Study of a single and mixed culture for the direct bio-conversion of sorghum carbohydrates to ethanol

Fusaium oxysporum F3 alone or in mixed culture with Saccharomyces cerevisiae 2541 fermented soluble and insoluble carbohydrates of sweet sorghum stalk directly to ethanol. Both microorganisms were first grown aerobically and fermented sorghum stalk to ethanol thereafter. During fermentation, insoluble carbohydrates were hydrolysed to soluble sugars by the celluloytic system of F. oxysporum. Ethanol yields as high as 24.4 and 33.5 g/100 g dry stalks were obtained by F. oxysporum and the mixed culture respectively, representing a theoretical yield enhancement of 11.6% and 53.6% respectively. The corresponding ethanol concentrations in the fermentation medium were 4.6% and 6.4% (w/v). These results clearly demonstrated that a large portion of insoluble carbohydrate from sorghum was converted by simultaneous saccharification and fermentation to ethanol, making the process promising for bioethanol production.     

A novel cost-effective technology to convert sucrose and homocelluloses in sweet sorghum stalks into ethanol

Background

Sweet sorghum is regarded as a very promising energy crop for ethanol production because it not only supplies grain and sugar, but also offers lignocellulosic resource. Cost-competitive ethanol production requires bioconversion of all carbohydrates in stalks including of both sucrose and lignocellulose hydrolyzed into fermentable sugars. However, it is still a main challenge to reduce ethanol production cost and improve feasibility of industrial application. An integration of the different operations within the whole process is a potential solution.

Results

An integrated process combined advanced solid-state fermentation technology (ASSF) and alkaline pretreatment was presented in this work. Soluble sugars in sweet sorghum stalks were firstly converted into ethanol by ASSF using crushed stalks directly. Then, the operation combining ethanol distillation and alkaline pretreatment was performed in one distillation-reactor simultaneously. The corresponding investigation indicated that the addition of alkali did not affect the ethanol recovery. The effect of three alkalis, NaOH, KOH and Ca(OH)₂ on pretreatment were investigated. The results indicated the delignification of lignocellulose by NaOH and KOH was more significant than that by Ca(OH)₂, and the highest removal of xylan was caused by NaOH. Moreover, an optimized alkali loading of 10% (w/w DM) NaOH was determined. Under this favorable pretreatment condition, enzymatic hydrolysis of sweet sorghum bagasse following pretreatment was investigated. 92.0% of glucan and 53.3% of xylan conversion were obtained at enzyme loading of 10 FPU/g glucan. The fermentation of hydrolyzed slurry was performed using an engineered stain, Zymomonas mobilis TSH-01. A mass balance of the overall process was calculated, and 91.9 kg was achieved from one tonne of fresh sweet sorghum stalk.

Conclusions

A low energy-consumption integrated technology for ethanol production from sweet sorghum stalks was presented in this work. Energy consumption for raw materials preparation and pretreatment were reduced or avoided in our process. Based on this technology, the recalcitrance of lignocellulose was destructed via a cost-efficient process and all sugars in sweet sorghum stalks lignocellulose were hydrolysed into fermentable sugars. Bioconversion of fermentable sugars released from sweet sorghum bagasse into different products except ethanol, such as butanol, biogas, and chemicals was feasible to operate under low energy-consumption conditions.

     

Impact of Storage Time on the Juice and Sugars Extracted from Chopped and Whole Stalk Sweet Pearl Millet and Sweet Sorghum Biomass

Since they have a high concentrations of fermentable sugars, sweet pearl millet and sweet sorghum are two interesting crops for bioethanol production. However, if the juice is not extracted from the biomass immediately after harvest, the biomass has to be transported and stored for further juice extraction. This delay could affect the amount of juice extracted and its sugar concentration. This paper presents the results of 3 years of experiments where different storage modes (chopped and whole stalks) and various storage time (0 to 14 days) were applied on two different crop species (sweet pearl millet and sweet sorghum). Storing sweet pearl millet as whole stalks for 2 weeks resulted in a water-soluble carbohydrate (WSC) concentration decrease of 52 %, while no significant decrease of the WSC concentration was observed for sweet sorghum. Whole stalks storage is much more efficient than storing the biomass chopped to avoid a rapid sugar loss. However, more juice can be extracted from stored chopped biomass than from stored whole stalks biomass. If the juice cannot be extracted quickly after the harvest, the biomass can be stored as whole stalks to avoid rapid sugar deterioration, especially for sweet sorghum.     

Simultaneous saccharification and fermentation of sweet sorghum carbohydrates to ethanol in a fed-batch process

Summary The simultaneous saccharification and fermentation (SSF) of sweet sorghum carbohydrates to ethanol by Fusarium oxysporum F3 alone or in mixed culture with Saccharomyces cerevisiae 2541 or Zymomonas mobilis CP4 in a fed-batch fermentation process was studied. While SSF was adequately carried out by the first microorganism the process achieved its maximum value by the mixed culture of the fungus and yeast. Under optimum conditions, ethanol yields and concentrations as high as 29.7 g of ethanol per 100 g of dry sorghum stalk and 7.5 % (w/v) respectively were obtained. These values together with the high yield of sorghum crop in Greece make this process promising and worthy of further investigation for the production of fuel bioethanol.     

先进固体发酵技术(ASSF)生产甜高粱乙醇

介绍了利用高产能源作物甜高粱生产燃料乙醇的先进固态发酵(ASSF)技术,从甜高粱茎秆保存、菌种、反应器,到固体发酵过程的数学模拟和工程放大进行了系统研究。筛选出高效产乙醇的菌种CGMCC1949,固体发酵时间低于30 h,乙醇收率高于92%;优选出贮存甜高粱茎秆的有效方法,通过抑菌处理,厌氧贮存200 d糖分损失小于5%;对固态发酵过程进行了数学模拟,设计并优化了固体发酵设备,成功进行了工程放大试验,并且基于ASPEN软件对该技术进行了技术经济评价,结果表明ASSF法生产甜高粱乙醇在技术、工程和经济上均具有充分的可行性和明显优势。     

Fermentation of Detoxified Acid-Hydrolyzed Pyrolytic Anhydrosugars into Bioethanol with <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> 2.399

Pyrolysate obtained from the pyrolysis of waste cotton is a source of fermentable sugars that could be fermented into bioethanol fuel and other chemicals via microbial fermentation. However, pyrolysate is a complex mixture of fermentable and non-fermentable substrates causing inhibition of the microbial growth. The aim of this study was to detoxify the hydrolysate and then ferment it into bio-ethanol fuel in shake flasks and fermenter applying yeast strain Saccharomyces cerevisiae 2.399. Pyrolysate was hydrolyzed to glucose with 0.2 M sulfuric acid, neutralized with Ba(OH)₂ followed by treatment with ethyl acetate and activated carbon to remove fermentation inhibitors. The effect of various fermentation parameters such as inoculum concentration, pH and hydrolysate glucose was evaluated in shake flasks for optimum ethanol fermentation. With respect to inoculum concentration, 20% v/v inoculum i.e. 8.0 × 10⁸–1.2 × 10⁹ cells/mL was the optimum level for producing 8.62 ± 0.33 g/L ethanol at 9 h of fermentation with a maximum yield of 0.46 g ethanol/g glucose. The optimum pH for hydrolysate glucose fermentation was found to be 6.0 that produced 8.57 ± 0.66 g/L ethanol. Maximum ethanol concentration, 14.78 g/L was obtained for 4% hydrolysate glucose concentration after 16 h of fermentation. Scale-up studies in stirred fermenter produced much higher productivity (1.32 g/L/h^–1) compared to shake flask fermentation (0.92 g/L/h^–1). The yield of ethanol reached a maximum of 91% and 89% of the theoretical yield of ethanol in shake flasks and fermenter, respectively. The complex of integrated models of development was applied, that has been successfully tested previously for the mathematical analysis of the fermentation processes.     

Evaluation and Modeling of Bioethanol Yield Efficiency from Sweet Sorghum Juice

One of the challenges with using sweet sorghum as an energy crop is that although fermentation of the juice to ethanol does not require enzymes, the juice can easily spoil. One strategy to avoid spoilage is to harvest the juice in the field, place it into a tanker for transport, and add the yeast immediately to initiate the fermentation process to begin during transport. Hence, it is also important to understand how the fermentation process is influenced by pH, temperature, and dissolved oxygen, since these parameters would not be “controlled” during transport. A full factorial design was applied to examine and optimize yield efficiency of ethanol production for the fermentation of sweet sorghum juice. Bioethanol yield efficiency was modeled using a linear equation. Under optimal pH (5.5), temperature (28 °C), and dissolved oxygen (0%) conditions, a maximum theoretical yield efficiency of 0.75 was achieved for bioethanol produced from M81E variety of sweet sorghum.     

Application of simultaneous saccharification and fermentation (SSF) from viscosity reducing of raw sweet potato for bioethanol production at laboratory, pilot and industrial scales

The aim of this work was to research a bioprocess for bioethanol production from raw sweet potato by Saccharomyces cerevisiae at laboratory, pilot and industrial scales. The fermentation mode, inoculum size and pressure from different gases were determined in laboratory. The maximum ethanol concentration, average ethanol productivity rate and yield of ethanol after fermentation in laboratory scale (128.51 g/L, 4.76 g/L/h and 91.4%) were satisfactory with small decrease at pilot scale (109.06 g/L, 4.89 g/L/h and 91.24%) and industrial scale (97.94 g/L, 4.19 g/L/h and 91.27%). When scaled up, the viscosity caused resistance to fermentation parameters, 1.56 AUG/g (sweet potato mash) of xylanase decreased the viscosity from approximately 30000 to 500 cp. Overall, sweet potato is a attractive feedstock for be bioethanol production from both the economic standpoints and environmentally friendly.     

Produced by a Strain of Unidentified Actinomycetes sp.

The possibility of using two kinds of sorghum as raw materials in consolidated bioprocessing bioethanol production using Flammulina velutipes was investigated. Enzymatic saccharification of sweet sorghum was not as high as in brown mid-rib (bmr) mutated sorghum, but the amount of ethanol production was higher. Ethanol production from bmr mutated sorghum significantly increased when saccharification enzymes were added to the culture.     

EVALUATION OF BIOETHANOL PRODUCTION FROM CAROB PODS BY Zymomonas mobilis AND Saccharomyces cerevisiae IN SOLID SUBMERGED FERMENTATION

Bioethanol production from carob pods has attracted many researchers due to its high sugar content. Both Zymomonas mobilis and Saccharomyces cerevisiae have been used previously for this purpose in submerged and solid-state fermentation. Since extraction of sugars from the carob pod particles is a costly process, solid-state and solid submerged fermentations, which do not require the sugar extraction step, may be economical processes for bioethanol production. The aim of this study is to evaluate the bioethanol production in solid submerged fermentation from carob pods. The maximum ethanol production of 0.42 g g^?1 initial sugar was obtained for Z. mobilis at 30°C, initial pH 5.3, and inoculum size of 5% v/v, 9 g carob powder per 50 mL of culture media, agitation rate 0 rpm, and fermentation time of 40 hr. The maximum ethanol production for S. cerevisiae was 0.40 g g^?1 initial sugar under the same condition. The results obtained in this research are comparable to those of Z. mobilis and S. cerevisiae performance in other culture mediums from various agricultural sources. Accordingly, solid submerged fermentation has a potential to be an economical process for bioethanol production from carob pods.     

L(+)-Lactic acid from sweet sorghum by submerged and solid-state fermentations

Sweet sorghum was used as the raw material for the lactate production by a strain of Lactobacillus paracasei. The submerged conversion of sugar juice obtained from sweet sorghum by extraction could be accomplished with the same efficiency as observed in a control experiment with MRS-glucose medium (final lactate concentration of 88–106 g/l, lactate yield of 91–95%, duration of the fermentation of 24–32 h). Finely ground stalks of sorghum served as the substrate in the solid-state fermentation. The lactate accumulation in the solid medium and the lactate yield were optimized up to values comparable with the results from the submerged fermentation (final lactate concentration of 90 g/kg, lactate yield of 91–95%). However, the duration of the fermentation amounted to 120–200 h in the solid-state process. The data from a series of experiments performed at variable values of temperatures between 30°C and 36°C and initial sugar concentrations between 60 g/kg and 115 g/kg, and degrees of moisture between 78% and 82% was the basis of a polynomial multidimensional regression. As a result, simple three-dimensional model functions were obtained for the maximum productivity of lactate formation, the lactate yield and the time required for a 90% conversion.     

Intermediate-Scale, Semicontinuous Solid-Phase Fermentation Process for Production of Fuel Ethanol from Sweet Sorghum

A novel, semicontinuous solid-phase fermentation system was used to produce fuel ethanol from sweet sorghum. The process was at an intermediate scale. In the process, dried and shredded sweet sorghum was rehydrated to 70% moisture, acidified to pH 2.0 to 3.0, and either pasteurized (12 h at 70 to 80°C) or not pasteurized before spray inoculation with a broth culture of Saccharomyces cerevisiae. Fermented pulp exited the semicontinuous fermentor after a retention time of 72 h and contained approximately 6% (vol/vol) ethanol. Ethanol yields from dry sweet sorghum were 176 to 179 liters/10³ kg (85% of theoretical). Production costs for a greatly scaled-up (×1,400) conceptual version of this system were projected by calculation to average $0.47/liter for 95% ethanol. The calculated energy balance (energy output/energy input ratio) was estimated to be 1.05 when pasteurization was included and 1.31 when pasteurization was omitted. In calculating the energy balances, the output energy of the protein feed byproduct and the input energy for growing the sweet sorghum were not considered. A design for the scaled-up plant (farm scale) is provided.     

Efficient Production of Lactic Acid from Sweet Sorghum Juice by a Newly Isolated Lactobacillus salivarius CGMCC 7.75

Sweet sorghum juice was a cheap and renewable resource, and also a potential carbon source for the fermentation production of lactic acid (LA) by a lactic acid bacterium. One newly isolated strain Lactobacillus salivarius CGMCC 7.75 showed the ability to produce the highest yield and optical purity of LA from sweet sorghum juice. Studies of feeding different concentrations of sweet sorghum juice and nitrogen source suggested the optimal concentrations of fermentation were 325 ml l⁻¹ and 20 g l⁻¹, respectively. This combination produced 142.49 g l⁻¹ LA with a productivity level of 0.90 g of LA per gram of sugars consumed. The results indicated the high LA concentration achieved using L. salivarius CGMCC 7.75 not only gives cheap industrial product, but also broaden the application of sweet sorghum.

Electronic supplementary material

The online version of this article (doi:10.1007/s12088-013-0377-0) contains supplementary material, which is available to authorized users.     

Impacts of main factors on bioethanol fermentation from stalk juice of sweet sorghum by immobilized Saccharomyces cerevisiae (CICC 1308)

In order to attain a higher ethanol yield and faster ethanol fermentation rate, orthogonal experiments of ethanol fermentation with immobilized yeast from stalk juice of sweet sorghum were carried out in the shaking flasks to investigate the effect of main factors, namely, fermentation temperature, agitation rate, particles stuffing rate and pH on ethanol yield and CO(2) weight loss rate. The range analysis and analysis of variance (ANOVA) were applied for the results of orthogonal experiments. Results showed that the optimal condition for bioethanol fermentation should be A(4)B(3)C(3)D(4), namely, fermentation temperature, agitation rate, particles stuffing rate and pH were 37 degrees C, 200rpm, 25% and 5.0, respectively. The verification experiments were carried out in shaking flasks and 5L bioreactor at the corresponding parameters. The results of verification experiments in the shaking flasks showed that ethanol yield and CO(2) weight loss rate were 98.07% and 1.020gh(-1), respectively. The results of ethanol fermentation in the 5L bioreactor showed that ethanol yield and fermentation time were 93.24% and 11h, respectively. As a result, it could be concluded that the determined optimal condition A(4)B(3)C(3)D(4) was suitable and reasonable for the ethanol fermentation by immobilized Saccharomyces cerevisiae. The conclusion in the research would be beneficial for application of ethanol fermentation by immobilized S. cerevisiae from stalk juice of sweet sorghum.     

High alcohol production by solid substrate fermentation from starchy substrates using thermotolerant Saccharomyces cerevisiae

Solid Substrate Fermentation system (SSF) was used to produce ethanol from various starchy substrates like sweet sorghum, sweet potato, wheat flour, rice starch, soluble starch and potato starch using thermotolerant yeast isolate (VS₃) by simultaneous saccharification and fermentation process. Alcohol produced was estimated by gas chromatography after an incubation time of 96 hrs at 37v°C and 42v°C. More ethanol was produced from rice starch and sweet sorghum. The maximum amount of ethanol produced from these substrates using VS₃ was 10 g/100 g and 3.5 g/100 g substrate (rice starch) and 8.2 g and 7.5 g/100 g substrate (sweet sorghum) at 37v°C and 42v°C respectively.     

Solid-state fermentation of sweet sorghum to ethanol

Solid-state fermentation of chopped sweet sorghum particles to ethanol was studied in static flasks using an ethanol tolerant yeast strain. The influence of various process parameters, such as temperature, yeast cell concentration, and moisture content, on the rate and extent of ethanol fermentation was investigated. Optimal values of these parameters were found to be 35 degrees C, 7 x 10(8) cells/g raw sorghum, and 70% moisture level, respectively.     

Comparison between solid-state and powder-state alkali pretreatment on saccharification and fermentation for bioethanol production from rice straw

The efficacy of different concentrations of NaOH (0.25%, 0.50%, 0.75%, and 1.00%) for the pretreatment of rice straw in solid and powder state in enzymatic saccharification and fermentation for the production of bioethanol was evaluated. A greater amount of biomass was recovered through solid-state pretreatment (3.74 g) from 5 g of rice straw. The highest increase in the volume of rice straw powder as a result of swelling was observed with 1.00% NaOH pretreatment (48.07%), which was statistically identical to 0.75% NaOH pretreatment (32.31%). The surface of rice straw was disrupted by the 0.75% NaOH and 1.00% NaOH pretreated samples as observed using field-emission scanning electron microscopy (FE-SEM) and atomic force microscopy (AFM). In Fourier-transform infrared (FT-IR) spectra, absorbance of hydroxyl groups at 1,050 cm^?1 due to the OH group of lignin was gradually decreased with the increase of NaOH concentration. The greatest amounts of glucose and ethanol were obtained in 1.00% NaOH solid-state pretreated and powder-state hydrolyzed samples (0.804 g g^?1 and 0.379 g g^?1, respectively), which was statistically similar to the use of 0.75% NaOH (0.763 g g^?1 and 0.358 g g^?1, respectively). Thus, solid-state pretreatment with 0.75% NaOH and powder-state hydrolysis appear to be suitable for fermentation and bioethanol production from rice straw.     

Changes of Saccharomyces cerevisiae cell membrane components and promotion to ethanol tolerance during the bioethanol fermentation

During bioethanol fermentation process, Saccharomyces cerevisiae cell membrane might provide main protection to tolerate accumulated ethanol, and S. cerevisiae cells might also remodel their membrane compositions or structure to try to adapt to or tolerate the ethanol stress. However, the exact changes and roles of S. cerevisiae cell membrane components during bioethanol fermentation still remains poorly understood. This study was performed to clarify changes and roles of S. cerevisiae cell membrane components during bioethanol fermentation. Both cell diameter and membrane integrity decreased as fermentation time lasting. Moreover, compared with cells at lag phase, cells at exponential and stationary phases had higher contents of ergosterol and oleic acid (C_18:1) but lower levels of hexadecanoic (C_16:0) and palmitelaidic (C_16:1) acids. Contents of most detected phospholipids presented an increase tendency during fermentation process. Increased contents of oleic acid and phospholipids containing unsaturated fatty acids might indicate enhanced cell membrane fluidity. Compared with cells at lag phase, cells at exponential and stationary phases had higher expressions of ACC1 and HFA1. However, OLE1 expression underwent an evident increase at exponential phase but a decrease at following stationary phase. These results indicated that during bioethanol fermentation process, yeast cells remodeled membrane and more changeable cell membrane contributed to acquiring higher ethanol tolerance of S. cerevisiae cells. These results highlighted our knowledge about relationship between the variation of cell membrane structure and compositions and ethanol tolerance, and would contribute to a better understanding of bioethanol fermentation process and construction of industrial ethanologenic strains with higher ethanol tolerance.     

Ethanol fermentation of mahula (Madhuca latifolia L.) flowers using free and immobilized yeast Saccharomyces cerevisiae

There is a growing interest to find alternate bioresources for production of ethanol, apart from cane/sugar beet molasses and starchy crops like sweet sorghum, cassava and sweet potato. Mahula (Madhuca latifolia L.) is a forest tree abundantly available in the Indian subcontinent and its flowers are very rich in fermentable sugars (28.1-36.3 g 100 g(-1)). Batch fermentation of fresh and 12-month-stored flowers with free (whole cells) and immobilized cells of Saccharomyces cerevisiae (strain CTCRI) was carried out in 2-l Erlenmeyer flasks. The ethanol yields were 193 and 148 g kg(-1) (using free cells) and 205 and 152 g kg(-1) (using immobilized cells) from fresh and 12-month-stored mahula flowers, respectively.     

Energy analysis of ethanol production from sweet sorghum

The Piedmont System is a collection of equipment for efficiently removing the juice from sweet sorghum stalks for the production of ethanol. The concept is to separate the whole stalks into pith and rind-leaf fractions, pass only the pith fraction through a screw press, and thus achieve an improvement in juice-expression efficiency and press capacity. An energy analysis was done for two options of this proposed harvesting/processing system: (Option 1) The juice is evaporated to syrup and used throughout the year to produce ethanol, and the by-products are used as cattle feed. (Option 2) The juice is fermented as it is harvested, and the by-products (along with other cellulosic materials) are used as feedstock for the remainder of the year. Energy ratios (energy output/energy input) of 0·9, 1·1 and 0·8 were found for sweet sorghum Option 1, sweet sorghum Option 2, and corn, respectively, as feedstocks for ethanol. If only liquid fuels are considered, the ratios are increased to 3·5, 7·9 and 4·5.