2025 Research results
1.Evaluation of the starch properties of raw rice and enzymatic digestibility of steamed rice grains for sake production by visual observation of swelling behavior in urea solutions
- Author
- OKUDA Masaki, JOYO Midori, TAKAHASHI Kei, and MUKAI Nobuhiko
- Abstract
- Background and Objectives: The facile low‐cost evaluation of rice starch properties may help assess the suitability of rice for sake production and other applications. Herein, the starch gelatinization properties of raw rice and enzymatic digestibility of steamed rice were simply evaluated by observing the swelling of raw rice grains in urea solutions.
Findings: Polished rice grains were soaked in urea solutions of varying concentrations to determine the concentration at which the rice grains started swelling or completely absorbed the solution. The swelling‐onset concentration was strongly positively correlated with the DSC‐determined gelatinization temperature and rapid visco analysis–determined pasting temperature of raw rice and negatively correlated with the enzymatic digestibility of steamed rice (R2 = 0.81–0.94) while being only slightly affected by the rice polishing ratio and moisture content.
Conclusions: The starch gelatinization properties of raw rice and enzymatic digestibility of steamed rice could be simply evaluated using the urea swelling test.
Significance and Novelty: The developed simple method does not require apparatus and flouring and provides results that are not influenced by rice moisture content and polishing ratio, thus holding promise for analyzing rice used in sake production and starch‐containing food materials. - Source
- Cereal Chemistry, 102, 537–547, (2025)
2.Evaluation of the starch properties of rice grains and enzymatic digestibility of steamed rice during sake brewing based on the swelling behavior of rice grains during heating
- Author
- OKUDA Masaki, JOYO Midori, TAKAHASHI Kei, and MUKAI Nobuhiko
- Abstract
- Background and Objectives: The physicochemical properties of cereal starch affect food quality and processing properties, for example, the properties of rice starch influence the suitability of rice for sake (traditional Japanese alcoholic beverage) production. This study reports a convenient method for evaluating the starch gelatinization and retrogradation properties of rice grains and enzymatic digestibility of steamed rice to determine the suitability of japonica rice for sake brewing.
Findings: The swelling properties of polished rice grains were observed during the heating of nonfloured (whole‐grain) samples. The swelling‐onset temperature and swelling ratio were significantly correlated with the amylopectin side‐chain structure of rice starch, starch gelatinization/retrogradation properties, and enzymatic digestibility of steamed rice under sake production–like conditions. Swelling‐onset temperatures at a swelling ratio of 1.2 were measured for 31 samples with a polishing ratio of 70%, and the strength of the corresponding correlations (R2 = 0.79–0.93) were comparable with those observed for gelatinization and pasting temperatures (determined by DSC and rapid visco analysis, respectively).
Conclusions: The starch properties and enzymatic digestibility of multiple steamed rice samples could be rapidly evaluated without flouring.
Significance and Novelty: The method facilitates the evaluation of rice used for sake brewing and analysis of starch properties in a wide range of food industries. - Source
- Cereal Chemistry, 102, 548–562, (2025)
3.Efficient genes identification via quantitative trait loci analysis by crossbreeding of sake and laboratory yeast
- Author
- KANAI Muneyoshi, SHIBATA Tomoko, ZHOU Yan, HAYASHI Risa, FUKUBA Ikuko, KOCHI Takayuki, TERAMOTO Satoko, SHIMOI Hitoshi, TAKAHASHI Hidekazu, and AKAO Takeshi
- Abstract
- Saccharomyces cerevisiae, a unicellular eukaryotic microorganism, includes various strains used in alcoholic beverage production, like sake, shochu/awamori, and wine yeasts. Despite being the same “Saccharomyces cerevisiae”, each strain has unique genes and mutations that make them suitable for specific production processes. We focused on sake yeast, Saccharomyces cerevisiae, suitable for sake making. To identify genes and mutations contributing to sake yeast’s characteristics more efficiently, we improved the quantitative trait loci (QTL) analysis system. This genetic statistical method used spore-separating haploid strains (F1 segregant haploids) from crossing sake yeast and laboratory yeast haploid strains. We increased the number of F1 segregant haploids for QTL analysis from 100 to 400 and set DNA markers uniformly across the genome (approximately 12 Mbp) at 5,267 locations using single nucleotide polymorphisms (SNPs) spaced about 3 kb apart. Additionally, a small-scale sake making test using 400 F1 segregant haploids and QTL analysis of ethanol concentration in sake sample identified the PBS2 gene and its causative mutation (amino acid substitution at position 545). The PBS2 gene was also implicated in producing organic acids (fumaric, succinic, and malic acids) and inorganic acids (phosphoric acid) for sake. These findings validated the improved QTL analysis system as effective genes screening method.
- Source
- Applied Microbiology and Biotechnology, 109, 84 (2025)
4.Novel method for predicting the risk of spoilage by lactic acid bacteria during the storage of Japanese sake
- Author
- TAKAHASHI Masayuki, MORIKAWA Kana, and AKAO Takeshi
- Abstract
- Lactobacillus acetotolerans and Fructilactobacillus fructivorans are responsible for the spoilage of Japanese sake, and sake brewers want to control these bacteria to prevent product-quality deterioration. Due to these bacteria's slow growth, a long time must pass before a product's spoilage can be identified by a culture-dependent method; a method enabling the rapid detection of spoilage is thus desired for maintaining sake's quality and reducing processing times. Herein, we developed a novel, simple method for determining the ori copy and ter copy numbers of both bacterial species by a single multiplex digital polymerase chain reaction (PCR). Our investigation about the ori and ter copy numbers of L. acetotolerans RIB9124 strain revealed that (i) the ter-region copy number was highly correlated with the colony-forming units, and (ii) the ori:ter ratio during log phase (mean: 1.36) was higher than that during non-log phase (mean: 1.16). The relationship between the ori:ter ratio and the bacterial doubling time was parabola-shaped, not linear. We constructed a model for estimating the two species' doubling times based on the ori:ter ratio and then created a generalized linear model with a probit link function using the ori:ter ratio, the doubling time predicted by the ori:ter ratio, and the ter copy number as parameters. The model was able to predict whether sake inoculated with either species would spoil within 4 months (spoilage-prediction error rate validated with 45 samples: 15.6 %). In light of these results, we suggest some criteria for judging the risk of Japanese sake spoiling within 4 months.
- Source
- Applied Food Research, 5(1) , 100835 (2025)
5.Activated carbon treatment to minimize VOC interferences in oxygen stable isotope analysis of beverages
- Author
- AKAMATSU Fumikazu, OSAFUNE Yukio, FUJITA Akiko, BOERZHIJIN Surina, TAKAHASHI Masayuki, and Atsuko ISOGAI
- Abstract
- Oxygen stable isotope analysis is a well-established technique for determining the geographical origin and authenticity of beverages. However, volatile organic compounds (VOCs) present in some beverages can hinder accurate analysis. This study presents a straightforward method to overcome this challenge. Granular activated carbon co-incubated with the beverage sample during CO2-H2O equilibration effectively removes VOCs across a range of beverages tested. This includes propan-1-ol, 2-methylpropan-1-ol, ethyl acetate, 3-methylbutan-1-ol, 3-methylbutyl acetate, and ethyl hexanoate, achieving an average reduction of 77%. Consequently, this method minimizes background noise interferences, preserving δ18O values of water in a wide range of beverages, such as sake, sparkling sake, wine, beer, cider, awamori, brandy, shochu, whisky, and fruit juices. Notably, grain-derived alcoholic beverages like sake and beer exhibit more negative δ18O values compared to fruit-derived alcoholic beverages such as wine and cider, reflecting the distinct source waters and production processes employed. This universally applicable approach minimizes VOC interferences during oxygen stable isotope analysis of aroma-rich beverages. Its inherent simplicity and cost-effectiveness facilitate seamless integration into existing analytical workflows, leading to enhanced efficiency and productivity. This practical solution has potential for brand protection and authenticity verificationwithin the beverage industry.
- Source
- Journal of Food Measurement and Characterization, 19(5), 3179-3187 (2025)
6.Combinatory breeding of sake yeast strains with mutations that enhance Ginjo aroma production
- Author
- GONG Yifeng, Norapat Klinkaewboonwong, HAYASHI Risa, ZHOU Yan, NISHIDA Ikuhisa, SAITO Rei, GOSHIMA Tetsuya, NISHI Tomoyuki, WATANABE Daisuke, HIRATA Dai, AKAO Takeshi, and OHYA Yoshikazu
- Abstract
- Isoamyl acetate and ethyl caproate are the primary aroma compounds responsible for the fruity fragrance characteristic of Ginjo sake. Simultaneous high-level production of both compounds is crucial to achieving a balanced aroma and complex flavor. Isoamyl acetate is predominantly produced by hda1∆/hda1∆ and LEU4(G516S)/LEU4(G516S), while ethyl caproate is produced in high quantities by FAS2(G1250S)/FAS2(G1250S). In this study, to maximize the production of both aroma compounds, genome editing was employed to generate sake yeast strains combining these mutations. After small-scale fermentation tests were conducted to evaluate the production of aroma compounds, we found that the isoamyl acetate–enhancing effect ofhda1∆ /hda1∆ was almost completely masked by FAS2(G1250S)/FAS2(G1250S). In contrast, the effects of LEU4(G516S)/LEU4(G516S) were not entirely masked by FAS2(G1250S)/FAS2(G1250S), resulting in 2.4- and 5.4-fold greater production of isoamyl acetate and ethyl caproate, respectively. This study highlights the utility of genome editing in the combinatorial breeding of sake yeast.
- Source
- Bioscience, Biotechnology, and Biochemistry, 89, 910-917 (2025)
7.Modeling the sensory characteristics of Japanese sake using the sake metabolome analysis method
- Author
- KOBAYASHI Takuji, KOMATSU-HATA Yuko, SAITO Ryota, YAZAWA Hisashi, TAKAHASHI Masayuki, ODA Ken, and IWASHITA Kazuhiro
- Abstract
-
Background/Objectives:
The components of food and beverages are important elements that determine their palatability. Although the components of sake, a traditional Japanese alcoholic beverage, have been studied for many years, their correlation with sensory characteristics is unclear.Methods:
We investigate the correlation with the sake metabolome analysis method developed by our group using ultra-high-performance liquid chromatography quadrupole time-of-flight mass spectrometry. We constructed orthogonal projections to latent structure models to predict sensory evaluation data obtained through the quantitative descriptive analysis method from the sake metabolome data.Results:
For two years of study, 8 sensory evaluation models of the 2016 brewing year and 11 sensory evaluation models of the 2017 brewing year, including color, ethyl hexanoate, Hine-ka, Nama hine-ka, ethyl acetate, grainy/sweet aroma, sweetness, sourness, body, astringency, harsh taste/acrid taste, aftertaste, and overall quality, demonstrated a predictive performance with Q2 > 0.5. Liquid chromatography-based analytical data indicated that it is possible to predict not only taste but also aroma. Additionally, the generalization performance of the prediction models for sensory evaluation attributes common to both years was verified.Conclusions:
These results provide a new option for explaining the sensory characteristics of sake from its components and contribute to a deeper understanding of them. - Source
- Metabolites, 15 (8), 559 (2025)
8.Development and evaluation of rapid and simple assay method for α-amylase activity in rice koji
- Author
- ODA Ken, SHIMAMOTO Kazumi, YAMASAKI Risa, SHIMOJI Kazuhiko, BAKKE Mikio, SUZUKI Shigeya, GOSHIMA Tetsuya, and AKAO Takeshi
- Abstract
- A chromogenic substrate assay using 2-chloro-4-nitrophenyl-65-azido-65 -deoxy-β-maltopentaoside (G5 method) is one of the official analytical methods for α-amylase activity in rice koji for sake-brewing. Although it is useful because of its rapidity, simplicity, and accuracy, there are concerns about unavailability of the substrate in near future. In this study, an alternative assay using 4,6-O-ethylidene-α-4-nitrophenyl-maltoheptaoside (G7 method) was developed as a method that can be measured using the same procedure as the G5 method. α-Amylase in the sample and the coupled enzymes, glucoamylase and α-glucosidase, hydrolyzes the substrate, and the resulting chromophore, measured at 400 nm, was directly proportional to the α-amylase activity. As the G7 method exhibited good precision, repeatability, and linearity with the G5 method for 41 kinds of rice koji (y = 1.096x, R2 = 0.996), it is useful for the determination of α-amylase activity in rice koji. When switching the G5 method to the G7 method, it should be noted that the G7 method tends to exhibit ca. 10% higher values than the G5 method as an average. Moreover, the ratios of the measurement values between the G7 and G5 methods can vary from 1.02 to 1.38 depending on the rice koji samples. Evaluation of Km values suggested that the catalytic properties of α-amylase for the G7 and G5 methods can vary among rice koji samples. Therefore, if the G7 method is used instead of the G5 method, the comparison of assay results of the two methods for the target koji sample is preferable to ensure data compatibility.
- Source
- Journal of Food Science, 90, e70344 (2025)
9.The relationship between glucose concentrations and overall ratings of sakes at the final-stage sensory evaluations of the Annual Japan sake contests
- Author
- FUJITA Akiko, and AKAMATSU Fumikazu
- Abstract
- To investigate the effect of the glucose concentrations on the overall ratings of ginjo-shu at the final-stage sensory evaluations of the Annual Japan Sake Contests, we analyzed the chemical components and sensory evaluation results of 1,285 entries from the past three years. We found that higher glucose concentrations tended to correlate with better overall ratings, while higher acidity and amino acid levels tended to correlate with worse overall ratings. Conversely, the differences in glucose concentration and other components between sequential sake samples did not significantly affect the overall rating. These results suggest that the judging panel at the final-stage sensory evaluation of the Annual Japan Sake Contests evaluated each sake independently and performed separate sensory assessments based on established criteria of flavor harmony and character.
- Source
- J. Brew. Soc. Japan, 120, 463-468 (2025)
10.Analysis of sake components presented to the Japan Sake Awards 2024
- Author
- ISOGAI Atsuko, FUJITA Akiko, AKAMATSU Fumikazu, BOERZHIJIN Surina, KAMIMOTO Maki, TAKAHASHI Masayuki, NISHIMOTO Masaki, TERAMOTO Satoko, IKEDA Yuriko, NITTA Hidemi, OGUSHI Kensuke, and FUKUDA Hisashi
- Source
- Report of the Research Institute of Brewing, 197, 1-17 (2025)
11.Review of the 47th Honkaku Shochu and Awamori Contest in 2023
- Author
- MUKAI Nobuhiko, ISOGAI Atsuko, OKUDA Masaki, TAKAHASHI Kei, OSAFUNE Yukio, IIZUKA Sachiko, and FUKUDA Hisashi
- Source
- Report of the Research Institute of Brewing, 197, 18-36 (2025)
12.Analysis of beer components presented to craft beer contest of Japan Brewers Association in 2025
- Author
- KUSAKA Kazutaka, TSUJII Masayuki, YAZAWA Hisashi, SUDA Shinri, KAMIMOTO Maki, and TERAMOTO Satoko
- Source
- Report of the Research Institute of Brewing, 197, 37-45 (2025)
13.The increase in the cell volume and number of nuclei of the Koji-kin Aspergillus oryzae contributes to their high enzyme productivity
- Author
- ITANI Ayaka, MOTOMURA Haruto, ODA Ken, YAMASHITA Hideyuki, SAKAI Kanae, KUSUMOTO Kenichi, SHIGETO Shinsuke, ICHIKAWA Takehiko, Hosain Mohammad Mubarak, HUKUMA Takeshi, KATAYAMA Takuya, MARUYAMA Jun-ichi, MASUO Shunsuke, TAKAYA Naoki, and TAKESHITA Norio
- Abstract
- Aspergillus oryzae is widely employed in traditional Japanese brewing, such as sake and soy sauce production, due to its potent carbohydrate- and protein-degrading capabilities. Additionally, it serves as an efficient host for heterologous expression and is increasingly utilized in the enzyme production industry. Although genome and transcriptional regulatory analyses have progressed in investigating the factors underlying the utility of A. oryzae, these factors remain incompletely understood. Our study revealed that the hyphae of A. oryzae undergo thickening during cultivation, accompanied by a tenfold increase in the number of nuclei within apical cells. These thick hyphae with an increased number of nuclei emerge abruptly via branching, exhibit rapid growth, and predominantly occupy the colony periphery. Hyphal cell volume and nuclear number are correlated, and the emergence of thick hyphae is associated with enhanced enzymatic activity and protein secretion capacity. Through transcriptomic analyses of mycelia or individual hyphae and SNP analyses of industrial strains from distinct clades, we identified a Ca²⁺ channel and a glycosyl transferase that contribute to the increase in cell volume. In addition to possessing the potential for cell volume expansion, the environment must also support the conditions that allow for an increase in nuclear number. Under nutrient-rich conditions, such as the presence of specific amino acids, the TOR pathway is activated, resulting in increased ribosome production and enhanced mitochondrial activity, which are likely to promote an increase in nuclear number. An increase in the number of nuclei per hyphal cell enhances transcription, translation, and enzyme secretion per cell, which is advantageous for industrial strains. Our study offers insights into the mechanisms by which the Koji-kin A. oryzae developed this unique trait through long-term breeding.
- Source
- eLife, 14, RP107043 (2025)
14.Ribonucleoprotein-based CRISPR/Cas9 genome co-editing in Aspergillus luchuensis mut. Kawachii
- Author
- KARASHIMA Takefumi, ODA Ken, FUTAGAMI Taiki, HOKAZONO Hideki, and TAKASHITA Hideharu
- Abstract
- In this study, we established a ribonucleoprotein-based clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) genome co-editing method for the white koji fungus, Aspergillus luchuensis mut. kawachii. To introduce the single guide RNA-Cas9 ribonucleoprotein complex into protoplast cells of A. luchuensis mut. kawachii, we investigated the conditions for protoplast preparation using Yatalase -Plus-. Subsequently, we employed the ribonucleoprotein-based method to knockout the ATP sulfurylase-encoding sC gene, which imparts selenate resistance in the model strain NBRC 4308 and the industrial strain No. 8046. Furthermore, we explored genome co-editing by simultaneously targeting sC along with either the orotidine 5'-phosphate decarboxylase-encoding pyrG gene or the transcriptional activator of protease genes-encoding prtR gene in NBRC 4308. The transformants were selected in medium containing selenate, resulting in the successful generation of pyrG- and prtR-knockout strains. Similarly, transformants were selected on medium containing selenate, resulting in the successful generation of prtR-knockout strain in No. 8046. These results demonstrate that the ribonucleoprotein-based genome co-editing method is applicable not only to the model strain but also to industrial strains, making it a promising approach for manipulating A. luchuensis mut. kawachii.
- Source
- Journal of Bioscience and Bioengineering, 140, 298-305 (2025)
15.Breeding of the sake yeast K1801 with normal checkpoint integrity by isolating spontaneous cdc55 revertants
- Author
- GOSHIMA Tetsuya, NISHIDA Ikuhisa, SHIMOI Hitoshi, AKAO Takeshi, and HIRATA Dai
- Abstract
- Sake yeast strain K1801 with high productivity of ethyl caproate is widely used for high-quality sake brewing; however, K1801 has a defect in the spindle assembly checkpoint, which is responsible for the mutation R48P of the PP2A regulatory subunit Cdc55. Here, we successfully isolated spontaneous revertants of the cdc55 mutation with the excellent brewing property similar to K1801 and normal checkpoint integrity.
- Source
- Bioscience, Biotechnology, and Biochemistry, 89(9), 1400–1404 (2025)
16.Locus heterogeneity in the mechanism underlying the superior fermentation performances of phylogenetically distant sake yeasts
- Author
- MORI Kotaro, SEIKE Taisuke, AKAO Takeshi, TAKAO Yoshifumi, YAMADA Tasuku, TAKAHASHI Toshinari, and MATSUDA Fumio
- Abstract
- Sake yeasts, Saccharomyces cerevisiae strains isolated from Japanese sake fermentation tanks, exhibit superior fermentation performance to that of other yeast strains. Although the exceptional abilities of the modern sake yeast strain K701 have been extensively investigated, those of phylogenetically distant classical sake yeasts remain largely understudied. In this study, we aimed to clarify the mechanism underlying the superior fermentation ability of the classical sake yeast strain Km67 by comparing its genetic and physiological properties with those of the laboratory strains X2180 and K701. Km67 did not exhibit quiescence-specific phenotypes in sake mashes in the same manner as K701. RNA sequencing revealed similar trends between the transcriptomes of Km67 and K701 in sake mash, suggesting that the lack of quiescence entry was related to the higher fermentation ability of Km67. Genetic testing demonstrated that signals upstream of Rim15p were not conveyed downstream, indicating impaired fermentation repression mediated by Rim15p in Km67 cells. Protein phosphatase 2A (PP2AB55δ) activity declined following CDC55 deletion downstream of Rim15p, consequently reducing the fermentation performance of Km67. However, the extent of fermentation reduction upon CDC55 deletion was not as large as that with X2180 and K701. This suggested the presence of unidentified factors that regulated fermentation independently of PP2AB55δ or Rim15p in Km67. Thus, our findings demonstrate locus heterogeneity in the mechanism underlying fermentation abilities between the phylogenetically distant sake yeasts Km67 and K701.
- Source
- Journal of Bioscience and Bioengineering, 140(5), 306－313 (2025)
17.Study on the solubility characteristics (enzyme digestibility/starch content) of rice in sake mash
- Author
- HIRASAWA Ryotaro, OKUDA Masaki, and HIRATA Dai
- Abstract
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To investigate the dissolution characteristics of rice in sake mash during fermentation, small-scall sake brewing was carried out using 3 varieties of rice (Gohyakumangoku/GOM, Koshitanrei/KOS, and Koshihibuki/KIB) of 2 polishing ratios (50% and 70%).
Based on component measurements of the mash, the changes in total extract between polishing ratios showed that GOM and KIB of a 50% polishing ratio tended to be significantly higher than those of a 70% polishing ratio in the first half of the mash, but no difference in total extract was observed between the 2 polishing ratios in KOS. This suggests that the dissolution rate differs depending on the polishing ratio between KOS, GOM, and KIB. In addition, when comparing varieties, no significant differences were observed in total extract on the 20th day, at the end of the mash.
Steamed rice (including koji) was taken from the mash, and the Brix value per amount of starch was defined as the solubility potential (Brix value/amount of starch) for ease of solubility of the rice at the time of collection (solubility characteristics), and this value was calculated. KOS had a higher solubility potential than the other two varieties. In addition, in the first half of the mash, the solubility potential was higher at a 50% polishing ratio than at a 70% polishing ratio for both varieties, but there was no significant difference at the end of the mash (20th day). In addition, a strong correlation was found between the increase in the amount of total extract and the solubility potential, regardless of the polishing ratio. Furthermore, since the solubility potential was higher in mash with slower alcohol production than in mash with faster alcohol production, it was inferred that the alcohol in the mash may have affected the aging rate of the steamed rice (decrease in enzyme digestibility).
- Source
- J. Brew. Soc. Japan, 120(11),625-634 (2025)
18.Effect of fermentation factors on ethyl caproate production in beer: An orthogonal array approach
- Author
- MARUYAMA Hironori, ITO Daichi, FUKUBA Ikuko, KANAI Muneyoshi, and HASEGAWA Keishi
- Abstract
- This study investigated fermentation conditions that enhance ethyl caproate production in beer brewing using the Mie sake yeast strain BMK3, which is characterized by high ethyl caproate production and efficient maltose assimilation. A small-scale fermentation trial was conducted using an L9 orthogonal array comprising four fermentation factors—temperature, pitching rate, initial glucose concentration, and initial maltose concentration—each at three levels. A total of 18 fermentations were performed, and volatile components were analyzed based on their peak concentrations during fermentation. Four-way ANOVA revealed that fermentation temperature had the greatest influence on ethyl caproate and its precursor caproate, followed by glucose concentration. In contrast, glucose had less effect on isoamyl acetate production. To explore the impact of these fermentation factors on yeast metabolism, a wide-target analysis of water-soluble metabolites in the brewed beer was conducted using CE–MS and subjected to principal component analysis (PCA). PCA showed that temperature primarily influenced PC1, while glucose concentration contributed to PC2. Several metabolites associated with NADPH regeneration, which is essential for fatty acid biosynthesis, were more abundant under high-glucose conditions. These findings suggest that glucose levels affect ethyl caproate formation through modulation of yeast metabolic pathways, particularly those related to fatty acid synthesis.
- Source
- Journal of the American Society of Brewing Chemists, 83(4), 442-451 (2025)
19.Effects of Aspergillus oryzae-derived rice-koji protein on the sake metabolome
- Author
- KATAOKA Ryousuke, FUKUHARA Shinichiro, KAKIUCHI Shingo, KONO Minori, Sharon Marie Bahena-Garrido, KOMATSU-HATA Yuko, and IWASHITA Kazuhiro
- Abstract
- Rice-koji is produced by cultivating Aspergillus oryzae on steamed rice and plays a central role in the production of traditional Japanese alcoholic beverages such as sake and shochu, as well as fermented seasonings like miso and soy sauce. While some functions of the proteins produced by A. oryzae during rice-koji fermentation have been clarified, the comprehensive protein profile and their specific roles in sake brewing are still not fully understood. We hypothesized that proteins expressed by A. oryzae in rice-koji contribute in diverse ways to shaping the metabolic characteristics of sake. In this study, we performed proteomic analyses on four types of rice-koji to explore rice-koji proteins (RKPs) and identified 159 rkp genes. From this set, selected targets were used to construct single-gene disruptants (Δrkp), resulting in the generation of 85 distinct Δrkp strains. Sake brewing experiments using these disruptants were carried out to assess the impact of individual rkp gene deletions. The results revealed alterations in the mycelial biomass of A. oryzae in rice-koji and showed clear associations with variations in multiple sake components. Further metabolomic analysis demonstrated that disruption of 57 rkp genes led to notable changes in at least one sake-derived metabolite peak. Overall, our findings indicate that RKPs identified through proteomic analysis of rice-koji exert a wide range of effects on both the growth of A. oryzae and the metabolite composition of sake. These insights are expected to advance our understanding of how RKPs influence sake brewing and the broader biological roles of A. oryzae.
- Source
- Applied and Environmental Microbiology, 92(3), e0195525 (2026)
20.Aspergillus oryzae strains heterogeneously produce plant polysaccharide degradation-related enzymes
- Author
- FUJIWA Shimma, KATAOKA Ryousuke, IWASHITA Kazuhiro, and MATSUZAWA Tomohiko
- Abstract
- Aspergillus oryzae is used for brewing, and many strains with different brewing characteristics have been isolated. We compared enzymatic activities of eight A. oryzae strains, RIB40, 128, 143, 163, 301, 915, 1108, and 1178 strains, toward plant polysaccharides. The plant polysaccharide degradation-related enzymes produced by A. oryzae change depending on the monosaccharides and polysaccharides added to the culture medium. RIB915 produced more cellulolytic and hemicellulolytic enzymes, whereas RIB40, 128, and 301 produced less of these enzymes than the other strains. In addition, A. oryzae RIB128 produced different glycoside hydrolases in response to monosaccharides and polysaccharides compared with other strains. These results indicated that there is diversity in the production of plant polysaccharide degradation-related enzymes within A. oryzae species.
- Source
- Journal of the Bioscience and Bioengineering, 141(1), 12-22 (2026)