Flavor assessment of a lactic fermented vinegar described in Japanese books from the Edo period (1603-1867).

Aims: Rice vinegar is a traditional fermented seasoning in Japan, and its production remained unchanged for over 800 years until the Edo period. However, based on the available information regarding rice vinegar production methods from this period and the results of reproduction experiments, we speculated that unlike the modern-day acetic fermented vinegar, rice vinegar produced during the Edo period was lactic fermented. Main methods: To verify this assumption, we analyzed the flavor components of Honcho, a lactic fermented product prepared using a method described in books, including "Honchoshokkan" from the Edo period, by capillary electrophoresis/time-of-flight mass spectrometry, high-performance liquid chromatography, gas chromatography mass spectrometry, and taste sensor analysis. Sensory evaluation was also conducted to assess validation as a seasoning. Results: Honcho contains 2 % lactic acid, which gives it its acidity, and small amounts of other nonvolatile acids, but significantly lower levels of acetic acid (0.188 ± 0.015 g/100 mL, p < 0.01). It contains more than double the free amino acids of Kurozu, a modern rice vinegar, and more glutamic acid. Boiling to remove ethanol from yeast fermentation concentrated the free amino acids 1.5 times. Sensor taste analysis showed Honcho had weaker acidity but stronger umami taste than commercial rice vinegar. The volatile compounds related to acetic acid fermentation were significantly different between Honcho and Kurozu. Boiling increased Honcho's acidity, mainly through non-volatile acids. Significance: These findings provide evidence to indicate that Honcho was an acidic seasoning for heat-cooking, which is uncommon in Japanese cuisine today and is mentioned in Edo period books. This seasoning contains many amino acids, implying that it adds umami flavor, not only the sourness of modern vinegar.

Lactic acid induces HSPA1A expression through ERK1/2 activation.

Heat shock protein (HSP) A1A protects cells from various stressors. The concentrated liquid of the traditional Japanese rice black vinegar Kurozu increased HSPA1A expression in normal rat liver RLN-10 cells. Lactic acid, the primary component of concentrated Kurozu, induced HSPA1A expression in a concentration-dependent manner. Induction with 4 m m lactic acid increased HSPA1A expression by three times compared with that in the absence of lactic acid. The induction was inhibited by staurosporine or a selective MEK1/2 inhibitor (SL327). The phosphorylation of ERK1/2 was increased by lactic acid. These results suggest that lactic acid induces HSPA1A expression by activating ERK1/2. As well as lactate, 3,5-dihydroxybenzoic acid (DHBA), a ligand for G protein-coupled receptor 81 (GPR81), also induced HSPA1A at lower concentrations than lactate. The increased effect of DHBA on HSPA1A expression as compared with lactate may be related to the higher affinity of DHBA for GPR81 than of lactate.

The Open Clinical Tests for the Safety of Kurozu-containing Food / 日本補完代替医療学会誌

Two different types of clinical safety test of Kurozu-containing food were done. Under the informed consent, following tests were conducted, Test 1; Ten healthy adults were orally administrated 10 capslues/day of Kurozu for 4 weeks, Test 2; Ten patients with hepatitis C and 10 patients with mild hepatic dysfunction were orally administrated 2 capslues/day of Krozu for 4 weeks. In both the Test1 and Test2, Vital sign including Blood pressure etc, blood cell count, blood chemical analysis, and urinalysis were performed. In both tests, there were no abnormal findings. Furthermore, no side effect was observed. In conclusion, safety of Kurozu-containing food was confirmed.

Effects of Dietary Supplement Containing Kurozu Concentrate on Visceral Fat Accumulation / 日本補完代替医療学会誌

Objective: The present study was done to examine the effect of dietary supplement containing Kurozu concentrate on human visceral fat accumulation. Material and Methods: Sixteen subjects(BMI≧25) were orally administrated Kurozu concentrate for 8 weeks(low dose group: 500 mg/day, high dose group 1000 mg/day). Before and at weeks 8, hematological analysis, urinalysis and CT-scan to estimate the visceral fat accumulation were conducted. Results: In high dose group, following results were obtained; visceral fat decreased significantly(p=0.033), HDL-cholesterol level elevated significantly (p=0.034) and systolic blood pressure decreased significantly(p=0.004). In addition, no serious symptoms or abnormal hematological values were observed in both group.Conclusions: The dietary supplement containing Kurozu concentrate may be useful to prevent a metabolic syndrome.

Chiral amino acid analysis of Japanese traditional Kurozu and the developmental changes during earthenware jar fermentation processes.

Enantioselective amino acid metabolome analysis of the Japanese traditional black vinegars (amber rice vinegar, Kurozu) was performed using two-dimensional high-performance liquid chromatography combining a microbore-monolithic ODS column and narrowbore-enantioselective columns. d-Amino acids, the enantiomers of widely observed l-amino acids, are currently paid attention as novel physiologically active substances, and the foodstuffs and beverages containing high amounts of d-amino acids are the subjects of interest. In the present study, the amino acid enantiomers were determined by two-dimensional HPLC techniques after pre-column fluorescence derivatization with 4-fluoro-7-nitro-2,1,3-benzoxadiazole. In the first dimension, the amino acid enantiomers are separated as their d plus l mixtures by the reversed-phase mode, then the d-amino acids and their l-counterparts are separately determined in the second dimension by the enantioselective columns. As a result, large amounts of d-Ala (800-4000nmol/mL), d-Asp (200-400nmol/mL) and d-Glu (150-500nmol/mL) were observed in some of the traditionally produced Kurozu vinegars. Relatively large or small amounts of d-Ser (50-100nmol/mL), d-Leu (10-50nmol/mL) and d-allo-Ile (less than 20nmol/mL) were also present in these samples. Developmental changes in the d-amino acid amounts during the fermentation and aging processes have also been investigated.

Effects of a Dietary Supplement Containing Kurozu Concentrate on Body Fat and Energy Metabolism / 日本補完代替医療学会誌

<b>Objectives:</b> Kurozu has long been used as a traditional fermented food product mainly in East Asia, including Japan; various health functions of Kurozu have been reported. This study examined its effects on body fat and energy metabolism by a double-blind placebo-controlled trial.<br> <b>Methods:</b> Mildly obese adults took a dietary supplement containing Kurozu concentrate for 12 weeks with approximately 10 min of exercise every day.<br> <b>Results:</b> A decline in hip circumference was observed in the group consuming the dietary supplement containing Kurozu. It was shown that fat-derived energy consumption increased while suppressing carbohydrate-derived energy consumption. Therefore, it was revealed that energy was derived from the fat consumed, and a tendency to increase the energy consumption at the time of exercise was also observed. Furthermore, negative correlations were observed between the initial fat volume and both the variation of the total fat area and the variation of the subcutaneous fat area in the examination group. It was revealed that larger the amount of body fat, greater the effect of Kurozu in decreasing the amount of fat. In addition, no safety issues were observed 12 weeks after ingesting the dietary supplement containing Kurozu concentrate, and it was confirmed as a safe food product.<br> <b>Conclusion:</b> Appropriate exercise and intake of the dietary supplement containing Kurozu concentrate contributed toward the improvement of the metabolic syndrome.<br>

<b>Effects of Dietary Supplement Containing Kurozu Concentrate on Blood Pressure, QOL and TCM Constitution</b> / 日本補完代替医療学会誌

<b>Objectives</b>: The present study was examined the effects of dietary supplement containing Kurozu concentrate on Blood Pressure, QOL and Traditional Chinese Medicine (TCM) Constitution. <b>Methods</b>: Forty subjects with slightly high blood pressure were assigned randomly to 2 groups (active group and placebo group). Kurozu-containing foods or the placebo was administered to the subjects daily for 12 weeks. At week 0, 6 and 12, blood samples were collected, physical measurements were taken, and the quality of life (SF-36v2) and TCM constitution were examined. <b>Results</b>: Significant differences on blood pressure and TCM constitution between the groups were observed at week 12. The change of QOL between the groups was observed significantly at week 6. <b>Conclusion</b>: These findings suggest that foods containing Kurozu have hypotensive action and improve effects on constitutional problems. In addition, it was confirmed that foods containing Kurozu are safe to ingest.

Protective effects of fermented rice vinegar sediment (Kurozu moromimatsu) in a diethylnitrosamine-induced hepatocellular carcinoma animal model.

Kurozu moromimatsu is the sediment of Kurozu, a jar-fermented Japanese black vinegar produced from unpolished rice. Here, we examined the protective effects of Kurozu moromimatsu in a diethylnitrosamine-induced model of hepatocellular carcinoma. Thirty-two F344 rats were divided into two groups; the control group received basal CE-2 diet, and the Kurozu moromimatsu group received CE-2 diet containing Kurozu moromimatsu. At 16 weeks after initial intraperitoneal administration of diethylnitrosamine (150 mg/kg/week), serum was collected from half the rats. These rats were sacrificed and the liver was resected for histological examination of hematoxylin-eosin-stained sections and assay of matrix metalloproteinase-2 and matrix metalloproteinase-9 levels in tumor tissues. Glutathione S-transferase placental form-positive foci were evaluated by immunostaining for glutathione S-transferase placental form. The remaining rats were maintained for evaluation of survival. There were no significant differences of serum transaminases, tumor necrosis factor-alpha, and also no marked hepatic histological differences, between the two groups. However, the size of hepatocellular carcinomas was greatly decreased and the levels of activated matrix metalloproteinase-2 and -9 were significantly reduced in the Kurozu moromimatsu group. Further, survival was significantly prolonged in the Kurozu moromimatsu group compared with the control. These results indicate that Kurozu moromimatsu inhibited the growth of hepatocellular carcinoma.