Bioethanol Production Although CO2 is the most important greenhouse gas (GHG), several studies show that it is important to consider other GHGs as well. The continued use of fossil fuels to meet the majority of the world’s energy demand energy demand is threatened by increasing concentrations of CO2 In the atmosphere and concerns over global warming. The combustion of fossil fuels is responsible for 73% of the CO2 production. production . The heightened awareness of the global warming issue has increased interest in the development of methods to mitigate GHG emissions. Much of the current effort to control such emissions focuses on advancing technologies that: 1) Reduce energy consumption. 2) Increase the efficiency of energy conversion or utilization. 3) switch to lower carbon content fuels, 4) enhance natural sinks for CO2, 5) Capture and store CO2. Reducing use of fossil fuels would considerably reduce the amount of CO2 produced, as well as reduce the levels of pollutants . To reduce the net contribution of GHGs to the atmosphere, bioethanol bioethanol has been recognized as a potential alternative to petroleumderived transportation fuels.
Overview Ethanol is made from a variety of products such as grain, molasses, fruit, cobs, and shell; its production, excluding that of beverages, has been declining since the 1930s because of the low cost. With the oil crises of the 1970s, ethanol became established as an alternative fuel. In 1975, only 76*106 lit of proof industrial ethanol was produced by fermentation compared to 7.95*106 lit by synthesis corn ethanol production causes more soil erosion than any other crop grown and uses more nitrogen fertilizer than any other crop grown. These two environmental limitations also apply to sugar cane production in Brazil properties
Properties of Bioethanol Bioethanol has a higher octane number, broader flammability limits, higher flame speeds and higher heats of vaporization than gasoline . These properties allow for a higher compression ratio, shorter burn time and leaner burn engine, which lead to theoretical efficiency advantages over gasoline in an internal combustion engine. Disadvantages Disadvantages of bioethanol include its lower energy density than gasoline (bioethanol has 66% of the energy that gasoline has), its corrosiveness, low flame luminosity, lower
vapor pressure (making cold starts difficult), miscibility with water, and toxicity to ecosystems Ethanol is an oxygenated fuel that contains 35% oxygen, which reduces particulate and NOx emissions from combustion. Ethanol has a higher octane number (108), broader flammability limits, higher flame speeds and higher heats of vaporization. These properties allow for a higher An oxygenate fuel such as bioethanol provides a reasonable antiknock value. Also, as it contains oxygen, fuel combustion is more efficient, reducing hydrocarbons and particulates in exhaust gases. The presence of oxygen in bioethanol improves combustion and therefore reduces hydrocarbon, carbon monoxide, and particulate emissions; but oxygenated fuels also tend to increase nitrogen oxide emissions.
Blending of Ethanol with Gasoline The most popular blend for light-duty vehicles is known as E85, and contains 85% bioethanol and 15% gasoline. In Brazil, bioethanol for fuel is derived from sugar cane and is used pure or blended with gasoline in a mixture called gasohol (24% bioethanol, 76% gasoline) In several states of the United States, a small amount of bioethanol (10% by volume) is added to gasoline, known as gasohol or E10. Blends having higher concentrations of bioethanol in gasoline are also used, e.g. in flexible-fuel vehicles that can operate on blends of up to 85% bioethanol—E85 [17]. Some countries have exercised biofuel program involving both form bioethanol–gasoline blend program, e.g. the United States (E10 and for Flexible Fuel Vehicle (FFV) E85), Canada (E10 and for FFV E85), Sweden (E5 and for FFV E85), India (E5), Australia (E10), Thailand (E10), China (E10), Columbia (E10), Peru (E10), Paraguay (E7), and Brazil (E20, E25 and FFV any blend) [18].
Current status In 2006, global production of bioethanol reached 13.5 billion gallons, up from 12.1 billion gallons in 2005. Bioethanol currently accounts for more than 94% of global biofuel production, with the majority coming from sugar cane. About 60% of global bioethanol production comes from sugar cane and 40% from other crops. Brazil and the United States are the world leaders, which exploit sugar cane and corn, respectively, and they together account for about 70% of the world bioethanol production. However, the US and Brazil are not oil independent countries. According to the Renewable Fuels Association (RFA) 2007 figures. In the United States, 90% of bioethanol is derived from Corn
Bioethanol feedstock
Bioethanol feedstock’s can be conveniently classified into three types: 1) Sucrose containing feedstock (e.g. sugar beet, sweet sorghum and sugar cane), 2) Starchy materials (e.g. wheat, corn, and barley), 3) lignocellulosic biomass (e.g. wood, straw, and grasses).
Two-third of world sugar production is from sugar cane and one-third is from sugar Beet Compared to sugar cane, sugar beet requires 35–40% less water and fertilizer. The conversion of carbohydrates with 5 and 6 carbons into bioethanol is easier compared to starchy materials and lignocellulosic biomass because previous hydrolysis of the feedstock is not required since this disaccharide can be broken down by the yeast cells; in addition, the conditioning of the cane juice or molasses favors the hydrolysis of sucrose
Reference Mustafa Balat, Havva Balat, Cahide Oz. progress in Bioethanol processing, progress in energy and combustion science 34 (2008) 551-578
Yeast Used for the production of Bioethanol
Zymomonas mobilis Ethanol production from sugar cane molasses was analyzed under different culture conditions using Z. mobilis in batch fermentation. The total reducing sugars (TRS) concentrations in the molasses, temperature, and agitation and culture time effects were studied simultaneously through factorial design. The best conditions for ethanol production were 200 g L-1 of total reducing sugars in the molasses, temperature of 30 C and static culture and time of fermentation of 48 h, achieving 55.8 g L-1. The pH of the medium was kept constant during the experiments, showing that molasses presents a buffering effect.
Overview Zymomonas mobilis, a Gram-negative bacterium, have been attracting increasing attention for fuel ethanol. It is an osmo and ethanol tolerant bacterium and it has shown higher specific rates of glucose uptake and ethanol production. Z. mobilis may have a greater potential for industrial ethanol production from raw sugar, sugarcane juice and sugarcane syrup. Molasses is an agro-industrial by-product often used in alcohol distilleries due to the presence of fermentative sugars, being an optimal carbon source for the microorganism metabolism. 2.1. Microorganism and culture conditions the strain used were Z. mobilis ATCC 29191. Culture The strain was maintained on agar plates containing (per liter): 200 g glucose, 10 g yeast extract, 5 g peptone, 1 g (NH4)2SO4, 2 g KH2PO4, 0.5 g MgSO4 Æ 7H2O and 0.5 g FeSO4 (Merck). The culture medium was sterilized at 121 _C for 15 min. The cultures were maintained at 4 C and renewed every five weeks. The inoculum culture was grown composed with sucrose at 200 g L-1 and the components mentioned previously. The cell concentration was standardized to 0.2 g L-1, determined by turbidimetry at k = 605 nm. The batch fermentations were carried out in duplicate in the sugar cane molasses culture medium, in the different culture conditions, according to the experimental design. Analytical methods After each fermentation, the culture was centrifuged (10, 0000 rpm for 15 min) and the biomass concentration was determined by measuring the turbidity of diluted sample at
605 nm using a standard curve of absorbance against dry cell mass. The total reducing sugars (TRS) were quantified Ethanol was determined by Gas Chromatography (GC) Shimadzu, using a DBWAX column (30.0 · 0.25 cm) with a flux of 40 ml min_1 and isopropanol as an internal standard. 3. Results and discussion
Optimize the ethanol production Four relevant factors for the fermentative process were selected. The variables, studied simultaneously were: TRS concentration in sugar cane molasses (150 and 250 g /L), temperature (25 and 35 C), agitation (180 oscillations per minute and static culture) and culture time (12 and 24 h), The main effects of the three factors concentration in sugar cane molasses, temperature, and are all positives, and agitation rate is negative. The growth time main effect is the most significant factorial design effect value for the production of ethanol. The inclusion of agitation rate reduces the average ethanol production. The results of the 3 variable showed that the condition of 200 g /L of TRS and temperature 30 C was the most favorable, achieving 54.83 g/L after a 48-hour-culture time. The time was a decisive factor, once the ethanol production increased to more than 60% from 24 to 48 h. By comparison reported a maximum ethanol concentration (55.3 g L_1) at 32.4 C, pH of 4.93 after 17.24 h from sago starch using Z. mobilis MTCC 92. Reported similar values (54 g/L) for Z. mobilis ZM4 from hydrolysed waste starch stream. In the central point, 250 g L_1 and 35 _C, ethanol production was an average of 31.15 g L_1. The decrease in ethanol production at high sugar concentration occurred due to an increase in the osmotic pressure that is one of the essential factors for by-products synthesis such as sorbitol and levan. The molasses was an industrial sucrose containing substrates that has been reported to contain substantial salt content. At 35 _C and 300 g L_1 sugars concentration on molasses obtained maximum sorbitol production by Z. mobilis ATCC 29191. The temperature of 40 _C was negative for fermentative process, resulting in lower productions, 4.6 g L_1. Numerous studies have shown that temperatures above 37 C are detrimental for ethanol production. Based on the results of 23 factorial designs, it was performed as a 22 factorial design, with central composite design, resulting in 11experiments (Table 3). In this stage the time was fixed in 48 h. With the central composite design it was possible to confirm that maximum ethanol concentration occurred at the central point, 55.8 g/L on average. These values are similar to the ones described for
ethanol production from sucrose and sago starch, which confirmed that the microorganism showed an optimal adaptation to the non-treated molasses. The ethanol productivity was a mean of 1.1 g L_1 h_1. Low biomass production is normally observed in Z. mobilis, and cell growth and fermentation are not linked. Approximately 2% of the carbon source is converted into biomass. This occurs due to Entner–Doudoroff pathway used by this microorganism. This pathway yields only a single mole of ATP per mole of sugar fermented, giving Zymomonas the lowest molar growth yield reported for a bacterium. The pH of the medium remained constant during the experiments, varying from 6.0 at the beginning to 5.6, on average. The pH has also been described as a factor that strongly interferes in the fermentative processes. However, according to molasses exhibits a buffering effect. This regulatory action depends of molasses chemical composition. The main stabilizer compounds of the pH are weak acids and amino acids that act in the acid range, mainly between pH 3.0 and 5.0, or phosphates, whose buffering effects occur in the range of 6.0 and 7.0.
Reference Fermentation of molasses by Zymomonas mobilis: Effects of temperature and sugar concentration on ethanol production M.L. Cazetta a,*, M.A.P.C. Celligoi b, J.B. Buzato b, I.S. Scarmino c
Kluyveromyces Marxianus Fuel ethanol production through yeast fermentation has generally not been found economically viable in hot climates. This is because of the high energy input required to maintain the fermentation process temperature at between 25 C and 35 C to maximize ethanol production and prevent irreversible heat-inactivation of the yeast cells. Yeasts, in general, grow at relatively low temperatures in comparison with bacteria, with upper temperature limits for thermo tolerant and thermophilic yeasts at 42 C and 45C respectively. Because of the advantages of thermo tolerant yeast in industrial fermentations, strains capable of growth and ethanol production above 40C have been actively sought, mainly through the screening of existing yeast strains a strain of Kluyveromyces marxianus growing up to 49 C and capable of producing alcohol at and above 40C. Members of the yeast genus Kluyveromyces were found to be more thermotolerant than Saccharomyces or Candida .
Strains the selection and optimization of a Saccharomyces yeast strain suitable for fermentation at a maximum temperature of 45 C but with incomplete sugar fermentation and a maximum of 4.5% (w/v) ethanol production from 15 to 20% The enrichment and isolation programme resulted in five thermo tolerant yeast cultures capable of growth at temperatures up to 52 C being isolated. These cultures were designated as IMB1, IMB2, IMB3, IMB4 and IMBS, and were believed to be different as they were selected according to minor morphological differences on plates, liquid culture and microscopic examination. The IDY Saccharomyces cerevisiae, in comparison, did not grow at temperatures above 40~
Molasses Fermentation The five yeast cultures fermented molasses and produced appreciable amounts of alcohol. The concentrations, however, were significantly less than that produced from glucose and it took a longer time for the fermentation to stop, as indicated by CO2 release. Maximum alcohol production of 5.6--6.0% (w/v) was at 40 C for all isolates.
However, little difference in alcohol concentration was observed at 37 C while a marked decrease in alcohol production, to 3.0 to 3.5% (w/v), was detected at 45 C. The IDY culture, in comparison, produced 6.0% (w/v) alcohol at both 37 C and 40 C and only 3.0% (w/v) at 45 C. Supplementation of molasses with P, K, Mg and Mn resulted in a significant increase in the alcohol concentrations produced for the five thermo tolerant isolates at
temperatures of 37 C, 40 C, and 45 C.
Molasses, however, is not as good a substrate as glucose, which may be due to the presence of some nonassimilatable reducing sugar complexes in the molasses or to the lack of some essential nutrient or element required by these cultures at elevated temperatures.
Supplement addition This latter view is supported by the fact that higher ethanol concentrations were produced by supplementing molasses with P, K, Mg and Mn. Similar increases in biomass yields on supplementation with selected elements have been noticed by the authors when working with thermophilic bacteria (data not reported). An increase in alcohol production and complete utilization of 15% glucose by thermo tolerant yeast strains growing at 43 C only when the other medium components were doubled to make them more readily available.
The requirement of magnesium at high temperatures for glycolytic enzyme production was suggested as a possible explanation. A high requirement for potassium at higher growth rates and temperatures in thermo tolerant yeasts. Interestingly, in the case of the IDY strain of Saccharomyces cerevisiae, the results show that it is capable of producing alcohol at the higher temperatures of 45 C and 50 C on both glucose and molasses. However, it should be noted that this strain had no ability to grow at these temperatures and it is only due to the fact that the inoculum were prepared at 30 C that we detected such alcohol production. No alcohol production would be detected if cells were required to grow aerobically at these temperatures before fermentation commenced. The results presented in this paper show the good potential in using these thermo tolerant IMB yeast cultures in industrial alcohol production and may create new areas of interest in biotechnological and industrial fermentation applications, particularly in hot regions.
Reference Isolation of thermotolerant, fermentative yeasts growing at 52 C and producing ethanol at 45 C and 50 C I.M. Banat*, P. Nigam and R. Marchant
