CN102703519A - Method for fermenting and producing cellulosic ethanol by means of pachysolen tannophilus - Google Patents
Method for fermenting and producing cellulosic ethanol by means of pachysolen tannophilus Download PDFInfo
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- CN102703519A CN102703519A CN2012102123280A CN201210212328A CN102703519A CN 102703519 A CN102703519 A CN 102703519A CN 2012102123280 A CN2012102123280 A CN 2012102123280A CN 201210212328 A CN201210212328 A CN 201210212328A CN 102703519 A CN102703519 A CN 102703519A
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Abstract
The invention provides a method for fermenting and producing ethanol by means of pachysolen tannophilus. According to the method, the pachysolen tannophilus is used as original strains; maize straw acid hydrolysate is used as a raw material; and the age, the liquid loading amount, the initial pH, the dissolved oxygen, the culture temperature, and the nitrogen sources of yeast which influences the fermentation and the production of the ethanol are researched.
Description
Technical field
The invention belongs to fermentation technical field, be specifically related to a kind of method of utilizing the pachysolen tannophilus fermentative production of ethanol.
Background technology
Ethanol is a kind of efficient, environmental protection, reproducible fuel and gasoline dope, greatly develops alcohol fuel, not only can effectively solve the deficient day by day difficult problem of the energy, can effectively reduce environmental pollution more.The alcohol production that China is traditional is raw material with grain generally, and not only cost height but also output are also received the restriction of grain resource, and output is limited, is difficult to satisfy the demands.In recent years, beginning the someone studies with agriculture and forestry organic waste materials such as agricultural crop straw, grass, barks to be raw material, to utilize microbial technique that cellulose conversion is become ethanol.Occurring in nature contains abundant cellulose series biomass, and only 700,000,000 tons nearby of agricultural crop straw YO of China are if utilize biofermentation technique; It is used for production of cellulosic ethanol; Not only realized the making full use of of agricultural waste resource, and reduced consumption, practiced thrift resource grain.Therefore, cellulosic ethanol is a kind of biofuel of tool development prospect.
At present, make cellulosic ethanol realize commercialization, need to solve two gordian techniquies, the one, improve cellulosic hydrolysis efficiency, the 2nd, improve mikrobe to cellulosic transformation efficiency.Mierocrystalline cellulose and semicellulose can generate wood sugar, glucose, pectinose etc. through hydrolysis, improve cellulosic hydrolysis efficiency, can effectively increase the content of a hexose pentose in the hydrolyzed solution, are cellulosic ethanol thereby make more sugar through microbial transformation.And can mikrobe efficiently utilize pentose and the hexose in the hydrolyzed solution simultaneously, and it is converted into ethanol, also is to utilize the cellulosic resource to produce the alcoholic acid key factor.
This patent is starting strain with the pachysolen tannophilus, is raw material with the corn straw acid hydrolysis liquid, and the yeast kind age, inoculum size, initial pH, dissolved oxygen, culture temperature, the nitrogenous source that influence fermentative production of ethanol are studied.
1. method of utilizing pachysolen tannophilus fermentative prodn cellulosic ethanol; It is characterized in that fermentation raw material is the acid hydrolysis liquid of corn straw; A certain amount of fermention medium of in 500 mL Erlenmeyer flasks, packing into during fermentation; Insert a certain amount of pachysolen tannophilus seed liquor, respectively in certain initial pH, rotating speed and certain culture temperature bottom fermentation regular hour.
2. method according to claim 1 is characterized in that the preparation method of raw material corn straw acid hydrolysis liquid is: corn straw is crushed to 40～60 orders, and the dilute sulphuric acid with 1.5%～3% soaks; Solid-to-liquid ratio 1:8; 121 ℃ of hydrolysis 1～2h, cooled and filtered is got filtrating.
3. method according to claim 1 is characterized in that fermention medium is the corn straw acid hydrolysis liquid that has added certain nitrogenous source, metals ion.
4. method according to claim 1 is characterized in that the making method of seed liquor is: the bacterium colony of slant culture is inserted seed culture medium cultivation 29～32 h, process seed liquor.
5. method according to claim 4 is characterized in that, seed culture medium (g/L): glucose 15, wood sugar 5, Tryptones 15, yeast extract paste 10, MgSO
40.3, CaCl
20.3, pH5.5, liquid amount 100 mL/500 mL Erlenmeyer flasks, 115 ℃ of sterilization 20 min.
6. method according to claim 1 is characterized in that it is 29～32 h that the best is planted age.
7. method according to claim 1 is characterized in that best liquid amount is the 150mL/500mL triangular flask.
8. method according to claim 1 is characterized in that optimum revolution is 90 r/min.
9. method according to claim 1 is characterized in that best initial pH is 5.0.
10. method according to claim 1 is characterized in that optimum culturing temperature is 30 ℃.
11. method according to claim 1 is characterized in that optimum nitrogen source is yeast extract paste 6 g/L.
Description of drawings
Best the confirming of age of planting of Fig. 1 seed;
The initial pH of the different substratum of Fig. 2 is to the influence of ethanol production;
Fig. 3 different rotating speeds and liquid amount are to the influence of ethanol production;
The different culture temperature of Fig. 4 are to the influence of ethanol production;
Fig. 5 different nitrogen sources is to the influence of ethanol production.
Embodiment
Following embodiment elaborates to the present invention, but to not restriction of the present invention.
This case study on implementation is used for selecting best age, the seed culture medium (g/L): glucose 15, wood sugar 5, Tryptones 15, yeast extract paste 10, MgSO of planting
40.3, CaCl
20.3, pH5.5, liquid amount 100 mL/500 mL Erlenmeyer flasks, 115 ℃ of sterilization 20 min.Pachysolen tannophilus bacterium colony on the inclined-plane is inserted in the seed culture medium, and the growth curve of drawing yeast culture is as shown in Figure 1.
Can be known that by Fig. 1 the lag period of pachysolen tannophilus is shorter, when being cultured to 29～32 h, barms is in the logarithmic growth end of term, and therefore selecting kind of an age is 29～32 h.
This case study on implementation is explained the influence of the initial pH of different substratum to ethanol production; Seed liquor is inserted in the fermention medium with 5% inoculum size; Initial pH is respectively 3～7; In 30 ℃, 100 r/min concussion cultivation, 48 h, the substratum liquid amount is 100 mL/500 mL triangular flasks, and the result is as shown in Figure 2.
PH mainly passes through to influence somatic cells membrane charge, membrane permeability and nutritive substance degree of ionization, thereby influences thalline to nutrient absorbing.Because the pH in the shake flask fermentation process is difficult to control, can only control the initial pH of fermented liquid.Can be known that by Fig. 2 when the initial pH of substratum was 3.0～5.0, alcoholic acid output was in a higher level, along with the continuation rising of pH, ethanol production has obvious decline, and initial pH is that 5.0 o'clock output is the highest, therefore selects pH5.0 as initial pH.
This case study on implementation explanation dissolved oxygen mainly is the influence to ethanol production of different liquid amounts of investigation and different rotating speeds to the influence of ethanol production.With in 5% the inoculum size access fermention medium, initial pH is 5.0, under 30 ℃ of conditions, cultivates 48 h with seed liquor.Liquid amount is respectively 90,110,130,150 mL/500 mL triangular flasks, and rotating speed is respectively 60,90,120,150 r/min, obtain two factors to ethanol production to influence the result as shown in Figure 3.
Pachysolen tannophilus is a kind of facultative anaerobe, and dissolved oxygen is bigger to growth and the metabolic effect of bacterium, can be known by Fig. 3, and when liquid amount was 90 mL/500 mL triangular flasks, rotating speed was big more, and dissolved oxygen is big more, and alcoholic acid output is low more on the contrary; And when liquid amount was respectively 110,130,150 mL/500 mL triangular flasks, along with the increase of rotating speed, ethanol production presented the trend of falling after rising.When liquid amount be 150 mL/500 mL triangular flasks, when rotating speed is 90 r/min, ethanol production is maximum, is 6.71 g/L, therefore 150 mL/500 mL triangular flasks are best liquid amount, and 90 r/min are optimum revolution.
This case study on implementation is explained the influence of different culture temperature to ethanol production; Seed liquor is inserted in the fermention medium with 5% inoculum size; Initial pH is 5.0; Liquid amount is 150 mL/500 mL triangular flasks, and rotating speed is 90 r/min, respectively at cultivating 48 h under 25 ℃, 28 ℃, 30 ℃, 32 ℃, 35 ℃, the 37 ℃ conditions.Obtain culture temperature to ethanol production to influence the result as shown in Figure 4.
Pachysolen tannophilus upgrowth situation when cultivating for 30 ℃ is best, and is also the fastest to the spending rate of sugar.Can be known that by Fig. 4 in 6 culture temperature, alcoholic acid output is also maximum in the time of 30 ℃, peak is 6.98 g/L, and therefore, 30 ℃ is the optimum temperuture of fermentative production of ethanol.
This case study on implementation explanation different nitrogen sources influences alcoholic acid; Seed liquor is inserted in the fermention medium with 5% inoculum size; Initial pH is 5.0, and liquid amount is 150 mL/500 mL triangular flasks, and rotating speed is 90 r/min; Adding concentration in the substratum respectively is ammonium sulfate, urea, peptone, yeast extract paste and the Carnis Bovis seu Bubali cream of 4,6,8,10,12 g/L, and it is as shown in Figure 5 to the influence of ethanol production to obtain different nitrogen sources.
Can know that by Fig. 5 along with the increase of urea concentration, ethanol production reduces gradually; Along with the increase of peptone, Carnis Bovis seu Bubali cream, yeast extract paste concentration, ethanol production is fallen after rising, and nitrogen concentration is 6 g/L during peak; Ethanol production when using organic nitrogen source in peptone, Carnis Bovis seu Bubali cream and the yeast extract paste 3 apparently higher than other two kinds inorganic nitrogen-sourced; Use 3 kinds of organic nitrogen source output difference little, using yeast extract paste, when concentration is 6 g/L; It is maximum that output reaches; Therefore peak 7.91 g/L confirm that optimum nitrogen source is a yeast extract paste, and concentration is 6 g/L.
Claims (10)
1. method of utilizing pachysolen tannophilus fermentative prodn cellulosic ethanol; It is characterized in that fermentation raw material is the acid hydrolysis liquid of corn straw; A certain amount of fermention medium of in 500 mL Erlenmeyer flasks, packing into during fermentation; Insert a certain amount of pachysolen tannophilus seed liquor, respectively in certain initial pH, rotating speed and certain culture temperature bottom fermentation regular hour.
2. method according to claim 1 is characterized in that the preparation method of raw material corn straw acid hydrolysis liquid is: corn straw is crushed to 40～60 orders, and the dilute sulphuric acid with 1.5%～3% soaks; Solid-to-liquid ratio 1:8; 121 ℃ of hydrolysis 1～2h, cooled and filtered is got filtrating.
3. method according to claim 1 is characterized in that fermention medium is the corn straw acid hydrolysis liquid that has added certain nitrogenous source.
4. method according to claim 1 is characterized in that the making method of seed liquor is: the bacterium colony of slant culture is inserted seed culture medium cultivation 29～32 h, process seed liquor.
5. method according to claim 4 is characterized in that seed culture medium: glucose 15 g/L, wood sugar 5 g/L, Tryptones 15 g/L, yeast extract paste 10 g/L, MgSO
40.3 g/L, CaCl
20.3 g/L, pH5.5, liquid amount 100 mL/500 mL Erlenmeyer flasks, 115 ℃ of sterilization 20 min.
6. method according to claim 1 is characterized in that be 29～32 h kind of age.
7. method according to claim 1 is characterized in that rotating speed is 90 r/min.
8. method according to claim 1 is characterized in that initial pH is 5.0.
9. method according to claim 1 is characterized in that culture temperature is 30 ℃.
10. method according to claim 1 is characterized in that nitrogenous source is yeast extract paste 6 g/L.
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Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN101085994A (en) * | 2006-06-06 | 2007-12-12 | 河南农业大学 | Ferment method of producing fuel ethanol using stalk material |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101085994A (en) * | 2006-06-06 | 2007-12-12 | 河南农业大学 | Ferment method of producing fuel ethanol using stalk material |
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| Title |
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| 潘春梅等: "玉米秸秆稀酸水解液发酵生产纤维素乙醇的研究", 《中国酿造》 * |
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Application publication date: 20121003 |