-octenal, leading to the formation of OOE-dG adduct. These findings demonstrated a new pathway for the formation of 4-oxo-2-alkenals during lipid peroxidation and might provide a mechanism for metal-catalyzed genotoxicity.

Syringic acid is one of the key skeletal structures of plant-derived,chemicals. The derivatives)of syringic acid have certain biological functions. In this study, a monoclonal antibody to syringic acid-based phytochemicals, was prepared and characterized. The obtained antibody reacted with methyl syringate, syringic acid, and leonurine. Methyl syringate is a characteristic compound found in manuka honey, other honey varieties, and plants. Manuka honey was fractionated using HPLC, and the reactivity of the fractions with the antibody was examined. The antibody reacted with the fraction in which methyl syringate was eluted. The amount of methyl syringate in honeys as estimated by ELISA using the antibody had a good linearity compared with that estimated by HPLC. These results suggest that the antibody is applicable for the immunochemical detection of syringic acid derivatives in plants and foods.

Manuka honey is known as one of the premium honeys because of its unique property: a potent antibacterial activity. Leptosperin, methyl syringate 4-O-beta-D-gentiobioside, has been specifically identified in manuka honey. Because leptosperin is relatively stable under warmer conditions, measuring leptosperin levels may be applied to authenticate manuka honey. In this study, an immunochromatographic separation and quantification of leptosperin techniques have been developed. The concentration of leptosperin measured by immunochromatography was significantly correlated with the concentration measured by high-performance liquid chromatography (HPLC) or enzyme-linked immunosorbent assay (ELISA). Because the immunochromatographic method is rapid and reliable, it could be applied to on-site quality control or inspection of honey samples by a beekeeper, a manufacturer, an inspector, a retailer, or a consumer. (C) 2015 Elsevier Ltd. All rights reserved.

Myeloperoxidase (MPO)-generated halogenating molecules, such as hypochlorous acid and hypobromous acid (HOBr), in inflammatory regions are postulated to contribute to disease progression. In this study, we showed that ergothioneine (EGT), derived from an edible mushroom, inhibited MPO activity as well as the formation of 8-bromo-2-deoxyguanosine in vitro. The HOBr scavenging effect of EGT is higher than those of ascorbic acid and glutathione. We initially observed that the administration of Coprinus comatus, an edible mushroom containing a high amount of EGT, inhibited the UV-B-induced inflammatory responses and DNA halogenation, suggesting that EGT is a promising anti-inflammatory agent from mushrooms.

Myeloperoxidase is an inflammatory enzyme that generates reactive hypochlorous acid in the presence of hydrogen peroxide and chloride ion. However, this enzyme also uses bromide ion or thiocyanate as a substrate to form hypobromous or hypothiocyanous acid, respectively. These species play important roles in host defense against the invasion of microorganisms. In contrast, these enzyme products modify biomolecules in hosts during excess inflammation, indicating that the action of myeloperoxidase is both beneficial and harmful. Myeloperoxidase uses other endogenous compounds, such as serotonin, urate, and <span style="font-variant: small-caps;">l</span>-tyrosine, as substrates. This broad-range specificity may have some biological implications. Target molecules of this enzyme and its products vary, including low-molecular weight thiols, proteins, nucleic acids, and lipids. The modified products represent biomarkers of myeloperoxidase action. Moderate inhibition of this enzyme might be critical for the prevention/modulation of excess, uncontrolled inflammatory events. Some phytochemicals inhibit myeloperoxidase, which might explain the reductive effect caused by the intake of vegetables and fruits on cardiovascular diseases.

© 2015 Elsevier Inc. All rights reserved. At the sites of inflammation, hypohalous acids, such as hypochlorous acid and hypobromous acid (HOBr), are produced by myeloperoxidase. These hypohalous acids rapidly react with the primary amino groups to produce haloamines, which are relatively stable and can diffuse long distances and cross the plasma membrane. In this study, we examined the effects of taurine, the most abundant free amino acid in the leukocyte cytosol, on the hypohalous acid-dependent formation of 8-chloro-2′-deoxyguanosine (8-CldG) and 8-bromo-2′-deoxyguanosine (8-BrdG). The reaction of taurine with HOBr yielded taurine bromamine, which is the most stable among other bromamines of amino acids. Taurine also enhanced the bromination of only dG among the four 2′-deoxynucleosides, whereas it inhibited the 8-CldG formation. The specificity of taurine for the enhanced formation of halogenated dG is completely different from that of nicotine, an enhancer of chlorination. The amount of dibrominated taurine (taurine dibromamine) closely correlated with the formation of 8-BrdG, suggesting that taurine dibromamine might be a plausible mediator for the dG bromination in vivo.

【目的】シカ肉の食資源化のため，シカ肉中の機能性アミノ酸であるカルニチン量について検討している。これまでシカ肉は牛肉，豚肉，鶏肉よりもカルニチンを多く含み，加熱調理では真空調理により80℃の加熱温度の場合にカルニチン損失が少ないことを明らかにした。本研究では，シカ肉の嗜好性の改善として，多穀麹を添加し熟成させたシカ肉を用いて，重量減少率，カルニチン含有量と物性の変化について検討を行った。<br> 【方法】ニホンジカのもも肉を用い，シカ肉重量に対し1%の多穀麹を添加したものと無添加のものを10℃で24時間熟成させた。各試料は真空包装後，多穀麹添加試料は90℃で，無添加試料は80℃,90℃,100℃でスチーム加熱を30分間行った。加熱時におけるシカ肉中芯温度測定と加熱前後のシカ肉重量変化から重量減少率を求めた。高速液体クロマトグラフィータンデム質量分析計（LC-MS/MS）を用いカルニチン量の比較を行った。クリープメーターを用い破断測定を行った。<br> 【結果】各加熱温度におけるシカ肉最終芯温は80℃加熱で70℃，90℃加熱で85℃，100℃加熱で90℃を示した。シカ肉重量減少率は80℃加熱で14.1%，90℃加熱で26.5%,100℃加熱で32.7%，1%多穀麹添加90℃加熱で24.4%を示した。L-カルニチン・アセチルカルニチンは，80℃加熱が90℃および100℃加熱より高い値を示した。物性測定では80℃加熱が100℃加熱のシカ肉より破断応力が小さく軟らかいという結果を得た。多穀麹1％添加試料は無添加試料より破断応力は小さく軟らかくなる傾向を示したが，カルニチン量において無添加試料より低い値を示す傾向を示した。

Leptosperin, a novel glycoside of methyl syringate, is exclusively present in manuka honey derived from the Leptospermum species Leptospermum scoparium. Quantification of leptosperin might thus be applicable for authentication of honey. The concentration of leptosperin has high linearity with antibacterial activity. We established a monoclonal antibody to leptosperin and characterized the antibody in detail by a competitive enzyme-linked immunosorbent assay (ELISA), comparing the results with those of the high-performance liquid chromatography (HPLC) method for validation. The antigen in manuka honey was confirmed as leptosperin by HPLC fractionation with quantitation by an ELISA. Leptosperin contents of 50 honey samples were analyzed by an established ELISA, which can handle 20 samples (duplicate) on one 96-well plate. Significant coincidence with the chemical quantitation was observed. Immunochemical quantitation of leptosperin would be an economical and facile method for the possible authentication of manuka honey, allowing many honey samples to be processed and analyzed by an ELISA simultaneously.

Manuka honey, obtained from Leptospermum scoparium flowers in New Zealand, has strong antibacterial properties. In this study, plausible authentication of the manuka honey was inspected by measuring leptosperin, methyl syringate 4-O-beta-D-gentiobiose, along with methyl syringate. Despite a gradual decrease in methyl syringate content over 30 days at 50 degrees C, even at moderate 37 degrees C, leptosperin remained stable. A considerable correlation between nonperoxide antibacterial activity and leptosperin content was observed in 20 certified manuka honey samples. Leptosperin and methyl syringate in manuka honey and related products were analyzed using HPLC connected with mass spectrometry. One noncertified brand displayed significant variations in the leptosperin and methyl syringate contents between two samples obtained from different regions. Therefore, certification is clearly required to protect consumers from disguised and/or low-quality honey. Because leptosperin is stable during storage and specific to manuka honey, its measurement may be applicable for manuka honey authentication.

Scope: The effect of food combination on metabolic profile in postprandial plasma has hardly been reported. We investigated the absorption and metabolism of quercetin and soy isoflavones in humans after combination meal consumption. Methods and results: Five healthy volunteers ingested sauteed onion and tofu, and the plasma metabolites of quercetin and isoflavones were analyzed. Quercetin and genistein were incubated with human intestinal Caco-2 cells and human hepatoma HepG2 cells to further analyze the influence of simultaneous supply to the small intestine and the liver. Glucuronosyl conjugates of quercetin and methylated quercetin were the major plasma metabolites in the case of onion intake. Plasma metabolites with the single serving of tofu were both glucuronide and sulfate metabolites of isoflavones. Interestingly, quercetin sulfate was only detected after the combined intake of sauteed onion and tofu, accompanied with a decrease in sulfated isoflavones. Besides, quercetin was shown as the preferential substance for phase II enzymes over genistein in both Caco-2 and HepG2 cells. Conclusion: These results indicate that, when flavonoids and isoflavonoids were ingested together, the metabolic conversions in the small intestine and/or the liver could be altered, resulting in the variation of the postprandial profiles of the plasma metabolites.

【目的】揚げ物調理として，短時間で食品に油脂の風味が付加でき，アクリルアミド生成抑制効果がある真空フライ調理法が注目されている。本研究では，真空フライ調理と常圧フライ調理における野菜を用いたチップスの外観，食感，栄養成分，酸化度，抗酸化力の比較から，真空フライ調理法による野菜チップスの品質について検討した。 <br>【方法】試料はゴボウ，ショウガ，ニンジン，ピーマン，リンゴとした。真空フライ調理法は真空フライヤー（アトラステクノサービス製）を用い，真空圧は－0.090MPaから－0.097MPaとした。各試料で水分含量3％程度となるまでの加熱温度と加熱時間を設定した。真空フライ調理の加熱温度は78～90℃で加熱時間は25～45分，常圧フライ調理の加熱温度は150℃から170℃で加熱時間は20～35分とした。野菜チップスの顕微鏡観察，表面測色，破断測定，官能評価，脂質酸化度，抗酸化力，ビタミンC含量の測定を行った。<br>【結果】真空フライ調理法による野菜チップスの外観は，常圧フライに比べて油膜が少なく，褐変が少なく生に近い色調を保っていた。また，真空フライの方が，破断エネルギーは大きいが，官能評価では食感・香り・色が良いと評価された。真空フライ調理法による野菜チップスの方が，脂質酸化物が少なく，ビタミンC含量は有意に高値を示した。一方，抗酸化力は常圧フライ法の方が高く、常圧フライによるチップスは、色調が黒く、アミノカルボニル反応が進行し生じた褐変物質が抗酸化機能を示した可能性が推察された。

Copyright © 2014, Japanese Society for Food Science and Technology For the purpose of promoting the utilization of venison as a functional food, we investigated the carnitine content and physical properties of venison subjected to different sous-vide (cooking by steam heating under vacuum) processing temperatures. The present work aimed to quantify free carnitine and acylcarnitine using liquid chromatography-tandem mass spectrometry. Compared to meat samples heated at 100°C, those heated at 80°C showed a slow temperature increase and a low rate of weight loss. Moreover, heating at 80°C resulted in greater hydrophilic L-carnitine and low molecular weight acetylcarnitine than heating at 100°C. On the other hand, levels of hydrophobic hexanoylcarnitine, myristoylcarnitine, and palmitoylcarnitine were greater at 100°C. Determinations of physical properties showed that samples heated at 80°C were significantly tenderer than those heated at 100°C. Furthermore, sensory evaluation results showed that samples heated at 80°C scored highly in the following three attributes?toughness (palatability); umami; and overall palatability. Based on these results, we propose that heating at 80°C is more desirable with respect to functionality and palatability for promoting the utilization of venison as a functional food.

For the purpose of promoting the utilization of venison as a functional food, we investigated the carnitine content in cooked venison. The present work aimed to quantitate free carnitine and acylcarnitine in cooked venison using liquid chromatography-tandem mass spectrometry [LC-MS/MS]. As a result of heat treatment, hydrophilic Lcarnitine showed a tendency to decrease, and hydrophobic acylcarnitine showed a tendency to increase, except under frying. However, only the acetylcarnitine showed a tendency to decrease because it is a low molecular weight compound that is connected to a two-carbon fatty acid. We propose that steaming is the most effective of the heat treatment methods tested in the present work for promoting the utilization of venison as a functional food. Copyright © 2014, Japanese Society for Food Science and Technology.

Serotonin, 5-hydroxytryptamine, is a systemic bioactive amine that acts in the gut and brain. As a substrate of myeloperoxidase in vitro, serotonin is oxidized to tryptamine-4,5-dione (TD), which is highly reactive with thiols. In this work, we successively prepared a monoclonal antibody to quinone-modified proteins and found that the antibody preferentially recognizes the TD-thiol adduct. Using the antibody, we observed that the chloride ion, the predominant physiological substrate for myeloperoxidase in vivo, is not competitive toward the enzyme catalyzed serotonin oxidation process, suggesting that serotonin is a plausible physiological substrate for the enzyme in vivo. Immunocytochemical analyses revealed that TO staining was observed in the cytosol of SH-SY5Y neuroblastoma cells while blot analyses showed that some cellular proteins were preferentially modified. Pull-down analyses confirmed that the cytoskeletal proteins tubulins, vimentin, and neurofilament-L were modified. When pure tubulins were exposed to micromolar levels of synthetic TD, self-polymerization was initially enhanced and then suppressed. These results suggest that serotonin oxidation by myeloperoxidase or the action of other oxidants could cause functional alteration of cellular proteins, which may be related to neurodegeneration processes or irritable bowel syndrome. (C) 2014 The Authors. Published by Elsevier B.V.

野生シカ肉の有効活用を目的として，オスジカ，メスジカの肉重量および栄養成分の差異について検討した。試料は，兵庫県丹波地域において2010年9月，11月，12月に捕獲したニホンジカを使用した。オスジカの平均体重は46.4 kg，肉重量は16.7 kg，歩留率は35.6％，栄養成分は100 gあたりタンパク質21.2 g，脂質0.4 gを示した。メスジカの平均体重は36.3 kg，肉重量は13.1 kg，歩留率は35.7％，タンパク質20.5 g，脂質0.7 gを示した。メスジカは，オスジカより小さいが，肉の歩留率は同等で，脂質量は増加傾向にあった。オスジカ，メスジカとも捕獲月による肉重量および栄養成分値の差異は小さかった。肉の部位別では，オスジカ，メスジカともモモとスネの重量割合が高く，肉の部位間での栄養的特徴の違いは小さかった。

本研究では，シカ肉の食資源としての有効利用を目的として，近年，機能性アミノ酸として注目されているカルニチンについての定性および定量的な検討を行った．<BR>カルニチンには，遊離カルニチンと脂肪酸が結合したアシルカルニチンとしても存在するが，アシル体の多くが微量で，今までの酵素法などを用いた測定法ではそれぞれを分別して検出することができなかった．本研究では，LC-MS/MSを用いることによって，遊離カルニチンおよびアシルカルニチンであるアセチルカルニチン，ヘキサノイルカルニチン，ミリストイルカルニチン，パルミトイルカルニチンの5種類をシカ肉から検出することができた．さらに，シカ肉と牛肉，豚肉，鶏肉とのカルニチン量の比較を行った結果，遊離およびアシルカルニチンのいずれにおいてもシカ肉と牛肉に多く含まれていた．遊離カルニチンと短・中鎖脂肪酸が結合したカルニチンにおいては，シカ肉に多く含まれており，長鎖脂肪酸が結合したカルニチンは，シカ肉よりも牛肉に多く含まれていた．本研究の測定方法を用いることによって，遊離カルニチンに加えてアシルカルニチン類をシカ肉から検出することが可能となった．さらに，シカ肉に遊離カルニチン，アセチルカルニチンが多く含まれることが示され，脳機能向上などの機能性が期待されるアセチルカルニチンを多く含むことから，シカ肉の機能性食品としての可能性を見出すことができた．

Sapovirus (SaV), a member of the family Caliciviridae, is an important cause of acute epidemic gastroenteritis in humans. Human SaV is genetically and antigenically diverse and can be classified into four genogroups (GI, GII, GIV, and GV) and 16 genotypes (7 GI [GI.1-7], 7 GII, [GII.1-7], 1 GIV and 1 GV), based on capsid sequence similarities. Monoclonal antibodies (MAbs) are powerful tools for examining viruses and proteins. PAI myeloma cells were fused with spleen cells from mice immunized with a single type of recombinant human SaV virus-like particles (VLPs) (GI.1, GI.5, GI.6, GII.3, GIV, or GV). Sixty-five hybrid clones producing MAbs were obtained. Twenty-four MAbs were characterized by ELISA, according to their cross-reactivity to each VLP (GI.1, GI.5, GI.6, GII.2, GII.3, GII.4, GII.7, GIV, and GV). The MAbs were classified by this method into: (i) MAbs broadly cross-reactive to all GI, GII, GIV and GV strains; (ii) those reactive in a genogroup-specific; and (iii) those reactive in a genotype-specific manner. Further analysis of three broadly cross-reactive MAbs with a competitive ELISA demonstrated that at least two different common epitopes are located on the capsid protein of human SaVs in the four genogroups. The MAbs generated and characterized in this study will be useful tools for further study of the antigenic and structural topography of the human SaV virion and for developing new diagnostic assays for human SaV. © 2012 The Societies and Wiley Publishing Asia Pty Ltd.

Background. Photoageing of skin is thought to be caused by protein denaturation, which can be induced by ultraviolet radiation. Previous studies have also reported that inflammation is related to protein denaturation; however, the influence of inflammation on skin ageing has not been explored in detail. Aim. To investigate the possible connection between inflammation and protein denaturation, which might lead to skin ageing, we focused on halogenated tyrosine as a denatured substance produced during the inflammation process. Methods. We measured halogenated tyrosine in aged human skin. Inflammatory cells and halogenated tyrosine were detected by immunohistochemistry using antibodies to mast-cell tryptase, neutrophilic myeloperoxidase and halogenated tyrosine. Finally, using elastic van Gieson ( EVG) staining, we investigated whether the sites of halogenated tyrosine coincided with the sites at which proteins were denatured. Results. Immunohistochemical analysis indicated that both inflammatory cells and halogenated tyrosines increased with ageing in both photoexposed and photoprotected skin. EVG staining confirmed that the localization of halogenated tyrosine was close to the sites at which protein was denatured. Conclusions. Our investigations indicate a possible connection between skin ageing and inflammation, suggesting that halogenated tyrosine could be a useful marker of ageing skin.

For immunological experiments on glycogens, anti-glycogen antibodies are indispensable to capture, detect, and visualize sugar molecules. An anti-glycogen monoclonal antibody (IV58B6) and newly constructed antibody (ESG1A9mAb) have a common immunoglobulin type (IgM) and binding ability to glycogens, but overall possess different binding features. Therefore, they may prove useful for the construction of an advanced system of quantitative ELISA based on their molecular structures. For this purpose, detailed information on the carbohydrate-specificities of ESG1A9mAb and IV58B6 is first required, but their fine specificities for various types of glycogens have not been elucidated. To overcome this problem, we performed interaction analysis by ELISA of ESG1A9mAb and IV58B6 toward 15 glucose polymers, that is, 5 enzymatically-synthesized glycogens (ESGs), 6 natural source glycogens (NSGs), 3 enzymatically digested glycogens (EDGs), and soluble starch. To provide a more detailed analysis, we determined the association constants (K-a) of the two antibodies toward these glycogens by surface plasmon resonance. The results indicated that the carbohydrate-binding properties toward NSGs of ESG1A9mAb and IV58B6 were similar, but markedly differed for ESGs and EDGs. ESG1A9mAb showed significant affinity for all the ESGs and NSGs tested, whereas IV58B6 had only slight affinity for ESGs, although the affinities were increased when the ESGs were enzymatically digested. This information should be helpful for the design of both in vitro and in vivo immunological assays. (C) 2012 Elsevier Ltd. All rights reserved.