Substrate For Ethanol Fermentation Research Articles

Production of chitosan-based composite film reinforced with lignin-rich lignocellulose nanofibers from rice husk

Lignocellulosic nanofibers (LCNFs), implying lignin-containing cellulose fibers, maintain the properties of both lignin and cellulose, which are hydrophobic and hydrophilic, respectively. The presence of hydrophobic lignin in LCNFs is expected to be an economical and attractive option that can improve the thermal and mechanical properties of polymers. Thus, this study was conducted to produce lignin-rich LCNFs from sugar-rich waste obtained from rice husks after acidic pretreatment. The LCNFs were produced from the lignin-rich solid fractions obtained after pretreatment and enzymatic hydrolysis, which were then incorporated as an additive into a chitosan-based film. The variations in lignin content in the range of approximately 50.6%–66.8% in differently obtained LCNFs gave significantly different optical strengths and mechanical properties. These controllable processes may allow for customized film formation. Additionally, the glucose-rich liquid fractions obtained after pretreatment and enzymatic hydrolysis were used as a substrate for ethanol fermentation to achieve total utilization of rice husk biomass waste. In conclusion, the lignin-rich biomass fraction holds promise as a suitable material for chitosan-LCNF film and has the potential to increase the economic feasibility of the biomaterial industry.

Utilization of Fruit Waste for Bioethanol Production by Co-cultures of Aspergillus niger and Saccharomyces cerevisiae

The purpose of this research was to study the effectiveness of simultaneous fermentation offruit waste with co-cultures of Aspergillus niger TISTR 3063 and Saccharomyces cerevisiaeTISTR 5606 in production of ethanol. The effect of fermentation temperature on ethanol yield was also observed. Pomelo and banana peels were selected as substrates and prepared by chopping into small rectangular pieces. Fermentation of batches of fruit waste was carriedout using a 250mL Erlenmeyer flask with glucose as a control. Analysis of the composition of the fruit waste included sugar, pH, TS, VS, ash, moisture, COD and TKN. From the results, it was found that maximum yields of 90.71% and 104.90% for pomelo and banana peel, respectively, were achieved at a temperature of 40°C within 24 h. The analysis also showed that fermentation temperature affected ethanol yield. When the fermentation temperature was raised from 30°C to 40°C, maximum ethanol yield from pomelo peel fermentation with10% of inoculum was increased from 73.86% to 90.71%, significant at p-0.05 Maximum yield from banana peel fermentation showed a similar trend. This study establishes the potential for upgrading fruit wastes such as pomelo and banana peels as high value substrates for ethanol production. Pomelo peel in particular shows high potential as a substrate for ethanol fermentation at 40°C for 24 h, with inoculum of 10% (w/w) of each fungus and yeast.

Fermentative Production of Ethanol from Acid Hydrolyzate of Rice Water Waste Using Saccharomyces cerevisiae: Experimental and Kinetic Studies

Rice water waste, a starch-bearing effluent waste stream, was utilized as an abundant and low-cost substrate for ethanol production. Prior to the fermentation process, rice water waste was hydrolyzed using dilute H2SO4 solution. To attain the highest concentration of sugars with a minimum amount of inhibitory by-products during the acid hydrolysis, different acid concentrations (2–6 v/v %), hydrolysis temperatures (60–100 °C) and reaction times (0–200 min) were examined. The maximum catalytic efficiency (ηE = 27.36) was obtained using 4% H2SO4 at 80 °C for 100 min; wherein, the concentration of reducing sugars, furfural and 5- hydroxymethylfurfural (HMF) in the hydrolyzate was 43.32, 0.29 and 0.17 g L−1, respectively. This hydrolyzate was utilized as a substrate for ethanol fermentation using Saccharomyces cerevisiae; the maximum ethanol concentration, yield, and cell density were 18.65 g L−1, 0.41 g ethanol g−1 sugar and 3.68 g L−1, respectively. The kinetics of the acid hydrolysis process was studied using “two-fraction” model. The kinetic constants indicated that the rate of release of reducing sugars from starch was high; while, the degradation rate of reducing sugar to furfural and HMF was negligible. Moreover, multi-response nonlinear regression analysis was performed on the experimental data to determine the kinetic parameters regarding cell growth, substrate utilization, and ethanol production.

A combination of evaporation and chemical preservation for long‐term storage of fresh sweet sorghum juice and subsequent bioethanol production

Evaporation and chemical preservation were combined for long‐term storage of sweet sorghum juice (SSJ). The preserved SSJ was further used as substrate for ethanol fermentation. Different chemicals including hydrogen peroxide, sodium hypochlorite, sodium benzoate, and methyl paraben were used as co‐preservatives and were added into concentrated SSJ that contains ~40% (w/v) of sugars. The effects of chemical preservatives on sugar storage and ethanol fermentation were compared. The results showed that methyl paraben has better sugar preservation ability and an acceptable yeast toxicity when compared with other chemicals. The addition of methyl paraben in concentrated SSJ was further evaluated and optimized. By adding 0.4 g/L of methyl paraben, 95.1% of fermentable sugars in SSJ were preserved after a 90‐day preservation at room temperature. Ethanol concentration of 121.1 ± 4.6 g/L with a yield of 0.45 ± 0.02 g/g was obtained using the preserved SSJ in batch fermentation. PRACTICAL APPLICATIONS: Ethanol production by sugar‐containing juices is of interest because it is more effective, cheaper, and easier than the lignocellulosic routes. However, the commercialization of the bioethanol plants that uses the sweet sorghum juice as a substrate has long suffered from the deterioration of sugars. As a consequence, the ethanol fermentation process is limited by the short intermittent substrate supplementation in order to satisfy the harvest of the fresh stalks. This work comes up with a novel “evaporation+chemical” preservation method. The synergistic effect of high concentration of sugars and methyl paraben preservative showed inspiring sugar preservation results. Most of the sugars in the sweet sorghum juice were preserved for at least 90 days at room temperature and can be used for ethanol fermentation effectively. The continuity of fermentation can be significantly improved, which showed promising results in year‐round operations.

Exogenous L-proline Increases Stress Tolerance of Yeast Pichia kudriavzevii Against Inhibitors in Lignocellulose Hydrolysates and Enhances its Ethanol Production

Three ethanologenic yeast isolates (E, R and T) capable to use both pentose and hexose sugars as substrate for ethanol fermentation was isolated. Thus, suggesting its potential for 2nd generation of ethanol production by using lignocellulose hydrolysate. Based on ITS sequence, those three isolates were identified as yeast Pichia kudriavzevii. Prior its utilization as fermentation substrate, lignocellulose is subjected to complex hidrolysis reactions, which results certain toxic compounds (furfural and organic acid). Our results indicate that P. kudriavzevii isolates were sensitive toward 0.15% furfural and 0.375% acetic acid. Interestingly, supplementation of 1.5% L-proline in the medium significantly enhanced the survival rate of all P. kudriavzevii isolates against the particular toxic compounds. Moreover, supplementation of L-proline also enhanced the production of ethanol in glucose as substrate, in a dose dependent manner. Indeed, addition of 1.5% L-proline could increase the ethanol yield of isolate E, R and T for approximately 57%, 53%, and 62%, respectively, as compared to that 0.5% L-proline. The highest ethanol production was exhibited by isolate T (14g/100mL) in 1.5% L-proline treatment. Our data indicate, that L-proline supplementation may potentially be applied to enhance ethanol production by P. kudriavzevii isolates, particularly for its future application in lignocellulose-based ethanol productions.

Effect of acid pretreatment on different parts of corn stalk for second generation ethanol production

In this study, the effects of different parts of corn stalk, including stem, leaf, flower, cob and husk on second generation ethanol production were evaluated. FTIR, XRD and SEM were performed to investigate the effect of dilute acid pretreatment. The bagasse obtained after pretreatment were further hydrolyzed by cellulase and used as the substrate for ethanol fermentation. As results, hemicelluloses fractions in different parts of corn stalk were dramatically removed and the solid fractions showed vivid compositions and crystallinities. Compared with other parts of corn stalk, the cob had higher sugar content and better enzymatic digestibility. The highest glucose yield of 94.2% and ethanol production of 24.0gL−1 were achieved when the cob was used as feedstock, while the glucose yield and the ethanol production were only 86.0% and 17.1gL−1 in the case of flower.

Sorghum as a renewable feedstock for production of fuels and industrial chemicals

Considerable efforts have been made in the USA and other countries to develop renewable feedstocks for production of fuels and chemicals. Among these, sorghum has attracted strong interest because of its many good characteristics such as rapid growth and high sugar accumulation, high biomass production potential, excellent nitrogen usage efficiency, wide adaptability, drought resistance, and water lodging tolerance and salinity resistance. The ability to withstand severe drought conditions and its high water usage efficiency make sorghum a good renewable feedstock suitable for cultivation in arid regions, such as the southern US and many areas in Africa and Asia. Sorghum varieties include grain sorghum, sweet sorghum, and biomass sorghum. Grain sorghum, having starch content equivalent to corn, has been considered as a feedstock for ethanol production. Its tannin content, however, may cause problems during enzyme hydrolysis. Sweet sorghum juice contains sucrose, glucose and fructose, which are readily fermentable by Saccharomyces cerevisiae and hence is a good substrate for ethanol fermentation. The enzyme invertase, however, needs to be added to convert sucrose to glucose and fructose if the juice is used for production of industrial chemicals in fermentation processes that employ microorganisms incapable of metabolizing sucrose. Biomass sorghum requires pretreatment prior to enzymatic hydrolysis to generate fermentable sugars to be used in the subsequent fermentation process. This report reviews the current knowledge on bioconversion of sorghum to fuels and chemicals and identifies areas that deserve further studies.

빛의 조사 기간에 따른 세 가지 미세조류의 지질 함량 변화와 균체의 당화 전환율 비교

본 연구에서는 빛의 조사기간을 변화시켜 세 종류의 미세조류(Nanochloris, Dunaliella tertiolecta, Tetraselmis)를 배양하고 이들의 성장속도 및 지질 함량을 분석하였다. 빛의 조사기간은 한국의 여름철과 겨울철의 일조시간을 반영해 각각 14.5시간과 7시간으로 설정하였다. 또한, 지질 추출 후 남은 미세조류를 당화시켜 포도당 전환율을 비교함으로써 미세조류의 바이오매스로써의 가능성을 가늠하고자 하였다. 실험 결과 D. tertiolecta가 다른 두 종의 미세조류보다 빛의 조사기간이 7시간일 때 최대 38% 높은 성장속도를 나타냈으며 지질함량은 최대 43.6% 정도 높은 결과를 보였다. 포도당으로의 당화 전환율도 D. tertiolecta가 최대 22% 높은 결과를 보였다. Microalgae have the advantages of being able to utilize the solar energy and culturing at a low cost. In particular, microalgae have a great potential in the production of biodiesel due to the high lipid content. Lipids produced from microalgae are converted to fatty acid methyl ester (FAME) by trans-esterification reaction and FAME is called a biodiesel in general. In addition, microalgae can also be utilized as a substrate for ethanol fermentation after saccharification reaction. In this study, three types of microalgae (Nanochloris, Dunaliella tertiolecta, Tetraselmis) were cultured and their lipid contents were compared. In addition, the effects of lighting period on the growth rate and lipid content were studied. Finally, the amounts of glucose produced from each saccharified microalgae were investigated. As a result, we demonstrated that D. tertiolecta has 43.6% higher lipid content and 22% higher glucose conversion than two others.

Enzymatic and Acid Hydrolysis of Sago Starch for Preparation of Ethanol Production

A series of studies on the hydrolysis of Sago starch for ethanol fermentation had been conducted. Hydrolysis of sago starch was carried out using sulfuric acid 2.5% and amylase(s) enzymes. The concentrations of sago starch used in this experiment were 5, 10, 15, 20, and 30% (w/v). The highest hydrolyzate containing reducing sugar was used as substrate for ethanol fermentation by Saccharomyces cerevisiae FNCC 3012. The results indicated that hydrolysis using 2.5% sulfuric acid for 120 min at 121 °C produced 6.6% (w/v) reducing sugar and hydrolysis using α-amylase and Dextrozyme DX produced more reducing sugar, 7% (w/v) and 17.1% (w/v), respectively. The fermentation of hydrolyzed sago starch by S. cerevisiae FNCC 3012 produced ethanol 7.98% (v/v).

The influence of nitrate on the physiology of the yeast Dekkera bruxellensis grown under oxygen limitation

A previous study showed that the use of nitrate by Dekkera bruxellensis might be an advantageous trait when ammonium is limited in sugarcane substrate for ethanol fermentation. The aim of the present work was to evaluate the influence of nitrate on the yeast physiology during cell growth in different carbon sources under oxygen limitation. If nitrate was the sole source of nitrogen, D. bruxellensis cells presented slower growth, diminished sugar consumption and growth-associated ethanol production, when compared to ammonium. These results were corroborated by the increased expression of genes involved in the pentose phosphate (PP) pathway, the tricarboxylic acid (TCA) cycle and ATP synthesis. The presence of ammonium in the mixed medium restored most parameters to the standard conditions. This work may open up a line of investigation to establish the connection between nitrate assimilation and energetic metabolism in D. bruxellensis and their influence on its fermentative capacity in oxygen-limited or oxygen-depleted conditions.

Evaluation of the waste from cassava starch production as a substrate for ethanol fermentation by Saccharomyces cerevisiae

Cassava starch production waste (cassava pulp) has been proposed as a high potential ethanolic fermentation substrate due to its high residual starch level and the small particle size of the lignocellulosic fibers. Saccharification of the residual starch from a 3% (w/v) dry weight basis (DS) of cassava pulp by α-amylase (100°C, 10 min) and glucoamylase (60°C, 2 h) resulted in a glucose yield of 22.6 g/l [67.8% (w/w) DS of cassava pulp] and in lignocellulosic fibers at 0.5 g/g DS cassava pulp. Pretreatment of the lignocellulosic fiber with dilute sulfuric acid and calcium hydroxide at 121°C, 15 lb/in2 for 30 min increased and decreased, respectively, its susceptibility to cellulase hydrolysis. Under the optimal conditions found, pretreatment of 6% (w/v) DS lignocellulosic fiber by 2% (w/v) H2SO4 for 30 min, followed by saccharification by cellulase (40°C, 9 h), yielded a glucose level of 26.6 g/l [79.8% (w/w) DS of the cassava pulp]. The starch and lignocellulosic fiber hydrolysates obtained from 30 g cassava pulp and 60 g H2SO4 pretreated lignocellulosic fiber were fermented by Saccharomyces cerevisiae, without the need for (NH4)2SO4 supplementation, to yield ethanol levels of 9.9 and 11.9 g/l, respectively, after 48 h.

Evaluation of wet air oxidation as a pretreatment strategy for bioethanol production from rice husk and process optimization

The pretreatment of rice husk by the wet air oxidation (WAO) technique was investigated by means of a statistically designed set of experiments. Reaction temperature, air pressure, and reaction time were the process parameters considered. WAO pretreatment of rice husk increased the cellulose content of the solid fraction by virtue of lignin removal and hemicellulose solubilization. The cellulose recovery was around 92%, while lignin recovery was in the tune of 8–20%, indicating oxidation of a bulk quantity of lignin. The liquid fraction was found to be rich in hexose and pentose sugars, which could be directly utilized as substrate for ethanol fermentation. The WAO process was optimized by multi-objective numerical optimization with the help of MINITAB 14 suite of statistical software, and an optimum WAO condition of 185 °C, 0.5 MPa, and 15 min was predicted and experimentally validated to give 67% (w/w) cellulose content in the solid fraction, along with 89% lignin removal, and 70% hemicellulose solubilization; 13.1 gl−1 glucose and 3.4 gl−1 xylose were detected in the liquid fraction. The high cellulose content and negligible residual lignin in the solid fraction would greatly facilitate subsequent enzymatic hydrolysis, and result in improved ethanol yields from rice husk.

Ethanol Production Potential from Oil Palm Empty Fruit Bunches in Southeast Asian Countries Considering Xylose Utilization

Much attention has been paid to fuel ethanol recently, but the use of food resources for fuel ethanol is problematic and the use of non-food resources is anticipated. We are developing ethanol production technology from lignocellulosic biomass subject to milling pretreatment and enzymatic hydrolysis, and also developed new yeast strains which can ferment xylose into ethanol efficiently. We estimated ethanol production potential from oil palm empty fruit bunches (EFB) in Indonesia, Malaysia, and Thailand with our experimental data. Even estimation based on moderate resource availability and without xylose utilization showed 1-2 % of gasoline consumption can be substituted by ethanol produced from EFB in Indonesia and Malaysia. If xylose can be also used as a substrate for ethanol fermentation, its contribution to production increase was significant. The use of other wastes generated in oil palm industry can increase ethanol production furthermore. The ethanol production potential from wasted oil palm trunk in Malaysia is comparable to that from EFB by our estimation which also based on the experimental data. Palm-oil-producing Southeast Asian countries have large ethanol production potential without utilizing food resources.

Investigation of the utility of pineapple juice and pineapple waste material as low-cost substrate for ethanol fermentation by Zymomonas mobilis

The utility of the juice of rotten or discarded pineapples and the waste material of the production of pineapple juice as low-cost substrates for ethanol production by Zymomonas mobilis was investigated. Z. mobilis ATCC 10988 produced 59.0g·l −1 ethanol in undiluted pineapple juice without nutritional supplementation and without the regulation of the pH while 42.5g·l −1 ethanol was obtained using a125g·l −1 sucrose medium supplemented with 10g·l −1 yeast extract and mineral salts. Ethanol fermentation using unhydrolyzed and enzymatically hydrolyzed pineapple waste material was also investigated under various culture conditions. When a 15% (v/v) dilution of unhydrolyzed waste material without nutritional supplementation was used, more than 3.5g·l −1 ethanol was produced. When the media containing 15, 30, and 40% (v/v) of the hydrolyzate consisting of a 60% (v/v) suspension of pineapple waste material were used, final concentrations of ethanol were 5.0g·l −1, 7.6g·l −1, and 9.3g·l −1, respectively. These results suggest that pineapple juice and the waste material can be useful low-cost substrates for ethanol production by Z. mobilis without supplementation with expensive organic nitrogen complexes such as yeast extract and without the regulation of the pH during cultivation, leading to the reduction in the production costs.

Acid hydrolysis of sweet potato for ethanol production

Studies were conducted to establish optimal conditions for the acid hydrolysis of sweet potato for maximal ethanol yield. The starch contents of two sweet potato cultivars (Georgia Red and TG-4), based on fresh weight, were 21.1 +/- 0.6% and 27.5 +/- 1.6%, respectively. The results of acid hydrolysis experiments showed the following: (1) both hydrolysis rate and hydroxymethylfurfural (HMF) concentration were a function of HCL concentration, temperature, and time; (2) the reducing sugars were rapidly formed with elevated concentrations of HCl and temperature, but also destroyed quickly; and (3) HMF concentration increased significantly with the concentration of HCl, temperature, and hydrolysis time. Maximum reducing sugar value of 84.2 DE and 0.056% HMF (based on wet weight) was achieved after heating 8% SPS for 15 min in 1N HCl at 110 degrees C. Degraded 8% SPS (1N HCl, 97 degrees C for 20 min or 110 degrees C for 10 min) was utilized as substrate for ethanol fermentation and 3.8% ethanol (v/v) was produced from 1400 mL fermented wort. This is equal to 41.6 g ethanol (200 proof) from 400 g of fresh sweet potato tuber (Georgia Red) or an ethanol yield potential of 431 gal of 200-proof ethanol/acre (from 500 bushel tubers/acre).