Masahiro Koyama

Phenolic compounds responsible for the superoxide dismutase-like activity in high-Brix apple vinegar.

High-Brix apple vinegar (HBAV) with palatable drinking qualities has been developed using a greater amount of apple ingredients. In HBAV and in regular apple vinegar (RAV), constituents of 4 kinds of organic acids, 20 kinds of amino acids, 3 kinds of sugars, 4 kinds of minerals, and phenols were determined. These constituents, except for acetic acid, in HBAV are of higher abundance than in RAV. HBAV had a 7.1 times greater superoxide dismutase (SOD)-like activity compared with RAV. Those constituents, except for phenols, had very low SOD-like activity, and total phenol levels in HBAV were comparable to 181 mg of gallic acid equivalents/100 mL, which was 6.0 times more abundant than in RAV. Nine kinds of phenols including two kinds of hydroxycinnamates, two kinds of hydroxybenzoates, and five kinds of hydroxycinnamoyl quinates, originating from raw material were determined, but there were no ascorbic acid and flavonoids in HBAV. Chlorogenic acid, 4-p-coumaroylquinic acid, and caffeic acid were the three major phenols, and their content levels were 19.6, 13.5, and 0.76 mg in 100 mL of HBAV, respectively. Sum of contents of chlorogenic acid and the isomers was 24.0 mg/100 mL, and the percentage was 56.9% in the total identified phenols in HBAV. In RAV, only chlorogenic acid was determined as phenols, and the content was 3.1 mg/100 mL. SOD-like activities of the constituents of HBAV were obtained through high-accuracy assays using vinegar reconstitutions, and each contribution to the total SOD-like activity was found. As a result, 77.2% for all SOD-like activity of HBAV was reconstituted using the determined nine phenols and other constituents. Chlorogenic acids were the most effective, and the contribution to the total activity was 41.7%. The most abundant phenols, chlorogenic acids, were the most important contributors to the SOD-like activity. These SOD-active phenols originated from raw material and remained through the acetic acid fermentation processes. In the fermentation process of HBAV, the active constituents were well maintained, providing an advantage in the production of a phenol-rich product.

Purification and Identification of Antihypertensive Peptides from Fermented Buckwheat Sprouts

Buckwheat (Fagopyrum esculentum) is rich in antihypertensive compounds. This study investigated the effect of lactic-fermented buckwheat sprouts (neo-FBS) on level, identification, and potency of blood pressure-lowering (BPL) compounds. A single oral dose of 1.0 mg/kg body weight buckwheat sprouts (BS) in spontaneously hypertensive rats did not show significant BPL activity, whereas neo-FBS significantly decreased blood pressure. HPLC of neo-FBS identified two peaks absent in the profile of BS. The peak exhibiting potent BPL activity was fractionated, and six peptides (DVWY, FDART, FQ, VAE, VVG, and WTFR) and tyrosine were identified by LC-MS/MS and Edman degradation. Single oral dose administration of the peptides revealed significant BPL effect of all the peptides, with the most potent being DVWY, FQ, and VVG. DVWY, VAE, and WTFR are novel. This study demonstrates that lactic fermentation of BS produces new, highly potent antihypertensive peptides and increases active compounds GABA and tyrosine already present in BS.

Fermentative l-(+)-lactic acid production from non-sterilized rice washing drainage containing rice bran by a newly isolated lactic acid bacteria without any additions of nutrients

A newly isolated lactic acid producing bacterium Lactobacillus rhamnosus M-23, from a rice washing drainage storage tank was found to produce l-(+)-lactic acid from a non-sterilized mixture of rice washing drainage and rice bran without any additions of nutrients under the simultaneous saccharification and fermentation (SSF) process. The maximum lactate yield attained was 59 g/l with a productivity of 1.23 g/l/h and a product optical purity of 95% corresponding to a conversion of 0.85 g of lactic acid per gram of sugar equivalent.

Blood pressure-lowering peptides from neo-fermented buckwheat sprouts: a new approach to estimating ACE-inhibitory activity.

Neo-fermented buckwheat sprouts (neo-FBS) contain angiotensin-converting enzyme (ACE) inhibitors and vasodilators with blood pressure-lowering (BPL) properties in spontaneously hypertensive rats (SHRs). In this study, we investigated antihypertensive mechanisms of six BPL peptides isolated from neo-FBS (FBPs) by a vasorelaxation assay and conventional in vitro, in vivo, and a new ex vivo ACE inhibitory assays. Some FBPs demonstrated moderate endothelium-dependent vasorelaxation in SHR thoracic aorta and all FBPs mildly inhibited ACE in vitro. Orally administered FBPs strongly inhibited ACE in SHR tissues. To investigate detailed ACE-inhibitory mechanism of FBPs in living body tissues, we performed the ex vivo assay by using endothelium-denuded thoracic aorta rings isolated from SHRs, which demonstrated that FBPs at low concentration effectively inhibited ACE in thoracic aorta tissue and suppressed angiotensin II-mediated vasoconstriction directly associated with BPL. These results indicate that the main BPL mechanism of FBP was ACE inhibition in living body tissues, suggesting that high FBPs bioavailability including absorption, tissue affinity, and tissue accumulation was responsible for the superior ACE inhibition in vivo. We propose that our ex vivo assay is an efficient and reliable method for evaluating ACE-inhibitory mechanism responsible for BPL activity in vivo.

The Restorative Effects of Eucommia ulmoides Oliver Leaf Extract on Vascular Function in Spontaneously Hypertensive Rats

Eucommia ulmoides Oliv. leaf is a traditional Chinese antihypertensive and antidiabetic medicine. We examined the effects of chronic Eucommia leaf extract (ELE) administration on artery function and morphology in spontaneously hypertensive rats (SHRs). ELE was orally administered via normal diet ad libitum to six-week-old male SHRs at a concentration of 5% for seven weeks. Acetylcholine (ACh)-induced endothelium-dependent relaxation, sodium nitroprusside (SNP)-induced endothelium-independent relaxation, plasma nitric oxide (NO) levels, and media thickness were assessed. ELE significantly improved ACh-induced aortic endothelium-dependent relaxation but did not affect SNP-induced endothelium-independent relaxation in the SHRs, as compared to the animals receiving normal diet. Plasma NO levels and media thickness were significantly increased and decreased, respectively, in the ELE-treated SHRs. Therefore, long-term ELE administration may effectively improve vascular function by increasing plasma NO levels and bioavailability, and by preventing vascular hypertrophy in the SHR aorta.

Preventive effect of Eucommia leaf extract on aortic media hypertrophy in Wistar-Kyoto rats fed a high-fat diet

Eucommia ulmoides Oliver leaf extract (ELE) has been shown to have anti-hypertensive and anti-obesity effects in rats that are fed a high-fat diet (HFD). To explore the effects of chronic administration of ELE on body weight, blood pressure and aortic media thickness, 7-week-old male Wistar-Kyoto (WKY) rats were orally administered a normal diet, a 30% HFD, or a 5% ELE plus HFD ad libitum for 10 weeks. The HFD treatment caused mild obesity and hypertension in the normotensive rats, while rats receiving both ELE and the HFD had significantly lower body weights, less visceral and perirenal fat, lower blood pressure and thinner aortic media than the control rats receiving the HFD only. The plasma adiponectin/leptin ratio also improved in ELE-treated rats. Although plasma leptin levels were elevated in all HFD rats, adiponectin levels increased only in the ELE-treated rats. Anti-hypertensive and anti-obesity effects may be caused by the geniposidic acid (GEA) and/or asperuloside present in ELE. These findings suggest that chronic ELE administration prevents aortic media hypertrophy in early-stage obesity with hypertension. Long-term administration of ELE might inhibit the development of arteriosclerosis.

Extracellular protease derived from lactic acid bacteria stimulates the fermentative lactic acid production from the by-products of rice as a biomass refinery function

A lactic acid producing bacterium, Lactobacillus rhamnosus M-23, newly isolated from a rice washing drainage storage tank was found to produce l-(+)-lactic acid from a non-sterilized mixture of rice washing drainage and rice bran without any additions of nutrients under the simultaneous saccharification and fermentation (SSF) process. This strain has the ability to utilize the non-sterilized rice washing drainage and rice bran as a source of carbohydrate, saccharifying enzymes and nutrients for lactic acid production. Observation of extracellular protease activity in SSF culture broth showed that a higher protease activity was present in strain M-23 than in other isolated lactic acid producing bacteria (LABs). To investigate the structural changes of solid particles of rice washing drainage throughout LAB cultivation, scanning electron microscopic (SEM) observation and Fourier transform infrared-spectroscopy (FT-IR) analysis were performed. The results of the SEM observation showed that the surface material could be removed from solid particles of rice washing drainage treated by culture broth (supernatant) of strain M-23, thus exposing the crystal structure of the starch particle surface. The results of the FT-IR analysis revealed that the specific transmittance decrease of the CC and CO stretching and OH group of the solid particles of the rice washing drainage were highly correlated with the produced lactic acid concentration and extracellular protease activity, respectively. These results demonstrate the high lactic acid producing ability of strain M-23 from a non-sterilized mixture of rice washing drainage and rice bran under the SSF condition due to the removal of proteinaceous material and exposure of the starch particle surface by extracellular protease.

Antihypertensive effects of orally administered eggplant (Solanum melongena) rich in acetylcholine on spontaneously hypertensive rats

Our previous results (Nakamura et al., 2013, 2016) indicated that acetylcholine (ACh) in orally administered foods exerts antihypertensive effects. Eggplants (Solanum melongena) contain abundant ACh (Horiuchi et al., 2003), and their food functionality was discovered, using spontaneously hypertensive rats, by measuring blood pressure after oral administration of a suspension of lyophilized eggplant powder. We found that lyophilized eggplant powder induced significantly lowered acute and chronic blood pressure levels at very low doses of 0.0650 mg/kg body weight (b.w.) and 0.821 mg/(kg b.w.·day), respectively. Chronic administration suppressed adrenaline and noradrenaline excretion in the urine, and aorta assays showed that eggplant acted on the M3 muscarinic ACh receptor (M3 mAChR). ACh was conclusively shown to function as the main component of eggplant contributing to antihypertensive activity by suppressing sympathetic nervous activity via M3 mAChR. This report reveals a new food functionality of eggplant and its potential as a novel antihypertensive food.

Decreased Fatty Acid β -Oxidation Is the Main Cause of Fatty Liver Induced by Polyunsaturated Fatty Acid Deficiency in Mice

Insufficient intake of polyunsaturated fatty acids (PUFA) causes fatty liver. The mechanism responsible is primarily related to increased lipogenesis and decreased FA degradation based on rodent studies. However, these studies were limited by the fact that the typical PUFA-deficient diets contained insufficient amounts of long-chain FA, the PUFA-containing diets were primarily composed of n-3 PUFA-enriched oil, and the intake of PUFA was excessive compared with the physiological requirement. To address these issues, mice were fed a PUFA-deficient diet containing long-chain FA at a standard fed level and then were orally fed a n-3/n-6-balanced PUFA-containing oil [PUFA (+)] or a PUFA-deficient oil [PUFA (-)] at physiological relevant levels (0.1 mL/mouse/2d). We compared these groups and examined whether fatty liver in PUFA deficiency was attributable to both the effects of increased lipogenesis and decreased FA catabolism. Compared with the PUFA (+) group, the PUFA (-) group showed increases in liver triglyceride and serum FA content. Hepatic gene expression of several mitochondrial β-oxidation enzymes, the serum 3-hydroxybutyrate level, and DNA-binding ability of peroxisome proliferator-activated receptor α (PPARα) were increased in the PUFA (+) group, whereas these adaptive responses were significantly attenuated in the PUFA (-) group. The hepatic expression of typical lipogenesis genes did not differ between the groups. Therefore, fatty liver in PUFA deficiency is attributable to suppression of the FA-degrading system probably from decreased PPARα adaptive responsiveness, and PUFA may be an essential factor for PPARα functioning. This finding is helpful for managing clinical situations having a risk of PUFA deficiency.

Fermentation-induced changes in the concentrations of organic acids, amino acids, sugars, and minerals and superoxide dismutase-like activity in tomato vinegar

ABSTRACT Various raw materials are used to produce vinegars that contain functional compounds associated with disease prevention. We evaluated changes in functional compounds during tomato vinegar production and superoxide dismutase-like activity of tomato vinegar. Tomato vinegar contained abundant anti-hypertensive compounds, e.g., γ-aminobutyric acid and potassium derived from tomatoes and acetic acid and pyroglutamic acid produced during fermentation. It had stronger superoxide dismutase-like activity than commercial vinegars because of tomato-derived superoxide dismutase-like compounds, including phenolic acids, flavonoids, and glutathione. These data indicate that tomato vinegar is a candidate dietary supplement with potential preventive effects against cardiovascular diseases due to its anti-hypertensive and superoxide dismutase-like compounds.