Rice Koji Extract Research Articles

Preventive Roles of Rice-koji Extracts and Ergothioneine on Anxiety- and Pain-like Responses under Psychophysical Stress Conditions in Male Mice.

This study determined the effect of daily administration of Rice-koji on anxiety and nociception in mice subjected to repeated forced swim stress (FST). In a parallel experiment, it was determined whether ergothioneine (EGT) contained in Rice-koji displayed similar effects. Anxiety and nociception were assessed behaviorally using multiple procedures. c-Fos and FosB immunoreactivities were quantified to assess the effect of both treatments on neural responses in the paraventricular nucleus of the hypothalamus (PVN), nucleus raphe magnus (NRM), and lumbar spinal dorsal horn (DH). FST increased anxiety- and pain-like behaviors in the hindpaw. Rice-koji or EGT significantly prevented these behaviors after FST. In the absence of formalin, both treatments prevented decreased FosB expressions in the PVN after FST, while no effect was seen in the NRM and DH. In the presence of formalin, both treatments prevented changes in c-Fos and FosB expressions in all areas in FST mice. Further, in vitro experiments using SH-SY5Y cells were conducted. Rice-koji and EGT did not affect cell viability but changed the level of brain-derived neurotrophic factor. In conclusion, Rice-koji could reduce anxiety and pain associated with psychophysical stress, possibly mediated by the modulatory effects of EGT on neural functions in the brain.

Formation of taste-active pyroglutamyl peptide ethyl esters in sake by rice koji peptidases

Formation of taste-active pyroglutamyl (pGlu) peptide ethyl esters in sake was investigated: 2 enzymes (A and B) responsible for the esterification were purified from a rice koji extract. MADLI-TOF/TOF analysis after deglycosylation identified enzyme (A) as peptidase S28 (GenBank accession number OOO13707.1) and enzyme (B) as serine-type carboxypeptidase (accession number AO090010000534). Both enzymes hydrolyzed pGlu peptides and formed ethyl esters under sake mash conditions: acidic pH (3-4) and in ethanol (5%-20% v/v) aqueous solutions. Enzyme (A) formed pGlu penta-peptide ethyl esters from pGlu undeca-peptides by a prolyl endo-type reaction. Enzyme (B) formed (pGlu) deca-peptide and its ethyl esters from pGlu undeca-peptides in an exo-type reaction. We are the first to report the enzymatic ethyl esterification reaction in the formation of pGlu peptides by rice koji peptidases.

Identification of Rice Koji Extract Components that Increase β-Glucocerebrosidase Levels in Human Epidermal Keratinocytes.

Rice miso contains many ingredients derived from rice koji and has been a valuable source of nutrition since ancient times. We found that the consumption of rice miso led to improvements in the moisture content of cheek stratum corneum, skin viscoelasticity, and skin texture. Further, rice miso extract was found to increase the mRNA expression and activity of β-glucocerebrosidase (β-GCase), an enzyme involved in ceramide synthesis in the stratum corneum, in cultures. In this study, we identified the lipid-derived components of rice koji that increase the β-GCase activity in cultured human epidermal keratinocytes. The methanol fraction of rice koji extract induced an increase in the mRNA expression and activity of β-GCase in keratinocytes. The active fraction of rice koji was found to contain phosphatidic acid (PA) and lysophosphatidic acid (LPA). The total PA concentration in rice koji was 973.9 ng/mg dry weight, which was 17.5 times higher than that in steamed rice. Among the molecular species, PA_18:2/18:2 was the most frequently found. The total LPA concentration in rice koji was 29.6 ng/mg dry weight, and 2-LPA_18:2 was the most frequently found LPA. Since PA and LPA increase the mRNA expression and activity of β-GCase in keratinocytes, they are thought to be the active ingredients in rice koji that increase the β-GCase levels in human epidermal keratinocytes.

Correction to Rice Koji Extract Enhances Lipid Metabolism through Peroxisome Proliferator-Activated Receptor Alpha (PPARα) Activation in Mouse Liver.

Correction to Rice Koji Extract Enhances Lipid Metabolism through Peroxisome Proliferator-Activated Receptor Alpha (PPARα) Activation in Mouse Liver.

Rice Koji Extract Enhances Lipid Metabolism through Proliferator-Activated Receptor Alpha (PPARα) Activation in Mouse Liver.

Koji is made from grains fermented with Aspergillus oryzae and is essential for the production of many traditional Japanese foods. Many previous studies have shown that koji contributes to the improvement of dyslipidemia. However, little is known regarding the underlying mechanism of this effect. Furthermore, the compound contributing to the activation of lipid metabolism is unknown. We demonstrated that rice koji extract (RKE) induces the mRNA expression of peroxisome proliferator-activated receptor alpha (PPARα) target genes, which promotes lipid metabolism in murine hepatocytes. This effect was not observed in PPARα-KO hepatocytes. We also demonstrated that RKE contained linolenic acid (LIA), oleic acid (OA), and hydroxyoctadecadienoic acids (HODEs), which activate PPARα, using LC-MS analysis. Our findings suggest that RKE, containing LIA, OA, and HODEs, could be valuable in improving dyslipidemia via PPARα activation.

Extension of Chronological Life in <i>Saccharomyces cerevisiae</i> under Ethanol Stress by Thermally Processed Rice <i>Koji</i> Extracts

The effect of thermally processed rice koji extracts on survival of yeast Saccharomyces cerevisiae was examined in comparison with non-heated koji extract. In chronological life span (CLS) tests on high-sugar fermentation, the survivals of the yeast cells grown with heated koji extracts were higher than non-heated koji extract. Heat treatment of the extracts by autoclaving led to a loss in most of amino acids due to the Maillard reaction, although histidine contents slightly increased. In glucose-arginine mixtures, arginine was partly converted to histidine by autoclaving and the addition of histidine prolonged the CLS of yeast cells. The yeast cells grown with the non-heated extracts were more resistant to oxidative stress whereas the antioxidant activities were lower than those of the heated extracts. The yeast cells grown with the heated extracts were more tolerant to ethanol and had a higher reduction capacity in the late stationary phase when the cells were incubated in the presence of ethanol. Maillard reaction products elevated the levels of reactive oxygen species to yeast cells grown under ethanol stress in the late stationary phase. These results suggest that thermally processed koji extracts can act as a protectant against ethanol stress during the late stationary phase of yeast growth and extend the CLS due to the increase of histidine contents by autoclaving.

A novel method for measurement of α-amylase activity in rice koji by capillary isoelectric focusing

A capillary isoelectric focusing (CIEF) method has been developed for the analysis of α-amylase activity in rice koji. The standards of α-amylase and rice koji extracts were successfully separated by CIEF in a large pH gradient. The α-amylase from A. oryzae was found to have pI of 3.7, as determined from the migration times of six standard proteins. A simple method has been developed for the measurement of α-amylase activity in rice koji, and the results were compared with those obtained by the official method. The coefficient of variation (C.V.) of the α-amylase assay was less than 0.5% in both intra-day and inter-day experiments. This CIEF method can be applied to the routine quality control of α-amylase activity in rice koji for the brewing process in sake production.

米麹中の糖化力の新規測定法

A new method for measuring glucose-forming activity in rice koji is proposed. In this method, G 2-PNP (4-nitrophenyl O-α-D glucopyranosyl-(1-4)-β-D- glucopyranoside) is used as the substrate. Because both glucoamylase and a-glucosidase can hydrolyze the substrate, the results showed the sum total of these enzymes activities (that is, glucose-forming activity) in rice koji extract. The substrateis hydrolyzed by these enzymes and the resulted product (PNP) is measured to determine the amount of glucose-forming activity quantitatively. This method has the following advantages:(i) The presence of glucose in the sample does not influence the results of the measurement, and therefore there is no need to dialyze the rice koji extract prior to analysis.(ii)α-Amylase of Aspergillus oryzae in the sample does not affect the results of measurement because if cannot hydrolyze the substrate.(iii) The assay procedure is simple, and can be easily accomplished in a short time. The proposed method showed good correlation with the method generally used for glucoamylase assay when purified glucoamylase is used as a sample. The averages of recovery of glucoamylase and a-glucosidase added were 99.6% and 101.1%, respectively. Coefficients of variation in within-day and day-to-day experiments were within 1% and 2.8.%, respectively.

Molecular cloning of the glucoamylase gene of Aspergillus shirousami and its expression in Aspergillus oryzae.

The glucoamylase enzyme (GAase) gene from Aspergillus shirousami was cloned and sequenced from genomic and cDNA libraries. The genomic gene was located in the 5.4 kb EcoRI fragment. The deduced amino acid sequence of GAase contained 639 amino acid residues with a relative molecular mass of approximately 68,000 daltons (non-glycosylated form). The genomic gene of A. shirousami GAase was introduced into Aspergillus oryzae. These transformants had increased GAase and raw starch degradation (RSD) activity in culture media and in rice-koji extracts. Analysis by Southern, Northern, SDS-PAGE, and Western blot techniques confirmed the foreign gene was correctly transcribed, translated, and expressed in A. oryzae.