Ac ce pt ed m an us cr ip t Mongolian Journal of Chemistry Page 1 of 15 Hydrothermal treatment of rice straw for carbohydrate production Enkhtur Munkhbat1, Zhongfang Lei2 1Institute of Chemistry and Chemical Technology, Mongolian Academy of Sciences, Ulaanbaatar 13330, Mongolia 2Graduate School of Life and Environmental Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8572, Japan *Author to whom correspondence should be addressed Enkhtur Munkhbat Institute of Chemistry and Chemical Technology Mongolian Academy of Sciences Ulaanbaatar, Mongolia E-mail: munkhbate@mas.ac.mn ORCID: https://orcid.org/0000-0002-0894-9929 This article has been accepted for publication and undergone full peer review but has not been through the copyediting, typesetting, and proofreading process, which may lead to differences between this version and the official version of record. Please cite this article as: Munkhbat E, Lei Z. Hydrothermal treatment of rice straw for carbohydrate production. Mongolian Journal of Chemistry, 24(50), 2023, https://doi.org/10.5564/mjc.v24i50.2425 User Typewriter xx User Typewriter mailto:munkhbate@mas.ac.mn Ac ce pt ed m an us cr ip t Mongolian Journal of Chemistry Page 2 of 15 Hydrothermal treatment of rice straw for carbohydrate production Enkhtur Munkhbat1, https://orcid.org/0000-0002-0894-9929 Zhongfang Lei2, https://orcid.org/0000-0002-0479-6179 3 1Institute of Chemistry and Chemical Technology, Mongolian Academy of Sciences, Ulaanbaatar 6 13330, Mongolia 2Graduate School of Life and Environmental Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8572, Japan 9 ABSTRACT This study focused on the effect of hydrothermal (HT) treatment at 180 – 210 °C for holding 0 - 15 min on the solubilization of rice straw and the changes of HT residue. The optimum 12 treatment conditions for the highest solubilization and solid reduction of rice straw was 210 °C for holding 0 min. Under this condition, the extraction yield and total organic carbon (TOC) concentration of the HT liquid part were the highest, about 44% and 7850 mg/L, respectively. 15 The dry residue showed that the HT conditions above 200 °C for holding a short time were more efficient, which was confirmed by FT-IR and the changes of surface morphology under microscope. The reactor headspace could be an important factor because HT treatment with 18 a lower headspace (HTp210-0(15)) yielded more soluble carbohydrate under the test conditions. Also, energy input calculated based on the 1 ton removed hemicellulose (extraction yield) in the headspace experiments proved this finding. 21 Keywords: lignocellulosic biomass; hydrothermal treatment; cellulose recovery; biomass 24 digestibility https://orcid.org/0000-0002-0894-9929 https://orcid.org/0000-0002-0479-6179 Ac ce pt ed m an us cr ip t Mongolian Journal of Chemistry Page 3 of 15 INTRODUCTION 27 Increasing energy usage and the rapid depletion of fossil fuels require renewable energy development to reduce pollutions generated by fossil fuels. According to the Energy Information Administration of the United States (EIA), worldwide energy demand will 30 increase by 40% by 2040, reaching about 800 quadrillion British thermal unit, with the rising countries accounting for the majority of the demand increases [1]. Due to its use as a fuel and other value-added chemical production, biomass or biofuel has attracted increasing 33 attention among the numerous renewable energy sources such as geothermal, hydro, wind, and solar [2]. The fact that lignocellulosic biomass makes up a large portion of plant matter makes it the 36 most plentiful renewable resource on earth. It is a desirable feedstock for making chemicals and fuels since it is accessible and affordable. The three main components of lignocellulosic biomass are cellulose (40–50%), lignin (15–20%), and hemicellulose (25–35%). Rice 39 straw’s recalcitrant nature is one of the challenges for its biochemical conversion to bioethanol and methane. To convert biomass to biofuel, cellulose and hemicellulose molecules must be broken down into monomers or simple sugars. Rice straw fermentation 42 is difficult in practice, which creates a significant barrier for lignocellulosic biomass in the bioconversion process [3]. In a sugar platform bio refinery, pretreatment is an important step in increasing biomass digestibility. Several criteria define the goal of any pretreatment 45 procedure: (1) maximizing the final yield such as ethanol and other valuable products; (2) high amount lignin removal; and (3) reducing the formation of degradation products that can inhibit the action of produce biofuels [3, 4]. There are four main types of treatments, i.e., 48 physical, chemical, physicochemical, and biological methods in the hydrolysis of lignocellulosic biomass, and some of these treatment methods are very effective. Many chemical, thermal, and biological pretreatment procedures have been extensively 51 researched to increase lignocellulosic biomass susceptibility to later enzymatic hydrolysis [5]. The use of water as the primary reaction medium with no other chemical additives makes hydrothermal pretreatment one of the most effective pretreatment techniques in terms of 54 both practical and environmental considerations [6]. Hydrothermal (HT) processing of lignocellulosic materials has been studied under a variety of operational conditions in the past. HT treatment operating temperatures are typically 57 between 100 and 230 °C, though higher temperatures can be used. The efficiencies vary depending on the applied temperature and time, which are co-related factors. Generally, 180 – 210 °C with a short holding time (1-15 minutes) can achieve the best sugar refinery 60 [7, 8]. Some researchers suggested HT treatment with some chemicals such as acid and Ac ce pt ed m an us cr ip t Mongolian Journal of Chemistry Page 4 of 15 alkali addition. According to Imman et al. [9], the carbohydrate yield from HT treatment with acid and alkali were about 30 - 40% higher than that without chemical addition. However, 63 adding chemicals is not regarded as environmentally friendly. The liquid to solid ratio (LSR) of solid concentrations can range from 2 to 100 (w/w), with the most typical values being around 10 [5]. The interaction between HT temperature and holding time has a significant 66 impact on the selection of both the liquid and solid phases. It is widely assumed that HT treatment at a higher temperature for a short time will result in slightly better pentoses yield and less inhibitor formation [10, 11]. HT treatment at 200 - 210 °C for a short period is 69 effective: When corn stover was hydrothermally treated at 210 °C for 0 and 10 minutes, more than 90% of the xylan was solubilized [12, 13]. One of the most critical aspects in the process economics of commercializing lignocellulosic biomass conversion is energy 72 consumption. That’s why energy balance analysis is very important. According to He et al. [13], HT pretreatment gained energy about 2741 MJ/t-rice straw when the process was performed at 150 °C for 20 minutes, which was 300 MJ/t-rice straw more compared to the 75 methane production from no pretreatment group. In addition, the energy recovery from the HT and microwave pretreatment was 43 - 53% and 57 - 79%, respectively [14]. Many researchers investigated the HT treatment on various types of lignocellulosic 78 materials, but there is little information about HT reactor head space's influence on sugar recovery. This research aimed to determine the suitable HT treatment conditions for rice straw to achieve digestible sugars which can be used for maximal ethanol production. 81 EXPERIMENTAL Materials: In this investigation, rice straw was collected from a farm area in Tsukuba 84 (Ibaraki-ken, Japan) and cut up into small pieces and then be air-dried. The air-dried rice straw particles were milled for the experiment, with particle sizes ranging from 0.27 to 0.56 mm. Before use, the milled straw was kept in a plastic container in the dark at ambient 87 temperature. The original rice straw used in this study contains 92.56% total solids, 50.3% total carbohydrates, 27.8% lignin and 10.44% ash. Apparatus and procedure: In a 200 mL stainless-steel reactor, HT treatment was 90 performed. Rice straw was treated at 4 temperature levels in the range of 180 – 210 °C for 0 min, 5 min, 10 min, and 15 min, respectively. The temperature in the HT reactor was increased at 12 °C/min on average, and the pressure was around 1 bar. In addition, when it 93 reached the holding time, the heater was powered off, and a table fan was used to cool it. The average cooling rate was 2 °C/min. Nine different HT treatment conditions were performed in this study, which were labelled as HT180-10, HT180-15, HT190-10, HT200-5, 96 Ac ce pt ed m an us cr ip t Mongolian Journal of Chemistry Page 5 of 15 HT200-10, HT210-0, HTP210-0(5), HTP210-0(9), HTP210-0(15). The first six experimental tests were to find out the suitable HT condition, and the last three experiments were to check whether the reactor pressure had any influence on the sugar yield. The installed pressure 99 meter was used to read the reactor pressure, which was around 1 - 2 MPa depending on HT conditions. The treated rice straw was centrifuged after HT treatment, and the solid HT residue was rinsed with distilled water. The pH value, total organic carbon (TOC), volatile 102 fatty acids (VFAs), and total carbohydrate of the isolated supernatant were all measured. After being washed with deionized water, the solid residue from HT was dried at 105 ºC for 24 hours and used to calculate the total yield based on the weight difference [15]. For future 105 usage, the pretreated dry biomass was packaged in plastic bags and stored in the dark. Analytical methods: The National renewable energy laboratory (NREL) method was used to determine total solid (TS), volatile solid (VS), and calculate yield [15]. The concentration 108 of total soluble carbohydrates was measured using the phenol sulfuric acid technique with glucose as reference [16]. A pH meter was used to determine the pH value. Individual VFA species in the liquid from rice straw during HT treatment was determined using gas 111 chromatography with a flame ionized detector (GC-8A, Shimadzu Corporation, Japan). VFAs were calculated as the sum of acetic, propionic, iso-butyric, n-butyric, iso-valeric, and n-valeric acids. A TOC analyzer was used to determine the total organic carbon (TOC) of 114 the HT liquid component (TOC-V CSN, Shimadzu, Japan). The modified method was used to determine the amounts of lignin, cellulose, and hemicellulose in HT treated dry biomass [15, 17]. In brief, 0.3 g of solid residue was mixed 117 with 3 mL of 72% w/w H2SO4 on a laboratory shaker for 4.5 hours at ambient temperature (25 °C). The solution was then diluted to 4% and hydrolyzed overnight to convert cellulose to glucose. The liquid and solid components were then separated using vacuum filtration, 120 and the solid part was dried at 105 °C for lignin analysis. Acid-soluble lignin and total carbohydrate were determined in the separated liquid. All the trials were done three times and the average results were presented. The structural 123 morphology of HT treated biomass and the raw rice straw were observed by optical microscopy. The structural modifications during the HT treatment were also investigated using an FT-IR spectrophotometer. 126 The energy consumption was estimated according to Eq. 1 [13]. The rice straw disposal capacity was expected to be 1 ton in this study. 129 𝐸𝐻𝑇 = 𝑚𝑤𝑎 𝛾𝑤𝑎(𝑇𝐻𝑇 − 𝑇𝑎𝑡 ) + 𝑚𝑟𝑠 𝛾𝑟𝑠(𝑇𝐻𝑇 − 𝑇𝑎𝑡 ) (1) Ac ce pt ed m an us cr ip t Mongolian Journal of Chemistry Page 6 of 15 where EHT (MJ) is the heat consumption by HT reactor; mrs (t) is the disposal capacity of rice straw; mwa (t) is the water usage; γwa is the specific heat capacity of water (4.18 kJ/kg 132 °C); γrs is the specific heat capacity of rice straw (1.67 kJ/kg °C); THT (°C) is the HT treatment temperature (180-210 °C in this study); Tat is the temperature of the environment (25 °C in this study). 135 The out wall of the HT reactor would be supplied with thermal insulation material if it were implemented in practice; however, heat loss through the reactor wall during the HT process was ignored in this study. 138 RESULTS AND DISCUSSION Soluble products from HT treatment of rice straw 141 Changes of pH value and extraction yield: HT treatment is an effective approach for the solubilization of biomass because the breakdown of macromolecular components is temperature-dependent. The HT extraction yield and pH value of the liquid fraction from HT 144 treatment are shown in Fig. 1. The extraction yield reflects the amount of all dissolved components, including dissolved hemicellulose, cellulose, lignin, protein and other soluble compounds. The extraction yield varied from 31% (HT180-10) to 44% (HT210-0), and it was 147 slightly declined to 39% under HTp210-0(5). As can be observed from the findings, increasing the peak temperature helped dissolve rice straw. The maximum extraction yield was achieved at 210 °C. Under this HT temperature, an additional experiment was 150 conducted to check the influence of headspace pressure and it was adjusted by amount of straw. As the additional experiment`s result shows, the maximum HT treatment extraction yields were 44.3% in HTp210-0(15), 39% in HTp210-0(5), and 40% in HTp-210-0(9), 153 respectively. This observation agrees with Yu et al. [22] who found that the soluble yield was ~36% under 180 °C for 10 minute, which could be a little bit increased (~40%) at 200 °C. The pH values varied from 3.31 (HT200-5) to 4.31 (HT180-10). From Fig.1, the pH value 156 was decreased from 4.31 to 3.31 (HT200-5), then slightly increased to 3.55 at HT210, probably due to a higher temperature especially > 200°C can break down some organic acids [18]. Generally, when compared to the total extraction yield and pH value, a reverse 159 tendency was noticed: the increased extraction yield was accompanied by a decreased pH value, probably owing to the production of organic acids from the dissolved hemicellulose. Ac ce pt ed m an us cr ip t Mongolian Journal of Chemistry Page 7 of 15 162 When the reactor headspace was changed, the pH values were also detected to change under 210 °C, which were degreased to 3.84, and 3.56 under HTp210-0(5) and HTp210-165 0(15), respectively. The results show that HTp210-0(5) and HTp210-0(9) conditions cannot substantially break down hemicellulose to organic acids when compared to HT210-0(15). This means that the reactor headspace or pressure may influence the extraction yield and 168 the liquid pH value. Dissolved carbohydrate and TOC from rice straw by HT treatment: The total dissolved carbohydrate was determined using the phenol-sulfuric acid method. This method can 171 reflect all types of sugars such as xylose, glucose and others. Fig. 2 shows the production of total sugars in the liquid fraction from HT treatment, including monomeric and oligomeric sugars. The total carbohydrate concentration varied depending on the HT temperature and 174 holding time. The lowest value was 8.3% from HT180-10, which was increased up to 17.1% under HT210-0. The total carbohydrate in rice straw is mostly made up of easily soluble polysaccharides (hemicellulose and mono sugar), rather than crystalline cellulose that is 177 usually degraded at temperatures above 230 °C [19]. It means under HT210, the obtained dissolved total carbohydrate was mainly from the dissolved hemicellulose. Total TOC concentration in the liquid part of HT treated rice straw followed a similar pattern to total 180 carbohydrate concentration. The lowest value was 4927 mg/L from HT180-10, and the highest value was 7849 mg/L obtained under HT210-0. 31 36.3 42.9 43.9 43.6 44 39 40 44.3 4.31 3.93 3.85 3.31 3.35 3.55 3.84 3.85 3.56 3.0 4.0 5.0 6.0 7.0 10 20 30 40 50 p H v a lu e E x tr a c ti o n y ie ld ( % , w /w ) HT conditions Extract yields, % Extract pH value Fig. 1. Extraction yield and pH value of HT liquid fraction Ac ce pt ed m an us cr ip t Mongolian Journal of Chemistry Page 8 of 15 183 Based on the above results, the most suitable condition was determined as HT210-0, under which the highest carbohydrate yield and TOC were achieved. For the headspace experiment, the amount of total carbohydrate and TOC were lower in HTp-210-0(5) and 186 HTp-210-0(9), when compared to HTp-210-0(15). More specifically, the total carbohydrates were 12.9% and 15.1% with TOC being 6060 mg/L and 6156 mg/L when the HT treatment was conducted under HTp-210-0(5) and HTp-210-0(9), respectively. In contrast, the total 189 carbohydrate was 17.7% with TOC being 7024 mg/l under HTp-210-0(15). This observation also suggests that the reactor headspace effected on the extraction yield and liquid products. 192 Dissolved organic acids in liquid fraction from HT treatments: Under the hydrothermal condition, xylose molecules can be broken down with organic acid production, which can influence the liquid pH value [18]. The VFAs obtained by HT treatment of rice straw at the 195 appropriate peak temperatures are shown in Fig. 3. The total concentration of VFAs was increased with the increase in HT temperature, while it was slightly degreased when prolonging the holding time. A longer holding time might break down some organic acids. 198 The dominant acid was acetic acid from all test conditions, accounting for 43% or 333 mg/L in HT180-10 to 96% or 847 mg/L in HTp210-0 of the total VFAs in the HT liquid fraction. HT210-0 produced the most successful solubilization of rice straw of all the test conditions, 201 mainly to the degradation of hemicellulose into xylose, which was then degraded into acetic acid. 4927 5612 7127 6999 6787 7849 6060 6156 7024 8.3 12.4 15.9 16.1 16.8 17.1 12.9 15.1 17.7 6 11 16 21 26 0 2000 4000 6000 8000 10000 T o ta l c a rb o h y d ra te ( % , w /w ) T O C ( m g /L , v /w ) HT conditions TOC Total carbohydrate Fig. 2. Soluble carbohydrate production and dissolved total organic carbons (TOC) from HT liquid fraction Ac ce pt ed m an us cr ip t Mongolian Journal of Chemistry Page 9 of 15 Other VFA species were also detected. In the HT treatment liquid from HT180-10, there 204 were 93 mg/L propionic acid, 95 mg/L n-butyric acid, 70 mg/L iso butyric acid, 83 mg/l iso valeric acid and 92 mg/L valeric acid. The concentrations of these VFAs were detected to decrease when the HT temperature increased over 200 °C. Under HT200-5 condition, only 207 two VFAs were detected, i.e., acetic acid (829 mg/L) and n-butyric acid (37 mg/L), suggesting that a higher temperature is beneficial for VFAs decomposition. In addition, a lower VFAs was detected in the liquid from HTp210-0(5) and HTp210-0(9) in 210 comparison to HTp210-0(15). There were 3 noticeable unknown peaks from the gas chromatography results that need further confirmation by additional VFA standards. These unknown VFA products were observed to increase when HT treatment was conducted at 213 temperature over 200°C. They could be levulinic acid and formic acid, which are produced at high temperatures from furfural and 5-HMF. This observation agrees with the statement by Liu et al. [20, 18] who detected the increase of these acids in the HT liquid part when 216 temperature was increased to 200 °C. The effect of HT treatment on the solubilization of rice straw was studied. The HT treatment yielded various amounts of carbohydrate and other products from rice straw. HT210 was 219 found to have a considerable impact on rice straw solubilization, boosting dissolved carbohydrate production with lower pH while also increasing VFA production. This observation suggests that this HT temperature is more suitable for hemicellulose 222 decomposition from rice straw. The reactor headspace experiment found that a smaller Fig. 3. Changes in individual volatile fatty acids (VFAs) during hydrothermal treatment of rice straw at peak temperatures 0 200 400 600 800 1000 V F A ( m g /L ) HT conditions n valeric acid iso valeric acid n butyric acid iso butyric acid propionic acid acetic acid Ac ce pt ed m an us cr ip t Mongolian Journal of Chemistry Page 10 of 15 headspace (HTp210-0(15)) is more effective compared to HTp210-0(5) and HTp210-0(9), yielding higher extraction rate, total carbohydrate and TOC concentrations. 225 Solid residue fraction from rice straw by HT pretreatment Lignin and total carbohydrates: The solid fraction from rice straw by HT treatment was dried at 105 °C for 24 hours after being separated by centrifuge. This dry residue is important as 228 it becomes a cellulose-rich biomass that could be used to produce ethanol and other useful products after hydrolysis. Fig. 4 shows the contents of lignin and total carbohydrate in the HT treated dry biomass. The lignin content was detected as 27.8 to 48.0 % (w/w) in rice 231 straw and HT210-0, respectively. The breakdown of hemicellulose resulted in an increase in lignin content as the HT temperature goes up. Results show that HT210 condition can remove most of the hemicellulose. 234 Fig. 4. Changes in lignin and total carbohydrate contents in the HT treated dry biomass The total carbohydrates varied from 29.3 to 50.3%. The highest carbohydrate content was 237 detected in the raw rice straw that contains all types of sugars such as hemicellulose, cellulose and soluble sugars. The lowest total carbohydrate content was 29.3 % in the treated dry biomass after 210 °C for holding 0 minute. This observation may indicate that 240 much hemicellulose has been removed and the remained carbohydrate might be only glucose. Due to a lack of suitable conditions for the HPLC, the contents of xylose, glucose, galactose, and arabinose were not measured in this study. 243 The reactor headspace experiments also have some difference on lignin and total carbohydrate contents. The highest lignin content was 47.1% in HTp210-0(15) condition compared to other 2 conditions, indicating that condition is more efficient to decompose 246 hemicellulose. However, the carbohydrates of HTp210-0(5), HTp210-0(9) were higher than HTp210-0(15) and they were 32.3%, 34.9%, respectively. It might be due to some amount Ac ce pt ed m an us cr ip t Mongolian Journal of Chemistry Page 11 of 15 of hemicellulose residue without breakdown. From the headspace experiments, amount of 249 loaded sample including water should be greater than 80% of the reactor capacity. Under this condition, a higher pressure would be created to break down the lignocellulosic materials compared to the lower filled (like 50 - 70%) reactor. 252 Morphological changes of treated biomass: The morphological changes were observed by optic microscopy. The rice straw without treatment looks so smooth and without any significant damage. During the HT treatment process, the surface of the treated rice straw 255 particles became more open, considerably rougher, and displayed clearly porous structures, likely resulting in much more contact between water molecules and carbohydrates inside the straw particles. The most suitable conditions were HT210-0 and HT200-10 because the 258 smooth structure looked broken down. FT-IR analysis: Under various HT treatment conditions, the FT-IR spectrum were recorded to investigate chemical structural changes in rice straw. As illustrated in Fig. 5, the changes 261 in functional groups in the treated rice straw were particularly noticeable between the wavenumbers 600 cm-1 and 1800 cm-1. The signal at 1720 cm-1, corresponding to the C=O functional group, is a typical peak of ester connected acetyl, feruloyl, and ρ-coumaroyl 264 groups between hemicellulose and lignin. The disappearance of this peak above 200 °C shows that HT treatment may have eliminated hemicellulose by cleaving the lignin- hemicellulose ester link. The signal observed at approximately 1432 cm−1 corresponds to -267 CH2 bending of cellulose [21]. The lignin-hemicellulose bond`s peaks at 1320 cm-1 (C-O of syringyl ring) and 1245 cm-1 (C-O of guaiacyl ring) were diminished in HT210-0. The peak at 1245 cm-1 (assigned to β-ether bonds in lignin and between lignin and carbohydrates [20] 270 was declined in the FT-IR spectrum of treated rice straw above 200 °C. These observations support the chemical components and optical microscopy findings. 273 Fig. 5. FT-IR spectrum of original and treated rice straw Ac ce pt ed m an us cr ip t Mongolian Journal of Chemistry Page 12 of 15 Energy consumption by HT treatment: The energy consumption by HT treatment is one of the important factors that influence the energy efficiency of the whole system. A higher HT 276 temperature can easily break down the rice straw complex structure, while it may be economically infeasible when compared to lower temperature HT treatment in terms of energy efficiency. Energy input was calculated based on per ton of extraction yield, meaning 279 how much energy was required for per ton of extract from HT treatment of rice straw. The energy input was calculated from the results obtained under HTp210-0(5), HTp210-0(9), and HTp210-0(15) in order to well understand the effect of the HT reactor headspace. Table 282 1 summarizes the required energy for per ton extract when HT treatment was conducted under the above conditions. 285 Table 1. The energy input of HT treatment for the headspace experiment HT conditions Water used (ton) Rice straw used (ton) HT extraction yield (%) HT yield (ton) Energy input (MJ) for HT extraction Energy input for per ton extract (MJ) HTp210-0(5) 5 0.5 39.0 0.195 4021 20620 HTp210-0(9) 9 0.9 40.0 0.360 7238 20104 HTp210-(15) 15 1.5 44.3 0.665 12063 18153 These data did not include the energy needed for drying after HT treatment. The energy 288 consumption was calculated as 20,620 MJ, 20,104 MJ and 18,153 MJ by HTp-210-0(5), HTp-210-0(9) and HTp-210-0(15), respectively. The lowest energy was consumed under HTp210-0(15) condition due to its higher extraction yield than the other two HT210 291 conditions. This result also suggests that the HT reactor headspace is critically essential for the enhanced breakdown of rice straw when energy consumption is taken into consideration. However, a more detailed energy balance analysis is necessary when the final products 294 such as ethanol, methane and other useful products are considered, which might be different when different final products being concerned. 297 CONCLUSIONS In this study, we investigated the effects of HT treatment on rice straw solubilization and residue changes. In terms of achieving optimal results, the HT treatment conducted at 210 300 °C for 0 minutes yielded the best outcome, with a soluble carbohydrate yield of 44% and a total organic carbon (TOC) content of 17.1%. The temperature of HT treatment was found to exert a significant influence on the production of volatile fatty acids (VFAs), with acetic 303 Ac ce pt ed m an us cr ip t Mongolian Journal of Chemistry Page 13 of 15 acid being the predominant species in this condition. Moreover, this study showed that HT treatment demonstrated higher efficiency at temperatures above 200 °C and short holding times, which was supported by evidence from FT-IR spectra and morphological changes. 306 Furthermore, we observed that reducing the headspace in the reactor resulted in a more efficient recovery of carbohydrates from rice straw with lowest energy usage. 309 ACKNOWLEDGMENTS This research was supported by a JDS grant from the Japanese government's JICA program at the University of Tsukuba. 312 REFERENCES 1. International Energy Outlook 2020 (IEO2020) Center for Strategic and International Studies. October 14, 2020. https://www.eia.gov/outlooks/ieo/pdf/ieo2020.pdf. (accessed on July 22, 2021) 2. 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