2019 Research results

1. Characteristics of the volatile composition of Shochu in the IzuIslands

Author: Hisashi Fukuda, Osamu Yamada and Jinshun Han
Abstract: To identify characteristics of compounds of Shochu made in the Izu Islands, we analyzed 78 volatile compounds by head space solid phase micro extraction(SPME), and 6 volatile compounds by direct head space analysis. There was a significant difference in 44 volatile compounds of sweet potato shochu between that in the Izu Islands and the in Honkaku Shochu Contest. In a stepwise discriminant analysis procedure, all of samples were rightly classified by 4 volatile compounds (1-octen-3-ol, ethyl butyrate, Ethyl enanthate, and vanillin). Estimating the accuracy of the discriminant analysis by cross validation, the ratios of appropriate classification were 100%. For barley shochu in the Izu Islands and in the Honkaku Shochu Contest, there was a significant difference in 29 of 84 volatile compounds. A total of 67 of 68 samples were rightly classified by 4 volatile compounds (Nonanal, Ethyl acetate, Ethyl butyrate, and DMTS). When estimating the accuracy of the discriminant analysis, the ratios of appropriate classification were 98.5%. For blended shochu of sweet potato shochu and barley shochu in the Izu Islands, there was a significant difference in 42 volatile compounds between sweet potato shochu, barley shochu, and blended shochu. In a stepwise discriminant analysis procedure, 30 of 33 samples were rightly classified by 8 volatile compounds (Citronellol, Ethyl isobutyrate, Ethyl caproate, Isoamyl caproate, Ethyl caprate, Ethyl cinnamate, Diethyl succinate and Guaiacol).
Source: J. Brew. Soc. Japan, 114, 151-158 (2019) in Japanese

2. Analysis of 4-mercapto-4-methylpentan-2-one in sake presented at the Sake Contest

Author: Sachiko Iizuka, Atsuko Isogai, Ryoko Kanda, and Tsutomu Fujii
Abstract: Recently, "muscat-like", "litchi-like" or "citrus-like" odor characteristics have been indicated in sake presented at Sake Contests. As thiols were considered to contribute to these odor characteristics, we analyzed one of thiol compounds, 4-mercapto-4-methylpentan-2-one (4MMP), in sake presented at the Sake Contest in 2015. The content of 4MMP in 25 samples was 5～14 ng/L, which was higher than the perception threshold of 4MMP in sake (1.2 ng/L) previously reported. To study the influence of 4MMP on the odor property of sake, a sensory evaluation was performed with the addition of 4MMP to the sake. The 'sulfur' odor property was significantly enhanced by the addition of ≥ 8.0 ng/L of 4MMP. Though the addition of 16.0 ng/L of 4MMP significantly lowered the total apparaisal, it significantly enhanced 'fruity' odor properties different from the aroma of ethyl caproate and isoamyl acetate.
Source: J. Brew. Soc. Japan, 114, 93-101 (2019) in Japanese

3. Carbon Stable Isotopic Compositions of Citric Acid and Malic Acid in Japanese Apricot Liqueur Decrease as the Fruit Ripens

Author: Fumikazu Akamatsu, Yasuhisa Tsuchida, Takaaki Oe, Yuri Hisatsune, Yukari Igi, Tomokazu Hashiguchi, Tsutomu Fujii
Abstract: The carbon stable isotopic composition (δ¹³C) is often analyzed to quantify the addition of acidulants to Japanese apricot liqueur, but little is known about the variation in theδ¹³C values of the main organic acids arising from differences in the ripeness of Japanese apricots. We show that in Japanese apricot liqueur prepared using fruits at different stages of ripeness, theδ¹³C values of citric acid and malic acid ranged from -25.1‰ to -23.7‰ and from -22.3‰ to -19.7‰, respectively, and theδ¹³C values decreased as the fruit ripened. The averageδ¹³C value of citric acid from liqueurs was 0.7‰ higher than that from fresh fruits, whereas theδ¹³C values of malic acid showed no isotope discrimination. The variation inδ¹³C values of the main organic acids in Japanese apricot liqueurs will help detect acidulant addition and control authenticity.
Source: Food Chemistry, 277, 70-74 (2019)

4. The potential aroma and flavor compounds in Vitis sp. cv. Koshu and V. vinifera L. cv. Chardonnay under different environmental conditions

Author: Sharon Marie Bahena-Garrido, Tomoko Ohama, Yuka Suehiro, Yuko Hata, Atsuko Isogai, Kazuhiro Iwashita, Nami Goto-Yamamoto and Kazuya Koyama
Abstract: BACKGROUND

Koshu, a hybrid of Vitis vinifera L. and V. davidii Foex, is the most popular indigenous cultivar for wine production in Japan. However, little is known about the potential aroma compounds it contains and how environmental factors affect these. In this study, we obtained comprehensive profiles of the volatile (both glycosidically bound and free) and phenolic compounds that occur in koshu berries, and compared these with similar profiles for V. vinifera cv. chardonnay. We then compared the response of these two cultivars to bunch shading and the ripening-related phytohormone abscisic acid (ABA).

RESULTS

Koshu berries contained significantly higher concentrations of phenolic compounds, such as hydroxycinnamic acid derivatives, and some volatile phenols, such as 4-vinyl guaiacol and eugenol, than chardonnay berries, which are thought to contribute to the characteristics of koshu wine. In addition, koshu berries had a distinctly different terpenoid composition from chardonnay berries. Shading reduced the concentration of norisoprenoid in both cultivars, as well as several phenolic compounds, particularly their volatile derivatives in koshu berries. The exogenous application of ABA induced ripening and increased the concentrations of lipid derivatives, such as hexanol, octanol, 1-nonanol, and 1-octen-3-ol. Multivariate and discriminant analyses showed that the potential aroma and flavor compounds in the berries could be discriminated clearly based on cultivar and environmental cues, such as light exposure.

CONCLUSION

The unique secondary metabolite profiles of koshu and their different responses to environmental factors could be valuable for developing various types of koshu wines and new cultivars with improved quality and cultural characteristics.
Source: Journal of the science of food and agriculture, 99, 1926-1937 (2019)

5. Meiotic chromosomal recombination defect in sake yeasts

Author: Hitoshi Shimoi, Yuta Hanazumi, Natsuki Kawamura, Miwa Yamada, Shohei Shimizu, Taro Suzuki, Daisuke Watanabe, and Takeshi Akao
Abstract: Sake yeast strains are classified into Saccharomyces cerevisiae and have a heterothallic life cycle. This feature allows cross hybridization between two haploids to breed new strains with superior characteristics. However, cross hybridization of sake yeast is very difficult because only a few spores develop in a sporulation medium, and most of these spores do not germinate. We hypothesized that these features are attributable to chromosome recombination defect in meiosis, which leads to chromosome loss. To test this hypothesis, we examined meiotic recombination of sake yeast Kyokai no. 7 (K7) using the following three methods: (i) analysis of the segregation patterns of two heterozygous sites in the same chromosome in 100 haploid K7 strains; (ii) sequencing of the whole genomes of four haploid K7 strains and comparison of the bases derived from the heterozygosities; and (iii) construction of double heterozygous disruptants of CAN1 and URA3 on the chromosome V of K7 and the examination of the genotypes of haploids after sporulation. We could not detect any recombinant segregants in any of the experiments, which indicated defect in meiotic recombination in K7. Analyses after sporulation of the same double heterozygous disruptants of K6, K9, and K10 also indicated meiotic recombination defect in these strains. Although rapamycin treatment increased the sporulation efficiency of K7, it did not increase the meiotic recombination of the double heterozygous K7. Moreover, the spo13 disruptant of the K7 derivative produced two spore asci without meiotic recombination. These results suggest that sake yeasts have defects in meiotic recombination machinery.
Source: Journal of Bioscience and Bioengineering, 127, 190-196 (2019)

6. Nutrient signaling via the TORC1-greatwall-PP2A^B55δ pathway responsible for the high initial rates of alcoholic fermentation in sake yeast strains of Saccharomyces cerevisiae

Author

Daisuke Watanabe, Takuma Kajihara, Yukiko Sugimoto, Kenichi Takagi, Megumi Mizuno, Yan Zhou, Jiawen Chen, Kojiro Takeda, Hisashi Tatebe, Kazuhiro Shiozaki, Nobushige Nakazawa, Shingo Izawa, Takeshi Akao, Hitoshi Shimoi, Tatsuya Maeda, Hiroshi Takagi.

Abstract

Saccharomyces cerevisiae sake yeast strain Kyokai no. 7 (K7) and its relatives carry a homozygous loss-of-function mutation in the RIM15 gene, which encodes a Greatwall-family protein kinase. Disruption of RIM15 in non-sake yeast strains leads to improved alcoholic fermentation, indicating that the defect in Rim15p is associated with the enhanced fermentation performance of sake yeast cells. In order to understand how Rim15p mediates fermentation control, we here focused on target-of-rapamycin protein kinase complex 1 (TORC1) and protein phosphatase 2A with the ^B55δ regulatory subunit (PP2A^B55δ), complexes that are known to act upstream and downstream of Rim15p, respectively. Several lines of evidence, including our previous transcriptomic analysis data, suggested enhanced TORC1 signaling in sake yeast cells during sake fermentation. Fermentation tests of the TORC1-related mutants using a laboratory strain revealed that TORC1 signaling positively regulates the initial fermentation rate in a Rim15p-dependent manner. Deletion of the CDC55 gene encoding ^B55δ abolished the high fermentation performance of Rim15p-deficient laboratory yeast and sake yeast cells, indicating that PP2A^B55δ mediates the fermentation control by TORC1 and Rim15p. The TORC1-Greatwall-PP2A^B55δ pathway similarly affected the fermentation rate in the fission yeast Schizosaccharomyces pombe, strongly suggested that the evolutionarily conserved pathway governs alcoholic fermentation in yeasts.

It is likely that elevated PP2A^B55δ activity accounts for the high fermentation performance of sake yeast cells. Heterozygous loss-of-function mutations in CDC55 found in K7-related sake strains may indicate that the Rim15p-deficient phenotypes are disadvantageous to cell survival.

IMPORTANCE

The biochemical processes and enzymes responsible for glycolysis and alcoholic fermentation by the yeast S. cerevisiae have long been the subject of scientific research. Nevertheless, the factors determining fermentation performance in vivo are not fully understood. As a result, the industrial breeding of yeast strains has required empirical characterization of fermentation by screening numerous mutants through laborious fermentation tests. To establish a rational and efficient breeding strategy, key regulators of alcoholic fermentation need to be identified. In the present study, we focused on how sake yeast strains of S. cerevisiae have acquired high alcoholic fermentation performance. Our findings provide a rational molecular basis to design yeast strains with optimal fermentation performance for production of alcoholic beverages and bioethanol. In addition, as the evolutionarily conserved TORC1-Greatwall-PP2A^B55δ pathway plays a major role in the glycolytic control, our work may contribute to research on carbohydrate metabolism in higher eukaryotes.

Source

Applied and Environmental Microbiology, 85,e02083-18 (2019)

7. Glutelin subtype-dependent protein localization in rice grain evidenced by immunodetection analyses.

Author: Kei Takahashi, Hiromi Kohno, Tomomichi Kanabayashi, and Masaki Okuda
Abstract: Key message

GluA and GluB-4/5 glutelin subfamilies are mainly localized to outer region of the endosperm, particularly in its ventral side, in rice grain, but GluC is localized to throughout the endosperm.

The major seed storage protein in rice (Oryza sativa) is glutelin, which forms a vacuole-derived protein body type-II. Glutelins are encoded by multiple genes, and generally comprise four protein subfamilies, namely, GluA, GluB, GluC, and GluD: however, the localization pattern of glutelin in rice grains remains obscure. In this study, we investigated the localization pattern of five subtypes of the glutelin protein in rice grains using glutelin-subtype specific antibodies. Immunoblot analysis against sequentially polished rice flour fractions from three crop years and seven japonica rice varieties revealed that GluA was strongly localized in the outer region of the endosperm, including the subaleurone layer, whereas GluC was distributed throughout the endosperm. Among the glutelin subtypes, GluA was mostly found in the outer region of the rice grain, followed by GluB-4/5, GluB-1, GluD, and GluC. Immunofluorescence labeling microscopy analysis using immature rice seeds clearly revealed that the localization pattern of GluC and GluD was completely different from that of GluA and GluB. Expression levels of all glutelins, particularly GluA, GluB-1, and GluB-4/5, were stronger on the ventral than dorsal side in rice grains. These results provide strong and consistent evidence that glutelins localize to the rice grain in a subfamily-dependent manner.
Source: Plant Molecular Biology, 100, 3, 231-246 (2019)

8. Anti-diabetic effect of S-adenosylmethionine and α-glycerophosphocholine in KK-Ay mice

Author: Hanae Izu, Mayumi Okuda, Sachi Shibata, Tsutomu Fujii, and Kiminori Matsubara
Abstract: Six-week-old male KK-Ay mice received drinking water with S-adenosylmethionine (SAM), α-glycerophosphocholine (GPC), or SAM+GPC for 10 weeks. The serum glucose of SAM+GPC at 15 weeks old, total cholesterol of GPC at 12 weeks old, and triglyceride of GPC at 15 weeks old and of SAM at 16 weeks old were reduced. SAM+GPC reduced serum leptin and food intake.
Source: Biosci Biotechnol Biochem. 83(4), 747-750 (2019)

9. Analysis of Sake Components Presented to Sake Contest in 2018

Author: Tsutomu FUJII, Akiko FUJITA, Sachiko IIZUKA, Hanae IZU, Ryoko KANDA, Toru KISHIMOTO, Nami GOTO-YAMAMOTO
Source: Report of the Research Institute of Brewing, 191, 1-17 (2019) in Japanese

10. Review of the 41st Honkaku Shochu and Awamori Contest in 2018

Author: Nobuhiko MUKAI, Atsuko ISOGAI, Muneyoshi KANAI, Masayuki TAKAHASHIO, Yukio OSAFUNE, Akinori MUTO, Toshihiro YOSHIMURA, and Nami GOTO-YAMAMOTO
Source: Report of the Research Institute of Brewing, 191, 18-36 (2019) in Japanese

11. Analysis of Beer Components Presented to Craft Beer Contest of Japan Brewers Association in 2019

Author: Shinichi ITO, Kazutaka KUSAKA
Source: Report of the Research Institute of Brewing, 191, 37-43 (2019) in Japanese

12. Characteristics of the starch and protein composition of rice grains used for awamori

Author: Masaki Okuda, Midori Joyo, Hisashi Fukuda, Takako Tamamura
Abstract: Recently, the properties of Thai non-broken rice for the production of awamori have varied significantly among shipments imported from Thailand, and these variations make the production of awamori difficult. In the present study, we investigated the starch and protein properties of Thai rice used for awamori production compared with Nipponbare, which is used as a cultivar for sake making, because these properties have the possibilities of affecting the water absorption ratio and enzyme digestibility of steamed rice grains. The results of individual alkali solubility testing and DSC (= Differential Scanning Calorimeter) analyzing showed that the shipments of Thai rice have mixtures of different properties in gelatinization temperature and alkali solubility. The Thai rice of high gelatinization temperature had a lower content of short-chain (DP6-8 = Degree of polymerization ) of amylopectin structure than that of low temperature gelatinization and Nipponbare. This result indicated that the difference of the amylopectin structure was due to the starch synthase IIa activity from a previous report. The starch in Thai rice with a high gelatinization temperature retrogradated faster than that with a low gelatinization temperature. The analysis of protein composition showed no clear difference between the Thai rice with high gelatinization temperature and that with low gelatinization temperature although intense 60kDa protein bands were observed in Thai rice. From the above results, the mixture of cultivars in Thai rice may influence treatments of materials for the production of awamori.
Source: J. Brew. Soc. Japan, 114,585-595 (2019)

13. Effect of organic acids in alcoholic drinks on the umami of food paired with alcoholic drinks

Author: Akiko FUJITA, Kenichi TOMIHARA, Yuri HISATSUNE, Yukio HAMADA, Tsutomu FUJII
Abstract: While cheese is widely known to pair well with wine, it is presently also consumed as a complement to sake in Japan. Cheese contains high levels of umami constituents such as amino acids and peptides. Its rich taste decreases sensorially in pairing with wine in comparison with sake. In order to confirm this fact, the intensity of the aftertaste of some kinds of food in paring with alcoholic drinks was measured by instrumental taste sensor analysis. Intensity of the umami aftertaste of cheese, miso soup, noodle soup, monosodium glutamate, and a kokumi seasoning was lower in paring with white wines than with sake. Among the major chemical constituents that distinguish wine from sake, organic acids such as tartaric acids and malic acids reduced umami aftertaste intensity. In paired comparison tests conducted for a pair of sake samples with and without tartaric acid, more panelists selected the sake without tartaric acid as the sample which left a stronger umami aftertaste of cheese. These results suggest that organic acids in alcoholic drinks reduce umami aftertaste derived from amino acids and peptides of food in food and alcohol pairings.
Source: J. Brew. Soc. Japan, 114(8), 522-529 (2019)

14. Nutritional characterization of sake cake (sake-kasu) after heat-drying and freeze-drying.

Author: Hanae IZU, Sayo YAMASHITA, Hideyuki ARIMA, Tsutomu FUJII
Abstract: Sake cake contains rice-derived components, as well as cell components and metabolites of Aspergillus oryzae and Saccharomyces cerevisiae. In this study, the effect of food processing on sake cake (sake-kasu) ingredients was investigated. Sake cake, obtained through brewing liquefied rice, was heat-dried (HD) or freeze-dried (FD) and analyzed. There were no differences in the amounts of proteins, lipids, carbohydrates, vitamin B6, choline, betaine, nicotinic acid, β-glucan and resistant proteins in HD and FD. There was also no difference in the amount of hydrolyzed amino acids in HD and FD, but many free amino acids were observed in HD. S-adenosyl methionine (SAM) was found to be abundant in FD. Meanwhile, nucleic acid-related components were found to be increased in HD, which seems to be due to the degradation of microbial metabolites. When considering the health benefits of sake cake, it is necessary to pay attention to the effects of processing method.
Source: Biosci Biotechnol Biochem. 83(8), 1477-1483 (2019)

15. Sake cake (sake-kasu) ingestion increases branched-chain amino acids in the plasma, muscles, and brains of senescence-accelerated mice prone 8

Author: Hanae IZU, Sachi SHIBATA, Tsutomu FUJII, and Kiminori MATSUBARA
Abstract: To examine metabolic effects of sake cake ingestion, plasma and tissues were analyzed in senescence-accelerated mice prone 8 (SAMP8) fed a sake cake diet. As a result, branched-chain amino acids (BCAA) were found to be significantly higher in the plasma, gastrocnemius muscles and brains of the sake cake group than in the control group. Mice in the sake cake group showed stronger grip strength than the control group. High levels of circulating BCAA have been reported to be associated with pathological states, such as metabolic diseases, but the parameters of glucose and lipid metabolism were not affected between the two groups. Otherwise, pyridoxal was significantly higher and nicotinamide as well as 1-methylnicotinamide showed a tendency to be higher in the plasma of the sake cake group than in the control group. These findings indicate that intake of sake cake increases the levels of BCAA, vitamin B6, and vitamin B3.
Source: Biosci Biotechnol Biochem. 83(8), 1490-1497 (2019)

16. Histone deacetylases in sake yeast affect fermentation characteristics

Author: Kazuya Tomimoto, Takeshi Akao, Hisashi Fukuda
Abstract: Yeast histone deacetylases (HDAC) affect the production of alcoholic beverages. In this study, we evaluated the sake fermentation characteristics when using HDAC gene-disrupted yeast strain Kyokai No. 701. Flavor components of the sake product were significantly changed. RPD3 or HDA1 disruption increased twofold the amount of isoamyl acetate, and isoamyl alcohol levels also increased in the rpd3△ strain. To determine the contribution of Rpd3L and Rpd3S complexes to sake characteristics, a gene responsible for Rpd3L and/or Rpd3S formation was also disrupted. Disruption of DEP1 or SDS3 that is an essential component of Rpd3L led to increased isoamyl alcohol production similar to that of the rpd3△ strain, but the efficiency of isoamyl alcohol esterification was not affected. In addition, Rpd3 and Hda1 may regulate the responsiveness to oxygen in isoamyl acetate production. We conclude that HDAC genes regulate the production of flavor components during sake fermentation.
Source: Bioscience, Biotechnology, and Biochemistry, 83 (8), 1498-1505 (2019)

17. SKO1 deficiency extends chronological lifespan in Saccharomyces cerevisiae

Author: Koji Masumura, Sachi Matsukami, Kumiko Yonekita, Muneyoshi Kanai, Kazunori Kume, Dai Hirata & Masaki Mizunuma
Abstract: Sko1 plays a key role in the control of gene expression by osmotic and oxidative stress in yeast. We demonstrate that the decrease in chronological lifespan (CLS) of hog1Δ cells was suppressed by SKO1 deletion. sko1Δ single mutant cells were shown to have a longer CLS, thus implicating Sko1 in the regulation of their CLS.
Source: Bioscience, Biotechnology, and Biochemistry, 83(8), 1473-1476 (2019)

18. Analysis of metabolites in Japanese alcoholic beverage sake made from the sake rice Koshitanrei

Author: Eri ICHIKAWA, Shougo HIRATA, Yuko HATA, Hisashi YAZAWA, Hiroyasu TAMURA, Mitsuoki KANEOKE, Kazuhiro IWASHITA, Dai HIRATA
Abstract: In sake brewing, the steamed rice is used in two ways, added to sake-mash (as kake-mai) and making koji. The rice is an important determinant for the quality of sake, as the metabolites in sake affect its taste/aroma. The sake rice Koshitanrei (KOS) was developed in Niigata Prefecture by genetically crossing two sake rice, Gohyakumangoku and Yamadanishiki. However, the metabolites in sake from KOS have not been analyzed. Here, to investigate the characteristic metabolites in sake from KOS, we performed two types of small-scale sake fermentation tests changing only the rice used for kake-mai or total rice (both kake-mai and koji) by these three rice cultivars and examined the effect of KOS on sake metabolites by the metabolome analysis method using UPLC-QTOF-MS. We identified the peaks/metabolites, whose intensity in sake from KOS was higher/lower than those from the other cultivars. The brewing properties of KOS were partially characterized by this analysis.
Source: Bioscience, Biotechnology, and Biochemistry, 83, 1570-1582 (2019)

19. Genome editing to generate nonfoam-forming sake yeast strains

Author: Shunsuke Onuki, Mao Kashima, Toshikazu Yamada, Farzan Ghanegolmohammadi, Yan Zhou, Tetsuya Goshima, Jun-ichi Maruyama, Katsuhiko Kitamoto, Dai Hirata, Takeshi Akao, Yoshikazu Ohya
Abstract: Mutations frequently occur during breeding of sake yeasts and result in unexpected phenotypes. Here, genome editing tools were applied to develop an ideal nonfoam-forming sake yeast strain, K7GE01, which had homozygous awa1△/awa1△ deletion alleles that were responsible for nonfoam formation and few off-target mutations. High-dimensional morphological phenotyping revealed no detectable morphological differences between the genome-edited strain and its parent, while the canonical nonfoam-forming strain, K701, showed obvious morphological changes. Small-scale fermentation tests also showed differences between components of sake produced by K7GE01 and K701. The K7GE01 strain produced sake with significant differences in the concentrations of ethyl acetate, malic acid, lactic acid, and acetic acid, while K701 produced sake with more differences. Our results indicated genuine phenotypes of awa1△/awa1△ in sake yeast isolates and showed the usefulness of genome editing tools for sake yeast breeding.
Source: Bioscience, Biotechnology, and Biochemistry, 83 (8), 1583-1593 (2019)

20. Breeding of a cordycepin-resistant and adenosine kinase-deficient sake yeast strain that accumulates high levels of S-adenosylmethionine

Author: Muneyoshi Kanai, Norito Yasuda, Tomoko Morimoto, Satoko Yoshida, Nahoko Nishibori, Masaki Mizunuma, Tsutomu Fujii & Haruyuki Iefuji
Abstract: Adenosine kinase (ADO1)-deficient mutants can be obtained from cordycepin-resistant strains, and the disruption of ADO1 causes S-adenosylmethionine (SAM) accumulation. To breed a high-SAM-accumulating yeast strain without genetic manipulation for industrial purposes, we bred a cordycepin-resistant strain using sake yeast kyokai No. 9 as the parent strain with a mutation in adenosine kinase (ADO1) and acquired high-SAM-accumulating strain. In the bred strain (NY9-10), a single mutation (T258I) was present in the ADO1, and this mutation site is an ATP binding site and is highly conserved during evolution. Moreover, it was suggested that high accumulation of SAM and cordycepin resistance in NY9-10 was due to functional deficiency of ADO1 by this mutation. This strain is not a genetically-modified organism and can be employed for use in the food and medicine industry such as mass production and sake making.
Source: Bioscience, Biotechnology, and Biochemistry, 83(8), 1530-1537 (2019)

21. Investigation of relationship between sake-making parameters and sake metabolites using a newly developed sake metabolome analysis method

Author: Hisashi Yazawa, Masafumi Tokuoka, Hajime Kozato, Yutaro Mori, Miyuki Umeo, Rieko Toyoura, Ken Oda, Hisashi Fukuda, and Kazuhiro Iwashita
Abstract: We developed a sake metabolome analysis method using liquid chromatography-quadrupole/time-of-flight mass spectrometry to investigate the metabolome of various types of sakes and other alcohol beverages. Our method identified 198 compounds by comparison with standard metabolites. Using this method, we investigated the relationship between several sake-making parameters and sake metabolites by conducting combination experiments of these parameters using small-scale fermentation. The results indicated that all parameters significantly affected sake metabolites (P < 0.005) and most peaks were affected by multiple sake-making parameters. Interestingly, the effect of the rice cultivar on sake metabolites was higher for koji rice than for kake-rice. This result suggests that the rice cultivar used has a greater effect on the characteristics of Aspergillus oryzae compared to sake yeast and affects sake metabolites. In this study, we also evaluated the combined effect of several parameters. We demonstrated the different effects of each parameter on several amino acids. The results showed a new aspect of the science of sake making. For example, the amount of α-ethylglucoside, which can affect the taste of sake, was negatively correlated with α-glucosidase activity in koji (r=-0.84). In this study, various unidentified peaks were observed; detectable peaks can be increased by analyzing additional standard reagents. Investigating these unidentified peaks and accumulating datasets for sake-making parameters will give us insight into how to improve sake taste and quality.
Source: Journal of bioscience and bioengineering, 128, 183-190 (2019)

22. Increase in the Oxygen Stable Isotopic Composition of Water in Wine with Low Ethanol Yield

Author: Fumikazu AKAMATSU, Hideaki SHIMIZU, Aya KAMADA,Yukari IGI, Tsutomu FUJII, Nami GOTO-YAMAMOTO
Abstract: The stable isotopic composition of oxygen (δ¹⁸O) in wine is often analysed to determine the geographic origin of the wine and the amount of water dilution. However, little is known regarding the effects of two major winemaking techniques (the addition of acid (acidification) and sugar (chaptalization)) on theδ¹⁸O value of water in wine. Here we show that acidification and chaptalization have minor direct effects on theδ¹⁸O value but indirect effects based on the ethanol yield, which causes isotopic variation of up to 0.6‰. During fermentation,δ¹⁸O values increase at low ethanol yields, suggesting that yeast release water with a highδ¹⁸O value into wine when consuming sugars. Additionally, the ethanol yield is negatively correlated with the consumption of amino acids by the yeast, indicating that yeast growth decreases the ethanol yield. We therefore identify ethanol yield, which is decreased by the consumption of sugars by yeast for non-alcohol-fermentation processes as a potential factor leading to variations in theδ¹⁸O value of water during the winemaking process.
Source: Scientific Reports, 9, 11039 (2019)

23. Identification of enzymes from genus Trichoderma that can accelerate formation of ferulic acid and ethyl ferulate in collaboration with rice koji enzyme in sake mash.

Author: Toshihiko Ito, Anna Sato, Itsuki Takahashi, Takahito Ito, Kouto Takano, Koji Noge, Masaki Okuda, Katsumi Hashizume
Abstract: The enzymes responsible for acceleration of ferulic acid and ethyl ferulate formation in sake mash were studied. Ferulic acid and ethyl ferulate are formed during the sake brewing process from feruloylated glucuronoarabinoxylan. Cellulase reagent from genus Trichoderma was used instead of rice koji, because rice koji for sake brewing produces extremely low levels of xylan-degrading enzymes. A combination of the reagent with rice koji enzymes accelerated the formation of ferulic acid from a-rice powder. Addition of the reagent to sake mash increased ferulic acid and ethyl ferulate formation. The enzyme responsible for the accelerated formation was purified using a newly developed assay method and a-rice powder as a substrate. During the assay procedure, feruloylated oligosaccharide was converted to ferulic acid by feruloylesterase for HPLC analysis. Analysis of the N-terminal amino acid sequence of the purified samples was successfully conducted after pyroglutamyl aminopeptidase de-blocking. Purified enzymes were identified as members of the glycoside hydrolase family 10 (GH10) and family 11 (GH11) xylanases by BLASTP database research. The GH10 xylanase showed higher specific activity for a-rice powder and insoluble wheat arabinoxylan compared with GH11 xylanase; the GH11 xylanase showed higher specific activity for the other xylan substrates, especially glucuronoarabinoxylan.The GH10 xylanase showed higher accelerating activity than the GH11 xylanase in the sake mash. The results of this study provides useful knowledge on ferulic acid and ethyl ferulate formation in sake mash, the relative levels of these compounds and their influence on the sensory quality of sake.
Source: Journal of Bioscience and Bioengineering 128,177-182(2019)

24. Study for practical application of supported gold nanoparticles for removal of DMTS responsible for hineka in sake

Author: Atsuko Isogai, Haruno Murayama, Moemi Kimura, Takaaki Shinozaki, Eiji Yamamoto, Tsutomu Fujii, Sachiko Iizuka, Makoto Tokunaga
Abstract: Dimethyl trisulfide (DMTS) is one of the primary components responsible for "hineka", the stale aroma of Japanese sake. We have reported that gold nanoparticles supported on silica (Au/SiO2) were effective for the selective removal of DMTS by adsorption from model ethanol solutions and sake. In this study, we carried out the adsorptive desulfurization of several kinds of sake: daiginjo-shu, junmai-shu, and jukusei-shu (long aged sake) in 5 L-scales. The results of instrumental analyses and sensory evaluations suggested that the effect of Au/SiO2 was most remarkable for the daiginjo sample containing a high level of ginjo-ka components and DMTS; DMTS was significantly reduced while ginjo-ka remained. For the practical application of Au/SiO2, we established a large-scale production by optimizing the calcination conditions using a rotary electric furnace. We also confirmed that Au/SiO2 can be reused by heating at 300°C.
Source: J. Brew. Soc. Japan, 114, 779-786 (2019)

25. Effect of Vineyard Altitude on Phenological Events and Berry Components of 'Chardonnay' and 'Merlot' in Yamanashi

Author: Eri INOUE, Munekazu KISHIMOTO, Manabu NEMOTO, Koki WATANABE, Hiroshi SAITO, Kazuya KOYAMA, Nami GOTO-YAMAMOTO, Hidetoshi MINO, Tomohiro SUZUKI and Tohru OKUDA
Abstract: The aim of this study was to investigate the relationships between vineyard altitude and climate classification, phenological events, and berry components. Vineyards located at an altitude of 250 m to 750 m where 'Chardonnay' and 'Merlot' grapevines are cultivated in Yamanashi Prefecture were classified into regions III to V on the basis of the Winkler Index. As the altitude increased, budburst, flowering, veraison or coloration, and harvest were delayed, whereas malic acid and anthocyanin contents increased. The difference in air temperature owing to vineyard altitude greatly affected both phenological events and berry components.
Source: Journal of ASEV Japan, 30(3), 111-120, 2019

26. Characterization of a wild sake-brewing yeast isolated from barley malt in Toyama

Author: Hiroyasu Onaka, Jun-ichi Maruyama, Shumpei Asamizu, Mayumi Kuroiwa, Katsuhiko Kitamoto, Masato Yamada, Tetsuya Goshima, Takeshi Akao
Abstract: A wild sake-brewing yeast, Toyama yeast (Toyama-Umare-no-Koubo) was isolated from the malt of six-rowed barley harvested in Takaoka, Toyama. Molecular phylogenetic analysis revealed that Toyama yeast had the highest homology with Saccharomyces cerevisiae NBRC2114 (Kotobukiya whisky No. 1) in the internal transcribed spacers (ITS) region. Small-scale sake brewing test revealed that the mash (moromi) fermentation period for Toyama yeast was 3 days more than that for K701 yeast, and the alcohol concentration reached almost the same i.e. 18%. In the composition of organic acids in sake fermented by Toyama yeast, the amounts of acetic acid, malic acid, and succinic acid were considerably different from sake fermented by K701. Considering volatile contents, sake fermented by Toyama yeast also contained more higher alcohols and less esters than K701 sake. In amino acid content, sake fermentated by Toyama yeast was much higher than K701. These properties also appeared in organoleptic evaluation. Further, Toyama yeast sake has unique and distinctive properties characterized by differences in aroma and flavor such as acid smell, umami, heavy, zatsumi, and thin as compared to K701 sake. Commercialized brewing with Toyama yeast has been done five times in Narimasa Shuzo Breweries, the current brewing sake has become rich in flavor and has a sharp finish. Also, since this yeast is not a sake yeast but a wild yeast, it has been also used for the brewing of craft beer, wine, and whiskey.
Source: J. Brew. Soc. Japan, 114 (10) 645-653 (2019)