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

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.     

Late-maturing cooking rice Sensyuraku has excellent properties,equivalent to sake rice,for high-quality sake brewing

Low protein content and sufficient grain rigidity are desired properties for the rice used in high-quality sake brewing such as Daiginjo-shu (polishing ratio of the rice, less than 50%). Two kinds of rice, sake rice (SR) and cooking rice (CR), have been used for sake brewing. Compared with those of SR, analyses of CR for high-quality sake brewing using highly polished rice have been limited. Here we described the original screening of late-maturing CR Sensyuraku (SEN) as rice with low protein content and characterization of its properties for high-quality sake brewing. The protein content of SEN was lower than those of SR Gohyakumangoku (GOM) and CR Yukinosei (YUK), and its grain rigidity was higher than that of GOM. The excellent properties of SEN with respect to both water-adsorption and enzyme digestibility were confirmed using a Rapid Visco Analyzer (RVA). Further, we confirmed a clear taste of sake produced from SEN by sensory evaluation. Thus, SEN has excellent properties, equivalent to those of SR, for high-quality sake brewing.     

Effect of koji starter on metabolites in Japanese alcoholic beverage sake made from the sake rice Koshitanrei

ABSTRACT

In sake brewing, the steamed rice is used in 2 ways, added to sake-mash and making rice-koji. Rice-koji is made from the steamed rice by using koji starter, and its quality is an important determinant of the aroma/taste of sake. The sake rice Koshitanrei (KOS) was developed in Niigata Prefecture by crossing 2 sake rice varieties, Gohyakumangoku and Yamadanishiki. Recently, we reported the characteristic components/metabolites in sake made from KOS by conducting metabolome analysis using UPLC-QTOF-MS. In this study, to investigate the effect of koji starter and sake rice cultivars on the sake metabolites, we performed small-scale sake-making tests using the above 3 rice cultivars and 3 koji starters. Finally, we demonstrated that some of the characteristic components/metabolites of sake from KOS are affected by the koji starter. Thus, in addition to rice cultivar, koji starter plays an important role for establishment/maintenance of the quality of the final product.     

Host influence on germination and reproduction of the facultative hemi‐parasitic weed Rhamphicarpa fistulosa

Rice Vampireweed, Rhamphicarpa fistulosa, was a minor parasitic weed until recently when rice cultivation in sub‐Saharan Africa was expanded into marginal wetlands, that are the parasite's natural habitat. Unlike most of the parasitic weeds, R. fistulosa is facultative, meaning that the parasite is able to complete its life cycle without a host. However, when not connected to a host plant, its biomass and seed production is lower. Because very little is known regarding the germination ecology of the parasite, the main objective of our study was to identify the cues that favour germination. We hypothesised that, first, being a wetland species, germination of R. fistulosa is stimulated by light and high soil moisture. Second, we hypothesised that if host plant presence increases its reproductive output then a germination stimulatory effect from host presence is likely to have developed. A Petri‐dish and pot experiment showed that light and completely saturated soils were a requirement for germination, demonstrating that germination requirements of R. fistulosa are typical of species that grow in environments with fluctuating water levels. A pot experiment in which five infestation levels of R. fistulosa were installed in the absence and presence of a rice plant, showed that host plant presence resulted in a 3.7 times higher seed production rate and a 15% larger average seed size. Despite this reproductive advantage, a pot experiment with three rice cultivars, selected because of their difference in strigolactone production, showed that host plant presence, regardless of the development stage, did not influence the emergence rate of R. fistulosa. In a follow‐up study, the germination stimulation effect of root exudates collected from the same three rice cultivars and a treatment consisting of an artificial germination stimulant (GR24) was compared with a treatment consisting of plain water. In these treatments, seeds of R. fistulosa were compared with seeds of the obligate parasite Striga hermonthica. Germination of S. hermonthica was strongly advanced by the presence of root exudates and GR24 but was completely absent in water, whereas germination of R. fistulosa in all treatments was similar to that in plain water. The absence of a host recognition mechanism at the germination stage suggests that the regulation of germination through light and soil moisture is near optimal. Our finding might also indicate that for this facultative parasitic plant species, a more opportunistic germination strategy is superior. Implications of the findings for management of R. fistulosa in rice cultivation are discussed.     

Agronomic aspects of wetland rice cultivation and associated methane emissions

Wetland rice cultivation is one of the major sources of atmospheric methane (CH₄). Global rice production may increase by 65% between 1990 and 2025, causing an increase of methane emissions from a 92 Tg CH₄ y^–1 now to 131 Tg in 2025.Methane production depends strongly on the ratio oxidizing: reducing capacity of the soil. It can be influenced by e.g. addition of sulphate, which inhibits methanogenesis. The type and application mode of mineral fertilizers may also affect methane emissions. Addition of organic matter in the form of compost or straw causes an increase of methane emissions, but methane production is lower for materials with a low C/N ratio.High percolation rates in wetland rice soils and occasional drying up of the soil during the cultivation period depresses methane release. Water management practices aimed at reducing emissions are only feasible during specific periods in the rice growing season in flat lowland irrigated areas with high security of water availability and good control of the water supply. Intermittent drying of soils may not be possible on terraced rice lands.Assuming a 10 to 30% reduction in emission rates per unit harvested area, the global emission may amount to 93 Tg CH₄ y^– in 2025. A reduction of global emissions seems very difficult. To develop techniques for reducing CH₄ emissions from wetland rice fields, research is required concerning interactions between soil chemical and physical properties, and soil, water and crop management and methanogenesis. Such techniques should not adversely affect rice yields.     

Development of region-specific emission factors and estimation of methane emission from rice fields in the East, Southeast and South Asian countries

Rice cultivation areas in East, Southeast and South Asia account for 89% of the world total, and field measurements of methane (CH₄) emission from rice cultivation have been widely performed in this area. In this paper, we assembled most of the measurements and developed region‐specific CH₄ emission factors. Efforts were made in order to regionalize rice fields by climate and soil properties, and to incorporate the effect of organic input and water regime on emission. Data on rice cultivation areas of 1995 were collected at subdivision level (province, state, prefecture, etc.). Total emission from these areas was estimated at 25.1 Tg CH₄ year ^{? 1}, of which 7.67 Tg was emitted from China and 5.88 Tg from India. Irrigated and rainfed rice fields contributed 70.4 and 27.5% to the total emission, respectively. Deepwater rice fields had a very small share. A high‐resolution and quality emission distribution map was constructed as the emission was directly estimated at province level and below that, a 30‐second land‐use dataset was used in order to translate the emission to grid format. As the rice cultivation area in the study region accounts for 89% of the world total, extrapolating the estimate to the global scale indicates a global emission of 28.2 Tg CH₄ year ^{? 1}. The estimate was compared with country reports made by local scientists. For some countries – such as Indonesia, Myanmar, Thailand, Vietnam, Japan, South Korea, Pakistan and the Philippines – the results of this estimate agree reasonably well with their country reports (CV < 15%). For some other countries – such as China, India and Bangladesh – there is relatively large disagreement between our estimate and their country reports. The reasons for the discrepancies were discussed.     

Patterns of bird abundance and habitat use in rice fields of the Kanto Plain, central Japan

Rice fields, the most widespread cropland in Japan, provide important habitat for birds, but few studies have examined bird communities that are influenced by agriculture in rice fields. I studied the seasonal pattern of bird abundance and their habitat preferences in terms of agricultural habitats in the rice fields of the central Kanto Plain, central Japan. In total, 19 waterbird species and 31 landbird species were recorded over the study year. Waterbird occurrence was largely restricted to the cultivation season (i.e. the flooded season, May–August), with maximum abundance in August. Rice fields dried up during winter and supported few waterbirds except for irregular occurrences of little egrets (Egretta garzetta) and common snipes (Gallinago gallinago), which selectively used ditches with shallow water. Landbird abundance increased after the start of cultivation (May) to August, but more birds were attracted to the rice fields in midwinter (around January). For both waterbirds and landbirds, most species preferred levees and fallow grass fields, but avoided fields with rice vegetation. Unplowed fields after harvest were preferred by several species including skylarks (Alauda arvensis), while plowed fields were less attractive to most species. These results suggest the need for advanced agricultural practices to include improvement in habitat quality for birds.     

Identification of l-Dopa and Protocatechuic Acid as Main Precursor of Pigment of Brown Rice-koji

The rice koji for sake making usually shows white, but sometimes, during storage in the air it turns brown. This turn is accelerated by some wet conditions.

Three kinds of precursor of the pigment in browned rice-koji were isolated in the present work by column chromatography. A main precursor was adsorbed on alumina at pH 8.5 and eluted with 0.3 n acetic acid. The eluate was then treated by Dowex 50-X8 (H) column, eluted with water and 1 n HCl, and then identified as l-dopa after derived to 3,4-di-O-benzoyl-N-benzoyl-l-dopa ethyl ester. The second precursor was isolated from ethylether soluble fraction of the water eluate described above and identified as protocatechuic acid by some chemical properties. The third precursor was not isolated owing to its chemical unstableness, though it was clearly different from these two precursors.

Isolation of l-dopa from the rice koji made by culture of Aspergillus and also from sake and sake cake was first carried out in the present work.     

Seasonal changes in fluxes of methane and volatile sulfur compounds from rice paddies and their concentrations in soil water

Rice paddies emit not only methane but also several volatile sulfur compounds such as dimethyl sulfide (DMS: CH₃SCH₃). However, little is known about DMS emission from rice paddies. Fluxes of methane and DMS, and the concentrations of methane and several volatile sulfur compounds including hydrogen sulfide (H₂S), carbonyl disulfide (CS₂), methyl mercaptan (CH₃SH) and DMS in soil water and flood water were measured in four lysimeter rice paddies (2.5 × 4 m, depth 2.0 m) once per week throughout the entire cultivation period in 1995 in Tsukuba, Japan. The addition of exogenous organic matter (rice straw) was also examined for its influence on methane or DMS emissions. Methane fluxes greatly differed between treatments in which rice straw had been incorporated into the paddy soil (rice straw plot) and plots without rice straw (mineral fertilizer plot). The annual methane emission from the rice straw plots (37.7 g m^-2) was approximately 8 times higher than that from the mineral fertilizer plots (4.8 g m^-2). Application of rice straw had little influence on DMS fluxes. Significant diurnal and seasonal changes in DMS fluxes were observed. Peak DMS fluxes were found around noon. DMS was emitted from the flood water in the early growth stage of rice and began to be emitted from rice plants during the middle stage. DMS fluxes increased with the growth of rice plants and the highest flux, 15.1 µg m^-2 h^-1, was recorded before heading. DMS in the soil water was negligible during the entire cultivation period. These facts indicate that the DMS emitted from rice paddies is produced by metabolic processes in rice plants. The total amount of DMS emitted from rice paddies over the cultivated period was estimated to be approximately 5–6 mg m^-2. CH₃SH was emitted only from flood water during the first month after flooding.     

Assessment of the methane mitigation potentials of alternative water regimes in rice fields using a process‐based biogeochemistry model

Rice production is a substantial source of atmospheric CH₄, which is second only to CO₂ as a contributor to global warming. Since CH₄ is produced in anaerobic soil environments, water management is expected to be a practical measure to mitigate CH₄ emissions. In this study, we used a process‐based biogeochemistry model (DNDC‐Rice) to assess the CH₄ mitigation potentials of alternative water regimes (AWR) for rice fields at a regional scale. Before regional application, we tested DNDC‐Rice using site‐scale data from three rice fields in Japan with different water regimes. The observed CH₄ emissions were reduced by drainage of the fields, but were enhanced by organic amendments. DNDC‐Rice gave acceptable predictions of variation in daily CH₄ fluxes and seasonal CH₄ emissions due to changes in the water regime. For regional application, we constructed a GIS database at a 1 × 1 km mesh scale that contained data on rice field area, soil properties, daily weather, and farming management of each cell in the mesh, covering 3.2% of the rice fields in Japan's Hokkaido region. We ran DNDC‐Rice to simulate CH₄ emissions under five simulated water regimes: the conventional water regime and four AWR scenarios with gradually increasing drainage. We found that AWR can reduce CH₄ emission by up to 41% compared with the emission under conventional water regime. Including the changes in CO₂ and nitrous oxide emissions, potential mitigation of greenhouse gas (GHG) was 2.6 Mg CO₂ Eq. ha^?1 yr^?1. If this estimate is expanded to Japan's total rice fields, expected GHG mitigation is 4.3 Tg CO₂ Eq. yr^?1, which accounts for 0.32% of total GHG emissions from Japan. For a reliable national‐scale assessment, however, databases on soil, weather, and farming management must be constructed at a national scale, as these factors are widely variable between regions in Japan.     

Cell biology of the Koji mold Aspergillus oryzae

Koji mold, Aspergillus oryzae, has been used for the production of sake, miso, and soy sauce for more than one thousand years in Japan. Due to the importance, A. oryzae has been designated as the national micro-organism of Japan (Koku-kin). A. oryzae has been intensively studied in the past century, with most investigations focusing on breeding techniques and developing methods for Koji making for sake brewing. However, the understanding of fundamental biology of A. oryzae remains relatively limited compared with the yeast Saccharomyces cerevisiae. Therefore, we have focused on studying the cell biology including live cell imaging of organelles, protein vesicular trafficking, autophagy, and Woronin body functions using the available genomic information. In this review, I describe essential findings of cell biology of A. oryzae obtained in our study for a quarter of century. Understanding of the basic biology will be critical for not its biotechnological application, but also for an understanding of the fundamental biology of other filamentous fungi.     

Efficacy of Chlorella pyrenoidosa and Scenedesmus abundans for Nutrient Removal in Rice Mill Effluent (Paddy Soaked Water)

Microalgae are product of sustainable development owing to its ability to treat variety of wastewater effluents and thus produced biomass can serve as value added product for various commercial applications. This paper deals with the cultivation of microalgae species namely Chlorella pyrenoidosa and Scenedesmus abundans in rice mill effluent (i.e., paddy soaked water) for nutrient removal. In order to investigate the nutrient removal capability, microalgae are subjected to cultivation in both raw and autoclaved samples. The maximum phosphate removal by Scenedesmus abundans and Chlorella pyrenoidosa in raw sample was 98.3% and 97.6%, respectively, whereas, the removal of ammoniacal nitrogen by Scenedesmus abundans and Chlorella pyrenoidosa in raw sample was 92% and 90.3%, respectively. The growth (measured in terms of chlorophyll content) of Scenedesmus abundans and Chlorella pyrenoidosa in raw sample was 3.88 mg/l and 5.55 mg/l, respectively. The results indicate the suitability of microalgae cultivation in rice mill effluent treatment for nutrient removal.     

A review of studies and measures to improve the mycotoxicological safety of traditional Japanese mold-fermented foods

Miso (fermented soybean paste), shoyu (soy sauce) and sake (rice wine) are traditional moldfermented foods in Japan and have been consumed throughout much of its history. These have long been considered safe foods. In this contribution we review and summarize long-term studies to investigate potential problems with mycotoxin contamination of these products. The fungal cultures used for fermentation of these products are called “koji-molds” and mainly consist of strains ofAspergillus oryzae. A. oryzae belongs to theA. flavus group taxonomically, which is generally known to be a main producer of aflatoxins. Therefore, we studied the productivity of aflatoxins by various koji-molds, as well as the possibility of aflatoxin contamination of rice (which is used in the production of fermented foods), miso, shoyu and sake. Rice was found to be free from aflatoxins. Furthermore, none of the tested koji-molds produced any detectable levels of aflatoxins, consequently no aflatoxins were found in miso, shoyu, or sake. However, some koji-molds are known to produce cyclopiazonic acid (CPA) and kojic acid (KA). We studied the production of CPA and KA by various commercial koji-molds and identified some strains that produce relatively high amounts of CPA or KA. Consequently, we advised food industry not to use these strains. Although mycotoxin contamination of these products is therefore presently very low, further attempts should be made to completely eliminate CPA and KA from fermented foods.     

海南中南部地区旱稻(山兰稻)种质资源及保育模式

山兰稻是一类适应性很强的旱稻品种,对于开发节水性高产稻种具有重要意义。该研究调查了海南中南部地区山兰稻种质资源的组成,并选种5个山兰稻品种开展其农艺性状和生产性状的研究,以期了解山兰稻种质资源在该地区的分布及生产现状,并进一步为其保护模式提供理论基础。结果表明:调查共获得38份山兰稻种质资源,含25个品种,主要分布在保亭、琼中和五指山三个市县,在现有的品种中约有45.0%在种植利用。因此,山兰稻品种资源面临濒危,亟需保护;这25个品种可分为籼稻和粳稻2个类型,共聚成6个类群,不同区域间的种质资源具有亲缘性,其中3个独立聚集的品种有明显区域性分布的特征;5个品种容易种植,生产性状较一致,其全生育期约为138 d,与历史栽培记录时间一致,遗传特性稳定,单位产量为2.0~2.8t·hm-2,平均产量为2.4 t·hm-2。综上说明保护山兰稻品种的成本和技术难度不高,但其育种和栽培技术还需要进一步研究,种质资源的开发模式应结合生态保护、品种保育、社会需求等多角度进行综合考虑。     

The impact of changing irrigation practices in rice fields on frog populations of the Kanto Plain, central Japan

Rice fields are important substitute wetlands for frogs. Traditionally, rice fields in Japan were supplied with water and drained via shallow earth ditches (old-style). In the last 30 years, however, more than 80% of rice fields have been converted to a new irrigation system in which water is typically supplied through underground pipes via taps and is drained into deep, concrete-sided ditches (new-style). We compared the occurrence of frogs in paired areas of old- and new-style rice fields at six locations in Ibaraki Pref., central Japan, from May to August 1995. The Japanese tree frog (Hyla japonica) did not differ in abundance between the two types of rice fields, but the Japanese brown frog (Rana japonica) and the Tokyo daruma pond frog (Rana porosa porosa) preferred the old-style rice fields. These findings suggest that the status ofRana species has been adversely affected by the conversion of rice fields to the new irrigation system. Modification of the deep, U-shaped concrete ditches and the water management regime during the spawning season is needed to safeguard these species.     

Establishment of a cell-free translation system from rice callus extracts

ABSTRACT

Eukaryotic in vitro translation systems require large numbers of protein and RNA components and thereby rely on the use of cell extracts. Here we established a new in vitro translation system based on rice callus extract (RCE). We confirmed that RCE maintains its initial activity even after five freeze-thaw cycles and that the optimum temperature for translation is around 20°C. We demonstrated that the RCE system allows the synthesis of hERG, a large membrane protein, in the presence of liposomes. We also showed that the introduction of a bicistronic mRNA based on 2A peptide to RCE allowed the production of two distinct proteins from a single mRNA. Our new method thus facilitates laboratory-scale production of cell extracts, making it a useful tool for the in vitro synthesis of proteins for biochemical studies.     

The role of rice cultivation in changes in atmospheric methane concentration and the Global Methane Pledge

Resumption of the increase in atmospheric methane (CH₄) concentrations since 2007 is of global concern and may partly have resulted from emissions from rice cultivation. Estimates of CH₄ emissions from rice fields and abatement potential are essential to assess the contribution of improved rice management in achieving the targets of the Global Methane Pledge agreed upon by over 100 countries at COP26. However, the contribution of CH₄ emissions from rice fields to the resumed CH₄ growth and the global abatement potential remains unclear. In this study, we estimated the global CH₄ emissions from rice fields to be 27 ± 6 Tg CH₄ year⁻¹ in the recent decade (2008–2017) based on the 2019 Refinement to the 2006 IPCC Guidelines for National Greenhouse Gas Inventories. The trend of CH₄ emissions from rice cultivation showed an increase followed by no significant change and then, a stabilization over 1990–2020. Consequently, the contribution of CH₄ emissions from rice fields to the renewed increase in atmospheric CH₄ concentrations since 2007 was minor. We summarized the existing low-cost measures and showed that improved water and straw management could reduce one-third of global CH₄ emissions from rice fields. Straw returned as biochar could reduce CH₄ emissions by 12 Tg CH₄ year⁻¹, equivalent to 10% of the total reduction of all anthropogenic emissions. We conclude that other sectors than rice cultivation must have contributed to the renewed increase in atmospheric CH₄ concentrations, and that optimizing multiple mitigation measures in rice fields could contribute significantly to the abatement goal outlined in the Global Methane Pledge.     

Emerging Molecular Strategies for Improving Rice Drought Tolerance

Rice occupies a pre-eminent position as a food crop in the world. Its production, how- ever, entails up to 3000 liters of water per kilogram of grain produced. Such high demand makes rice prone to drought easily. Sustainable rice cultivation with limited water resources requires the deployment of a suitable strategy for better water use efficiency and improved drought tolerance. Several drought-related genes have been evaluated in rice for their mode of action in conferring drought tolerance. Manipulation of components of abscisic acid signal transduction, stomatal density, deposition of cuticular wax, and protein modification pathways are emerging as priority targets. Gene reprogramming by microRNAs is also being explored to achieve drought tolerance. Genetically dissected Quantitative Trait Loci (QTLs) and their constituent genes are being deployed to develop drought-tolerant rice varieties. Progressive research and challenges include a better understanding of crucial components of drought response and search for new targets and the deployment of improved varieties in the field.     

控释氮肥对抗除草剂转基因水稻田土壤甲烷排放的影响

采用温室盆栽和静态箱-气相色谱法,研究了控释氮肥对抗除草剂转基因水稻和亲本常规水稻稻田土壤甲烷(CH4)排放的影响。供试土壤为潴育型水稻土,氮肥种类为尿素和控释氮肥。结果表明,与对照(尿素)相比,控释氮肥提高了水稻分蘖数、株高、生物量及产量。水稻品种对CH4季节性排放规律没有明显影响,CH4排放通量基本表现为,自水稻移栽后逐渐升高,移栽后62—92 d出现峰值,而后逐渐降低至水稻收获。与对照相比,控释氮肥可显著降低CH4排放通量和全生育期累积排放量。抗除草剂转基因水稻稻田土壤CH4排放通量和累积排放量均显著低于亲本常规水稻。研究认为,一次性基施控释氮肥和种植抗除草剂转基因水稻对有效减缓稻田甲烷排放具有重要意义。     

Approaches to breeding for salinity tolerance - a case study on Porteresia coarctata

Cereals are the world's major source of food for human nutrition. Among these, rice (Oryza sativa) is the most prominent and represents the staple diet for more than two-fifths (2.4 billion) of the world's population, making it the most important food crop of the developing world (Anon., 2000a). Rice production in vast stretches of coastal areas is hampered due to high soil salinity. This is because rice is a glycophyte and it does not grow well under saline conditions. In order to increase rice production in these areas there is a need to develop rice varieties suited to saline environments. Research has shown that Porteresia coarctata, a highly salt tolerant wild relative of rice growing in estuarine soils, is an important material for transferring salt tolerant characteristics to rice. It is quite possible that Porteresia may be used as a parent for evolving better and truly salt resistant varieties. The inadequate results and the difficulties associated with conventional breeding techniques necessitate the use of the tools of crop biotechnology in unravelling some of the characteristics of Porteresia that have been highlighted in this report. In view of the limited resources available for increasing salinity tolerance to the breeders to wild rice germplasm, Porteresia is undoubtedly one of the key source species for elevating salinity tolerance in cultivated rice.