CA1143677A - Process for producing ethyl alcohol from amylaceous raw stock - Google Patents
Process for producing ethyl alcohol from amylaceous raw stockInfo
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- CA1143677A CA1143677A CA000349907A CA349907A CA1143677A CA 1143677 A CA1143677 A CA 1143677A CA 000349907 A CA000349907 A CA 000349907A CA 349907 A CA349907 A CA 349907A CA 1143677 A CA1143677 A CA 1143677A
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02E—REDUCTION OF GREENHOUSE GAS [GHG] EMISSIONS, RELATED TO ENERGY GENERATION, TRANSMISSION OR DISTRIBUTION
- Y02E50/00—Technologies for the production of fuel of non-fossil origin
- Y02E50/10—Biofuels, e.g. bio-diesel
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Abstract
PROCESS FOR PRODUCING ETHYL ALCOHOL FROM AMYLACEOUS
RAW STOCK
Abstract of the Disclosure The process for producing ethyl alcohol from amylace-ous raw stock consists in carrying out hydrolysis of said amylaceous raw stock such as grains of wheat, barley or rye, in the presence of amylolytic and cellulosolytic en-zymes.
Used as cellulosolytic enzymes is the preparation of the culture of a mould fungus Trichoderma k?ningii, con-taining a complex of enzymes, viz., C1-enzyme, endoglucana se, exoglucanase, cellobiase, xylanase, .beta.-glucosidase, protease, and a number of amylolytic enzymes.
RAW STOCK
Abstract of the Disclosure The process for producing ethyl alcohol from amylace-ous raw stock consists in carrying out hydrolysis of said amylaceous raw stock such as grains of wheat, barley or rye, in the presence of amylolytic and cellulosolytic en-zymes.
Used as cellulosolytic enzymes is the preparation of the culture of a mould fungus Trichoderma k?ningii, con-taining a complex of enzymes, viz., C1-enzyme, endoglucana se, exoglucanase, cellobiase, xylanase, .beta.-glucosidase, protease, and a number of amylolytic enzymes.
Description
~ ~ 4 ~ 7 PROCESS FOR PRODUCING ~THYL A~COHO~ FROM AMYLACEOUSRAW S~OCK
The present invention relates to the fermentation branah of eood indus~ry a~d is concerned with processes for producing ethyl alcohol from amylaceous raw stock, such as, say, wheat, barley, or rye.
Prior-art processes for producing ethyl alcohol from amylaceous raw stock are kno~n to be based on hydrolysis and fermentation, and to comprise the following stages:
/1/ crushing the corn grain3;
/2/ rendering the grains pastelike and grain cooking;
/3/ enzymic hydrolysis of cereal starch;
/4~ fermenting;
/5/ wash distilling.
There is ex~ensively applicable currently a process for producing ethyl alcohol ~rom amylaceous raw stock9 su¢h as rye, barley, or wheat by hydrolyzing in the presence of malt enzymes. Malt is known to comprise predominantly ~_ and ~ -amylases that h~drolyze s~arch to maltose and gluco-se, which are then yeast-fermented into alcohol.
The above-discussed process i~volves, however~ top-grade grains ~or malt preparation, which there~ore is a hi~hly critical product. In additiont non~amylaceous sub-stances, such as cellulose, hemicellu1ose9 and some others, contained in grains are not hydrolyzable by the malt ensymes.
3~'7~7 _ 2 _ ~ urther improvement in the ethyl alcohol production techniques has heretofore been aimed at searching ~or such microbial-origin enzymic preparations that would be able to hydrolyze starch, hemicellulose, cellulose, and some other substances contained in corn ~rains.
~ here have been proposed some processes for producing ethyl alcohol ~rom amylaceous raw stock by hydrolyzing the latter in the presence of amylolytic enz~mes produced by mould fungi &nd bacteria. Such ~ungial and bacterial enzymic preparations possess more potent complex of enzymes than malt and hence are capable of hydrolyzing the starch-con-taining grain portion somewhat more completely. On that account the yield o~ e~hyl alcohoi pe~ unit o~ grain raw stock is higher due to the e~ect o~ fungial- or bacterial-origin amylolytic enzymes. Substitution o~ malt for the above enzymic preparations in producing ethyl alcohol enables one to save to a great e~tent high-grade corn grains which have heretofore been spent in great a~ounts for malt preparation-However, the afore-mentioned enæymic preparations ob-tained from mould fungi and bacteria, can hydrolyze starch only and practically produce no hydrolyzing effect upon non-amylaceous carbohydrates, such as cellulose and hemicellulose 9 and also somc other polysaccharides-One prior-art process ~or producing ethyl alcohol is known to conzist in hDdrolyzing corn grainz in the prezenoe , 1 ~ ~ 3 ~ ~
of an enzymic preparation obtained ~rom the grown culture of Aspergillus ni~e.r /c~. USSR Inventor~s Certificate No. ~7,012, cl~C 1~c 7/04, published on February 20, 1941/-However, raw stock h~ydrolysis occurs in this case but in-completely, and a great proportion o:~ carbohydrates involv-ed remain unfermented. ~his is due to the fact that the above preparation fails to contain a required complex of enzymes.
One more prior-art process for producing ethyl alcohol from corn grains is known to involve the use of an enzymic preparation obtained from the grown culture of a mould .fungus named Aspergillus oryzae /cf. USSR Inventor~s Certi-.ficate ~oO119,852, cl C 12c 7/04, published in 1958/. Howe-ver, this preparation fails to improve hydrolysis, too.
An enzymic preparation of phosphatase has bee~ propos-ed for carrying out more complete hydrolysis of amylaceous raw stock /cf. USSR Inventor~s Certificate No.460,292,cl-C
12c 7/04, published on May 15, 197~/. But even this prepara-tion ~ails to bring about complete h~rdrolysis o~ amylaceous raw stockO
The eng~mes listed above are engaged only in hydrolysis of t'ne amylaceou~ portion o~ corn grains and produce practi-cally no e~ect upon such hardl~ hydrolyzable grain poly-saccharides as cellulose and hemicellulose whose hydrolyza-tes can be converted into ethyl alcohol.
~3~;~77 _ 4 ~
~ here have also been proposed some processes ~or pro-ducing ethyl alcohol involving the use of enzymes capable o~ hydrolyzing carbohydrates con~ained in the grain wall cells. ~rhe enzymes mentioned above are produced by ~richo-derma viride /c~- US Patent No.3,616,220, cl-C 12b 1/00 published on October ~6, 1971/~ AsperOillus niger /cf. Pa-tent of GDR No.77,176, cl~6a2~ 3/10 published on October 20, 1970/, Trichotecium roseum /c~. USSR Inventor~s Certi-~icate No.3169719, cl.C 12c 7/04 published on October 7, l971/. However, the above-mentioned producers ~ail to gene_ rate such an active complex o~ hydrolytic enzymes that is required ~or comple~e and thorough-going hydrolysis of grain raw stock-A process for saccharifying grain raw stock with ami~ture of malt and an enzymic preparation ~rom Aspergillus awamori mould fungus containing xylanase and~ -~lucanase, is protected by USSR Inventor~s Certificate No.467,9Z9, cl~G 12c 7/04 published on December 25, 1974. ~his prepara-kion hydrolyzes xylane and ~ -glucane~ whereby the yield of eth~l alcohol is increased by 2 to 2.5 per cent~ However, the application o~ the above en~ymic preparation enables one to substitute malt by 30 to 35 per cent only.
Known in the art are some methods o~ cellulose process-ing in the presence o~ highly puri~ied, concentrated or immobilized enzymes obtained from the cultures o~ microor-.
~3~77 ganisms grown by the submerged cultivation method, e.g~,~richoderma viride cult~re /c~. US Patent No. 3,542,580, cl.C 1~d 13~04 pub].ished ~ebrua.ry 15, 1972/. Canadian Pat-ent ~o.975,313, cl. 195-16 pu~lished on February 23, 1972, discloses a process for treating starch and protein-con-taining vegetable material with proteolytic enzymes, whereas ~rench Patent No-2,382,497, cl. C 12c 11/12 publish-ed on November 3, 1978 deals with a process for producing ethyl alcohol by virtue o~ cellulose fermenting under the e~ect o~ a miæed culture at a temperature of 50 to 65C
and the pH of 7 to 8. However, this patent fails to speci~y what producers are employed-It is an e~sential object of the present invention tos~arch for such an enzymic preparation ~hat would contain the required complex of h~drolytic enzymes capable o~ hydro-lyzing the nonam~laceous portion of the raw stock into fer-mentable sugars, thereby increasing the yield o~ ethyl alcohol.
~ he above object is acoomplished due to the provision o~ a novel enzymic preparation~ containing a complex of hydrolytic enzymes produced by a mould fungi ~richoderma koningii which is capable o~ more complete hydrolysis of amylaceous raw stock-According to the present invention, a process for pro-ucing ebhyl aloohol ~rom amylaceous raw stock by ibs hydro-,.
~4~ 7~
l~sis under the e~fect of am~lolytic enzymes is characteriz-ed by that hydrolysis is carried out in the presence o~ an enzymic preparation o~ cellulase obtained ~rom the culture of ~richoderma koningii, containing a complex o~ hydrolytic enzymes, viz., C1-enzyme? endo- and exoglucanase, cellobiase, x~lanase, J~ -glucosidase, protease, and a number o~ amylo-lytic enæymes-The principal e~zyme~ making part of the afore-mention-ed proparation ~eature the ~ollowing activity /in terms o~
units per gramJ:
C1-enzyme/with respect to paper~ _ 100 to 125 endoglucanase - 15 to 20 exoglucanase - 3 to 4 cellobiase - 4 to 6 ~ylanase - 200 to 250 Optimum consumption o~ the above preparation equals 1 to 2 per cent of the entire mass of the amylaceous raw stock invol~ed due to the ~act ~hat in the abovesaid consumption limits more complete and economieally advantageous h~dro-lysis is attained ~ he present invention enables one to carry out deeper hydrolysis o~ grain polysaceharides and increase the ~ield of ethyl alcohol by 3 to 4 per cent as compared to the known processes. ~his is accounted for by the fact that the provi-sion of such enzymes as C1-enz~me, endoglucanase, exogluca-nase, oellobiase, and others in the proposed preparation, 3~;77 are capable of hydrolyzing grain polysaccharides o~ the nonamylaceous nature, such as cellulose and hemicellulose, into ~ermentable carbohydrates, with the result that the yield of ethyl alcohol is increased and the amount of lost unfermented sugars is diminished-The above-mentioned enzymic p~eparation of cellulase lS obtai~ed ~rom culti~ating a mould fungus ~richoderma koningii on a solid nutrient medium, containing the follow-ing components /in terms of mass per cent/:
wheat bran - 40 to 45 beet pomace - 20 to 25 malt sprouts - ~5 to ~0 sawdust - 5 to 10 Culti~ation is carried out at 30 to ~5C ~or 48 to 55 hours. On terminating the cultivation process the re-sultant enzymic preparation can be made use of in alcohol production or as the mould culture together with the re-mainder of the nutrient medium and mycelium, disintegrated and dried to a moisture content of 12 or 1~ per cent; or else in the form o~ a preparation obtained by precipitating the enzymes ~rom aqueous extracts of the mould fungus cul-ture, b~ organic solvents-Application of unpurified enzgmic preparation in the form of the culture of the above mould fungus is economi.cal-ly more reasonable, as this rules out e~penditures for iso-lati~g -the enzymes and prevents losses of their activity in .
,.
, "
1~3tj77 the course o~ isolation~
~ he process of the present i.n~ention is carried out as Pollows. Am~laceous raw stock, 9 .g.1 wheat grains, is ~ed to the crusher to be disintegrated there, wherefrom the stock enters the mixer-pre~oiler composed of two compart-ments. In the first compartment the crushed grains is mixed with water in a weight ratio o~ 1 4, whereupon the mixture is forwarded to ~he second compartment, where it is heated to 80 or 85C. While the mixture is being heated starch is rendered pasteliXe- Next the pastelike mass is delivered from the preboiler to the cooker arrangement, wherein the mass is cooked at a pressure of 0-4 to 0.5 MPa for 30 to 50 min. ~hen the cooked mass is delivered to the steam separator for getting it read of steam, and from there to the 1st-stage saccharifier to be cooled to the temperature of hydrolysis /58 to 60C/. Fed to the sacchari~ier in an is amount of 30 per cent an aqueous suspension of a mixture of the hydrolytic enzymes obtained from surface cultures o~
mould fungi7 the above suspension containing 1 per cent -amylase from Aspergillus.oryzae /in terms of starch mass/, 4 per cent glucoamylase ~rom Aspergillus awamori Jin terms of the starch mass/, and.1~5 per cent o~ a comp-lex preparation of cellulase from the culture o~ ~richo-derma Xoningii /in terms of whole mass of raw stock~ ~he above aqueous suspension is treated with formalin in a 0-02 mass per cent concentration-~ ~3~i77 It is in -the 1st-stage saccharifier that the cooked mass is lique~ied and partially ~ydrolyzed under the effect o~ the aLore-mentioned enzymes, the process proceeding at 58 to 60C ~or 10 min. From the 1st-stage saccharifier the mass is transferred to the 2nd-stage saccharifier to which are added the remaining 70 per cent of the above aqueous suspension o~ the hydrolytic enzymes grown on surface cul-tures of mould fungi, whereupon starch and nonamylaceous polysaccharides undergo thorough hydroly~is in the 2nd-stage saccharifier at 57 to 58C for 2 to 5 minutes. Ne~t the finally h~drolyzed mass is transferred from the 2nd-stage saccharifier to the heat e~changer to be cooled to the fermentation temperature /30C/ and from there to the fermenter vat, into which is also introduced yeast ~accha~
romyces cerevisiae XII in an amount of 6 to 8 per cent of the fermenter ~at use~ul capacity. ~hereupon the sacchari-fied mass undergoes fermentatio~ under the effect of the above-mentioned yeast, the fermentation process occurring at 28 to 30C for 4B to 50 hours-The ~ermentation process over, the mature yeast washis fed to the rectifying still. A 4-per cent increase in the yield of ethyl alcohol per unit o~ the original amylace-ous stock is attained as compared to the ethyl alcohol pro-ducing process diispensing with the proposed preparation-~ he enzymic preparation of cellulose made use of forhydrolysis, is obtained by surface cultivatine o~ s mould .
3f~
_ 10 _ ~ungus Trichoderma koningii on a solid nutrient medium, containing the Eollowing components /in terms of mass per cen ~:
wheat bran - 45 malt sprouts - 25 beet pomace ~ 25 . sawdust - 5 . The moisture content of the nutrient medium is within ,'. 60 to 65 per cent, the cultivation occurring at 30 to 35C
~or 48 to 55 hours.
The finished culture o~ ~richoderma k'oningii fungus is in ~act an enz,ymic preparation of cellulase. Such'pre-~' paration contains a complex o:E hydrolytic enz~es, mostly cellulosoly~ic enz~mes and xylanase Eeaturing the following activit,y ~i~ terms o~ units per gram~::
~. C1-enz~me /with respect to paper/ - 100 to 125 . endoglucanase - 15 to 20 ;~ exoglucanase - ~ to 4 cellobiasa - 4 to 6 x 'xylanase _ 200 to 250 he thus-obtained enz~mic preparation o~ cellulase is ~; used for h,ydrol~sis in t~e form of a ~ungus culture "per ; sef~ without isolating and puri~ing with organic solvents, .~ an advantag~ous feature that rules out costly and sophis-ticated stage o~ isolating the enz~mes producing cellulase effect and pr0vents the loss of their ~ermenting activity ,;, .
.,, :
., .
...
,, ,,, : , -.
. . .
., ~3t;,~7 _ 11 _ while being isolated.
To promote understanding o~ the present invention given below are some examples o~ its practical embodiment carried out under laboratory conditions.
Fxample 1 50 g ground wheat grains is placed in a dry Erlenmeyer flask 0-5 1 in capacity, into which is added 200 ml water, whereupon the contents are stirred and rendered pastelike in a boiling water bath ~or 40 min. ~hen the ~lask lS re-moved from the bath and closed with glass covers, whereupon the ~lask is placed in a digester where its contents are cooked at a pressure of 0-15 MPa ~or 90 min. A~ter cooking the flask is removed ~rom the digester, and added thereto ~or the cooked mass to liquef~ is 30 per cent o~ a mixture o~ enzymes, aontainlng the following components /in terms o~ per cent o~ starch mass~ CC-am~lase from Aspergillus oryzae~, 1; glucoamylase ~rom Aspergillus awamori, 4; and the cellulase preparation ~rom ~richoderma k~oni~gii, 1.5 /in terms o~ mass o~ grains/, the above preparation con-taining a complex of hydrol~tic enzymes, i-e-, C1-enzyme, endoglucanase, exoglucanase, cellobiase, xylanass, B -clu-cosidase, protease 9 and a number of amylolytic enzymes. ~he mass is ¢ooled to 58C, and bhe rest o~ the mixture o~ en-zymes /70 per cent/ is added thereto, whereupon the mass is subjected to h~drol~sis at the above-mentioned tempera-ture ~or an hour-1~L43~77 _ 12 _After the h~drolysis the flask contents are cooled to 30C and doped with a suspension of yeast Saccharomyces cerevisiae XII /6 per cent of ~he mass/, whereupon some ~ormalin /0.02 per cent o~ the mas~/ is added ~or sterility.
Then the ~lask is closed and placed in a temperature-con-trolled cabinet for fermentation, which occurs at ~0G
within 72 hours. Next ethyl alcohol is distilled ~rom the resultant yeast wash in a distiller-~he yield o:~ ethyl alcohol equals 19.2~ ml from 50 g wheat containing 56.4 per cent starch, i.e. 104 per cent.
Used in Example 1 is the preparation of cellulase ~eaturing the following activity o~ the basal enzymes /in terms of units per gram/:
C1-enzyme /with respect ~o paper/ - 125 endoglucanase - 17 exoglucanase - 3 cellobiase _ 5 xylanase - ~19 ~he above preparation is obtained b~ cultivating a mould fungus ~richoderma koningii on a solid nutrient medi-um, containing the ~ollowing components /in terms o~ mass per cent/:
wheat bran - 45 malt sprouts - 25 beet pomace - 25 sawdust - 5 ,, ,.
,, 3f~77 The thus-obtained fungus culture is used in a native state 9 i.e., without isolating the enzymes from the nutri-ent medium and their puri~ying, which ~o a great extent simplifies production and utilization of the preparation-Exampla 2 Wheat grain~ /in an amount of 1000 k~ are continuous-ly fed to the grain crusher ~to be disintegrated there, whereupon the crushed grains are ~ed to the mixer-preboiler, composed o~ two compartments. Water is also fed to the 1st compartment in an amount four times the grain mass. Prelimi-nary heat treatment o~ grains occurs in the 1st compart-ment, whereas final heat treatment of grains, i.e., render-ing them pas~elike, is carried out in the 2nd compartment at 80 to 85C for 2 to 5 minutes. For ~inal heat treatment of the pastelake mass it is fed to the cooker arrangement to be coo~ed at a pressure of 0.5 MPa for 50 minutes.
Thereupon the mass is transferred to the steam separator for getting the mass read o~ steam, wherefrom the ~ass is forwarded to the 1st-stage saccharifier. On ¢ooling the mass to 58 or 60C addsd thereto is an aqueous suspension /30 per cent of the whole amount/ o~ a mixture of hydro-lytic enzymes o~ surface cultures of mould fungi, containing 1 per cent ~ -am~lase ~rom Aspergillus oryzae, 4 per cent glucoamylase from Aspergillus awamori /both in ~erms o~
starch mass/, and 1.5 per cent /in terms of preparation mass/ o~ the cellula~e preparation obtaine~ ~rom the culture ' , of a mould fungus Trichoderma koningii o~ the composition as in ~xample 1. Then the aqueous suspension is treated with an antiseptic, say, ~ormalin in a concentration of 0.02 mass per cent- The process o~ hydrolysis in the 1st-stage sacchari~ier takes 10 minutes, whereupon the mass is transferred to the 2nd~stage sacchariXier to which is added the remaining 70 per cent of the above aqueous suspension of the hydrolytic enzymes of surface cultures. It is in the ~-2nd-stage saccharifier that starch and nonamylaceous pol~-saccharides undergo thorough hydrol~sis at 57 to 58C ~or 5 minutes. Hydrolysis ovsr, the mass is let cool to 30C in a heat exchanger and trans~erred to the ~ermenter vat, to which is added yeast Saccharomyces cere~isiae XII /6 to 8 per cent o~ the fermenter use~ul capacity/, where the mass is ~ermented at 28 to 30C ~or 72 hours. ~he fermentation procèss over, the mature yeast wash is fed to the recti~-ing still to obtain ethyl alcohol there~rom- The ~ield of ethyl alcohol per ton o~ starch e~uals 66.56 dal~ i.e-, by 4 per cent higher as compared to the control procefis carried out according to the known method, that is5 without the pro-posed preparation. An increased ethyl alcohol yield is at-tained due to ~ermenting the carbohydratss resulting ~rom hydrol~sis o~ the nonam~laceous portion o~ corn grains-
The present invention relates to the fermentation branah of eood indus~ry a~d is concerned with processes for producing ethyl alcohol from amylaceous raw stock, such as, say, wheat, barley, or rye.
Prior-art processes for producing ethyl alcohol from amylaceous raw stock are kno~n to be based on hydrolysis and fermentation, and to comprise the following stages:
/1/ crushing the corn grain3;
/2/ rendering the grains pastelike and grain cooking;
/3/ enzymic hydrolysis of cereal starch;
/4~ fermenting;
/5/ wash distilling.
There is ex~ensively applicable currently a process for producing ethyl alcohol ~rom amylaceous raw stock9 su¢h as rye, barley, or wheat by hydrolyzing in the presence of malt enzymes. Malt is known to comprise predominantly ~_ and ~ -amylases that h~drolyze s~arch to maltose and gluco-se, which are then yeast-fermented into alcohol.
The above-discussed process i~volves, however~ top-grade grains ~or malt preparation, which there~ore is a hi~hly critical product. In additiont non~amylaceous sub-stances, such as cellulose, hemicellu1ose9 and some others, contained in grains are not hydrolyzable by the malt ensymes.
3~'7~7 _ 2 _ ~ urther improvement in the ethyl alcohol production techniques has heretofore been aimed at searching ~or such microbial-origin enzymic preparations that would be able to hydrolyze starch, hemicellulose, cellulose, and some other substances contained in corn ~rains.
~ here have been proposed some processes for producing ethyl alcohol ~rom amylaceous raw stock by hydrolyzing the latter in the presence of amylolytic enz~mes produced by mould fungi &nd bacteria. Such ~ungial and bacterial enzymic preparations possess more potent complex of enzymes than malt and hence are capable of hydrolyzing the starch-con-taining grain portion somewhat more completely. On that account the yield o~ e~hyl alcohoi pe~ unit o~ grain raw stock is higher due to the e~ect o~ fungial- or bacterial-origin amylolytic enzymes. Substitution o~ malt for the above enzymic preparations in producing ethyl alcohol enables one to save to a great e~tent high-grade corn grains which have heretofore been spent in great a~ounts for malt preparation-However, the afore-mentioned enæymic preparations ob-tained from mould fungi and bacteria, can hydrolyze starch only and practically produce no hydrolyzing effect upon non-amylaceous carbohydrates, such as cellulose and hemicellulose 9 and also somc other polysaccharides-One prior-art process ~or producing ethyl alcohol is known to conzist in hDdrolyzing corn grainz in the prezenoe , 1 ~ ~ 3 ~ ~
of an enzymic preparation obtained ~rom the grown culture of Aspergillus ni~e.r /c~. USSR Inventor~s Certificate No. ~7,012, cl~C 1~c 7/04, published on February 20, 1941/-However, raw stock h~ydrolysis occurs in this case but in-completely, and a great proportion o:~ carbohydrates involv-ed remain unfermented. ~his is due to the fact that the above preparation fails to contain a required complex of enzymes.
One more prior-art process for producing ethyl alcohol from corn grains is known to involve the use of an enzymic preparation obtained from the grown culture of a mould .fungus named Aspergillus oryzae /cf. USSR Inventor~s Certi-.ficate ~oO119,852, cl C 12c 7/04, published in 1958/. Howe-ver, this preparation fails to improve hydrolysis, too.
An enzymic preparation of phosphatase has bee~ propos-ed for carrying out more complete hydrolysis of amylaceous raw stock /cf. USSR Inventor~s Certificate No.460,292,cl-C
12c 7/04, published on May 15, 197~/. But even this prepara-tion ~ails to bring about complete h~rdrolysis o~ amylaceous raw stockO
The eng~mes listed above are engaged only in hydrolysis of t'ne amylaceou~ portion o~ corn grains and produce practi-cally no e~ect upon such hardl~ hydrolyzable grain poly-saccharides as cellulose and hemicellulose whose hydrolyza-tes can be converted into ethyl alcohol.
~3~;~77 _ 4 ~
~ here have also been proposed some processes ~or pro-ducing ethyl alcohol involving the use of enzymes capable o~ hydrolyzing carbohydrates con~ained in the grain wall cells. ~rhe enzymes mentioned above are produced by ~richo-derma viride /c~- US Patent No.3,616,220, cl-C 12b 1/00 published on October ~6, 1971/~ AsperOillus niger /cf. Pa-tent of GDR No.77,176, cl~6a2~ 3/10 published on October 20, 1970/, Trichotecium roseum /c~. USSR Inventor~s Certi-~icate No.3169719, cl.C 12c 7/04 published on October 7, l971/. However, the above-mentioned producers ~ail to gene_ rate such an active complex o~ hydrolytic enzymes that is required ~or comple~e and thorough-going hydrolysis of grain raw stock-A process for saccharifying grain raw stock with ami~ture of malt and an enzymic preparation ~rom Aspergillus awamori mould fungus containing xylanase and~ -~lucanase, is protected by USSR Inventor~s Certificate No.467,9Z9, cl~G 12c 7/04 published on December 25, 1974. ~his prepara-kion hydrolyzes xylane and ~ -glucane~ whereby the yield of eth~l alcohol is increased by 2 to 2.5 per cent~ However, the application o~ the above en~ymic preparation enables one to substitute malt by 30 to 35 per cent only.
Known in the art are some methods o~ cellulose process-ing in the presence o~ highly puri~ied, concentrated or immobilized enzymes obtained from the cultures o~ microor-.
~3~77 ganisms grown by the submerged cultivation method, e.g~,~richoderma viride cult~re /c~. US Patent No. 3,542,580, cl.C 1~d 13~04 pub].ished ~ebrua.ry 15, 1972/. Canadian Pat-ent ~o.975,313, cl. 195-16 pu~lished on February 23, 1972, discloses a process for treating starch and protein-con-taining vegetable material with proteolytic enzymes, whereas ~rench Patent No-2,382,497, cl. C 12c 11/12 publish-ed on November 3, 1978 deals with a process for producing ethyl alcohol by virtue o~ cellulose fermenting under the e~ect o~ a miæed culture at a temperature of 50 to 65C
and the pH of 7 to 8. However, this patent fails to speci~y what producers are employed-It is an e~sential object of the present invention tos~arch for such an enzymic preparation ~hat would contain the required complex of h~drolytic enzymes capable o~ hydro-lyzing the nonam~laceous portion of the raw stock into fer-mentable sugars, thereby increasing the yield o~ ethyl alcohol.
~ he above object is acoomplished due to the provision o~ a novel enzymic preparation~ containing a complex of hydrolytic enzymes produced by a mould fungi ~richoderma koningii which is capable o~ more complete hydrolysis of amylaceous raw stock-According to the present invention, a process for pro-ucing ebhyl aloohol ~rom amylaceous raw stock by ibs hydro-,.
~4~ 7~
l~sis under the e~fect of am~lolytic enzymes is characteriz-ed by that hydrolysis is carried out in the presence o~ an enzymic preparation o~ cellulase obtained ~rom the culture of ~richoderma koningii, containing a complex o~ hydrolytic enzymes, viz., C1-enzyme? endo- and exoglucanase, cellobiase, x~lanase, J~ -glucosidase, protease, and a number o~ amylo-lytic enæymes-The principal e~zyme~ making part of the afore-mention-ed proparation ~eature the ~ollowing activity /in terms o~
units per gramJ:
C1-enzyme/with respect to paper~ _ 100 to 125 endoglucanase - 15 to 20 exoglucanase - 3 to 4 cellobiase - 4 to 6 ~ylanase - 200 to 250 Optimum consumption o~ the above preparation equals 1 to 2 per cent of the entire mass of the amylaceous raw stock invol~ed due to the ~act ~hat in the abovesaid consumption limits more complete and economieally advantageous h~dro-lysis is attained ~ he present invention enables one to carry out deeper hydrolysis o~ grain polysaceharides and increase the ~ield of ethyl alcohol by 3 to 4 per cent as compared to the known processes. ~his is accounted for by the fact that the provi-sion of such enzymes as C1-enz~me, endoglucanase, exogluca-nase, oellobiase, and others in the proposed preparation, 3~;77 are capable of hydrolyzing grain polysaccharides o~ the nonamylaceous nature, such as cellulose and hemicellulose, into ~ermentable carbohydrates, with the result that the yield of ethyl alcohol is increased and the amount of lost unfermented sugars is diminished-The above-mentioned enzymic p~eparation of cellulase lS obtai~ed ~rom culti~ating a mould fungus ~richoderma koningii on a solid nutrient medium, containing the follow-ing components /in terms of mass per cent/:
wheat bran - 40 to 45 beet pomace - 20 to 25 malt sprouts - ~5 to ~0 sawdust - 5 to 10 Culti~ation is carried out at 30 to ~5C ~or 48 to 55 hours. On terminating the cultivation process the re-sultant enzymic preparation can be made use of in alcohol production or as the mould culture together with the re-mainder of the nutrient medium and mycelium, disintegrated and dried to a moisture content of 12 or 1~ per cent; or else in the form o~ a preparation obtained by precipitating the enzymes ~rom aqueous extracts of the mould fungus cul-ture, b~ organic solvents-Application of unpurified enzgmic preparation in the form of the culture of the above mould fungus is economi.cal-ly more reasonable, as this rules out e~penditures for iso-lati~g -the enzymes and prevents losses of their activity in .
,.
, "
1~3tj77 the course o~ isolation~
~ he process of the present i.n~ention is carried out as Pollows. Am~laceous raw stock, 9 .g.1 wheat grains, is ~ed to the crusher to be disintegrated there, wherefrom the stock enters the mixer-pre~oiler composed of two compart-ments. In the first compartment the crushed grains is mixed with water in a weight ratio o~ 1 4, whereupon the mixture is forwarded to ~he second compartment, where it is heated to 80 or 85C. While the mixture is being heated starch is rendered pasteliXe- Next the pastelike mass is delivered from the preboiler to the cooker arrangement, wherein the mass is cooked at a pressure of 0-4 to 0.5 MPa for 30 to 50 min. ~hen the cooked mass is delivered to the steam separator for getting it read of steam, and from there to the 1st-stage saccharifier to be cooled to the temperature of hydrolysis /58 to 60C/. Fed to the sacchari~ier in an is amount of 30 per cent an aqueous suspension of a mixture of the hydrolytic enzymes obtained from surface cultures o~
mould fungi7 the above suspension containing 1 per cent -amylase from Aspergillus.oryzae /in terms of starch mass/, 4 per cent glucoamylase ~rom Aspergillus awamori Jin terms of the starch mass/, and.1~5 per cent o~ a comp-lex preparation of cellulase from the culture o~ ~richo-derma Xoningii /in terms of whole mass of raw stock~ ~he above aqueous suspension is treated with formalin in a 0-02 mass per cent concentration-~ ~3~i77 It is in -the 1st-stage saccharifier that the cooked mass is lique~ied and partially ~ydrolyzed under the effect o~ the aLore-mentioned enzymes, the process proceeding at 58 to 60C ~or 10 min. From the 1st-stage saccharifier the mass is transferred to the 2nd-stage saccharifier to which are added the remaining 70 per cent of the above aqueous suspension o~ the hydrolytic enzymes grown on surface cul-tures of mould fungi, whereupon starch and nonamylaceous polysaccharides undergo thorough hydroly~is in the 2nd-stage saccharifier at 57 to 58C for 2 to 5 minutes. Ne~t the finally h~drolyzed mass is transferred from the 2nd-stage saccharifier to the heat e~changer to be cooled to the fermentation temperature /30C/ and from there to the fermenter vat, into which is also introduced yeast ~accha~
romyces cerevisiae XII in an amount of 6 to 8 per cent of the fermenter ~at use~ul capacity. ~hereupon the sacchari-fied mass undergoes fermentatio~ under the effect of the above-mentioned yeast, the fermentation process occurring at 28 to 30C for 4B to 50 hours-The ~ermentation process over, the mature yeast washis fed to the rectifying still. A 4-per cent increase in the yield of ethyl alcohol per unit o~ the original amylace-ous stock is attained as compared to the ethyl alcohol pro-ducing process diispensing with the proposed preparation-~ he enzymic preparation of cellulose made use of forhydrolysis, is obtained by surface cultivatine o~ s mould .
3f~
_ 10 _ ~ungus Trichoderma koningii on a solid nutrient medium, containing the Eollowing components /in terms of mass per cen ~:
wheat bran - 45 malt sprouts - 25 beet pomace ~ 25 . sawdust - 5 . The moisture content of the nutrient medium is within ,'. 60 to 65 per cent, the cultivation occurring at 30 to 35C
~or 48 to 55 hours.
The finished culture o~ ~richoderma k'oningii fungus is in ~act an enz,ymic preparation of cellulase. Such'pre-~' paration contains a complex o:E hydrolytic enz~es, mostly cellulosoly~ic enz~mes and xylanase Eeaturing the following activit,y ~i~ terms o~ units per gram~::
~. C1-enz~me /with respect to paper/ - 100 to 125 . endoglucanase - 15 to 20 ;~ exoglucanase - ~ to 4 cellobiasa - 4 to 6 x 'xylanase _ 200 to 250 he thus-obtained enz~mic preparation o~ cellulase is ~; used for h,ydrol~sis in t~e form of a ~ungus culture "per ; sef~ without isolating and puri~ing with organic solvents, .~ an advantag~ous feature that rules out costly and sophis-ticated stage o~ isolating the enz~mes producing cellulase effect and pr0vents the loss of their ~ermenting activity ,;, .
.,, :
., .
...
,, ,,, : , -.
. . .
., ~3t;,~7 _ 11 _ while being isolated.
To promote understanding o~ the present invention given below are some examples o~ its practical embodiment carried out under laboratory conditions.
Fxample 1 50 g ground wheat grains is placed in a dry Erlenmeyer flask 0-5 1 in capacity, into which is added 200 ml water, whereupon the contents are stirred and rendered pastelike in a boiling water bath ~or 40 min. ~hen the ~lask lS re-moved from the bath and closed with glass covers, whereupon the ~lask is placed in a digester where its contents are cooked at a pressure of 0-15 MPa ~or 90 min. A~ter cooking the flask is removed ~rom the digester, and added thereto ~or the cooked mass to liquef~ is 30 per cent o~ a mixture o~ enzymes, aontainlng the following components /in terms o~ per cent o~ starch mass~ CC-am~lase from Aspergillus oryzae~, 1; glucoamylase ~rom Aspergillus awamori, 4; and the cellulase preparation ~rom ~richoderma k~oni~gii, 1.5 /in terms o~ mass o~ grains/, the above preparation con-taining a complex of hydrol~tic enzymes, i-e-, C1-enzyme, endoglucanase, exoglucanase, cellobiase, xylanass, B -clu-cosidase, protease 9 and a number of amylolytic enzymes. ~he mass is ¢ooled to 58C, and bhe rest o~ the mixture o~ en-zymes /70 per cent/ is added thereto, whereupon the mass is subjected to h~drol~sis at the above-mentioned tempera-ture ~or an hour-1~L43~77 _ 12 _After the h~drolysis the flask contents are cooled to 30C and doped with a suspension of yeast Saccharomyces cerevisiae XII /6 per cent of ~he mass/, whereupon some ~ormalin /0.02 per cent o~ the mas~/ is added ~or sterility.
Then the ~lask is closed and placed in a temperature-con-trolled cabinet for fermentation, which occurs at ~0G
within 72 hours. Next ethyl alcohol is distilled ~rom the resultant yeast wash in a distiller-~he yield o:~ ethyl alcohol equals 19.2~ ml from 50 g wheat containing 56.4 per cent starch, i.e. 104 per cent.
Used in Example 1 is the preparation of cellulase ~eaturing the following activity o~ the basal enzymes /in terms of units per gram/:
C1-enzyme /with respect ~o paper/ - 125 endoglucanase - 17 exoglucanase - 3 cellobiase _ 5 xylanase - ~19 ~he above preparation is obtained b~ cultivating a mould fungus ~richoderma koningii on a solid nutrient medi-um, containing the ~ollowing components /in terms o~ mass per cent/:
wheat bran - 45 malt sprouts - 25 beet pomace - 25 sawdust - 5 ,, ,.
,, 3f~77 The thus-obtained fungus culture is used in a native state 9 i.e., without isolating the enzymes from the nutri-ent medium and their puri~ying, which ~o a great extent simplifies production and utilization of the preparation-Exampla 2 Wheat grain~ /in an amount of 1000 k~ are continuous-ly fed to the grain crusher ~to be disintegrated there, whereupon the crushed grains are ~ed to the mixer-preboiler, composed o~ two compartments. Water is also fed to the 1st compartment in an amount four times the grain mass. Prelimi-nary heat treatment o~ grains occurs in the 1st compart-ment, whereas final heat treatment of grains, i.e., render-ing them pas~elike, is carried out in the 2nd compartment at 80 to 85C for 2 to 5 minutes. For ~inal heat treatment of the pastelake mass it is fed to the cooker arrangement to be coo~ed at a pressure of 0.5 MPa for 50 minutes.
Thereupon the mass is transferred to the steam separator for getting the mass read o~ steam, wherefrom the ~ass is forwarded to the 1st-stage saccharifier. On ¢ooling the mass to 58 or 60C addsd thereto is an aqueous suspension /30 per cent of the whole amount/ o~ a mixture of hydro-lytic enzymes o~ surface cultures of mould fungi, containing 1 per cent ~ -am~lase ~rom Aspergillus oryzae, 4 per cent glucoamylase from Aspergillus awamori /both in ~erms o~
starch mass/, and 1.5 per cent /in terms of preparation mass/ o~ the cellula~e preparation obtaine~ ~rom the culture ' , of a mould fungus Trichoderma koningii o~ the composition as in ~xample 1. Then the aqueous suspension is treated with an antiseptic, say, ~ormalin in a concentration of 0.02 mass per cent- The process o~ hydrolysis in the 1st-stage sacchari~ier takes 10 minutes, whereupon the mass is transferred to the 2nd~stage sacchariXier to which is added the remaining 70 per cent of the above aqueous suspension of the hydrolytic enzymes of surface cultures. It is in the ~-2nd-stage saccharifier that starch and nonamylaceous pol~-saccharides undergo thorough hydrol~sis at 57 to 58C ~or 5 minutes. Hydrolysis ovsr, the mass is let cool to 30C in a heat exchanger and trans~erred to the ~ermenter vat, to which is added yeast Saccharomyces cere~isiae XII /6 to 8 per cent o~ the fermenter use~ul capacity/, where the mass is ~ermented at 28 to 30C ~or 72 hours. ~he fermentation procèss over, the mature yeast wash is fed to the recti~-ing still to obtain ethyl alcohol there~rom- The ~ield of ethyl alcohol per ton o~ starch e~uals 66.56 dal~ i.e-, by 4 per cent higher as compared to the control procefis carried out according to the known method, that is5 without the pro-posed preparation. An increased ethyl alcohol yield is at-tained due to ~ermenting the carbohydratss resulting ~rom hydrol~sis o~ the nonam~laceous portion o~ corn grains-
Claims (2)
1. A process for producing ethyl alcohol from amylaceous raw stock by virtue of hydrolysis of said raw stock under the effect of amilolytic enzymes and an enzymic preparation of cellulase obtained from the culture of a mould fungus Trichoderma k?nigii and containing a complex of hydrolytic enzymes, viz., C1-enzyme, exo-glucanase, endoglucanase, cellobiase, xylanase, .beta.-glucosidase, protease, and a number of amylolytic enzymes.
2. A process as claimed in Claim 1, wherein said enzymic preparation is taken in an amount of 1 to 2 mass per cent.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CA000349907A CA1143677A (en) | 1980-04-15 | 1980-04-15 | Process for producing ethyl alcohol from amylaceous raw stock |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CA000349907A CA1143677A (en) | 1980-04-15 | 1980-04-15 | Process for producing ethyl alcohol from amylaceous raw stock |
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| Publication Number | Publication Date |
|---|---|
| CA1143677A true CA1143677A (en) | 1983-03-29 |
Family
ID=4116715
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA000349907A Expired CA1143677A (en) | 1980-04-15 | 1980-04-15 | Process for producing ethyl alcohol from amylaceous raw stock |
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| CA (1) | CA1143677A (en) |
Cited By (15)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4810647A (en) * | 1985-08-07 | 1989-03-07 | Valpi | Process of producing ethanol and various other by-products from cereals |
| WO1992020777A1 (en) * | 1991-05-17 | 1992-11-26 | Solvay Enzymes, Inc. | Process for producing ethanol |
| WO2007035730A2 (en) | 2005-09-20 | 2007-03-29 | Novozymes North America, Inc. | Process of producing a fermentation product |
| US7244597B2 (en) | 2000-11-10 | 2007-07-17 | Novozymes A/S | Secondary liquefaction in ethanol production |
| CN100564534C (en) * | 2002-02-08 | 2009-12-02 | 金克克国际有限公司 | Produce the alcoholic acid method with carbon substrate |
| WO2010009515A1 (en) * | 2008-07-25 | 2010-01-28 | Applimex Systems Pty Ltd | Enzyme hydrolysis method |
| US7820419B2 (en) | 2005-02-07 | 2010-10-26 | Novozymes North America Inc. | Fermentation product production processes |
| US20110124070A1 (en) * | 2008-05-29 | 2011-05-26 | Gang Duan | Process for alcohol and co-product production from grain sorghum |
| US8409640B2 (en) | 2003-03-10 | 2013-04-02 | Poet Research, Inc. | Methods and systems for producing ethanol using raw starch and fractionation |
| US8450094B1 (en) | 2009-03-03 | 2013-05-28 | Poet Research, Inc. | System for management of yeast to facilitate the production of ethanol |
| US8470550B2 (en) | 2003-03-10 | 2013-06-25 | Poet Research, Inc. | Composition comprising raw starch for the production of ethanol |
| US8597919B2 (en) | 2005-10-10 | 2013-12-03 | Poet Research, Inc. | Methods and systems for producing ethanol using raw starch and selecting plant material |
| US8815552B2 (en) | 2009-03-03 | 2014-08-26 | Poet Research, Inc. | System for fermentation of biomass for the production of ethanol |
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Cited By (23)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4810647A (en) * | 1985-08-07 | 1989-03-07 | Valpi | Process of producing ethanol and various other by-products from cereals |
| WO1992020777A1 (en) * | 1991-05-17 | 1992-11-26 | Solvay Enzymes, Inc. | Process for producing ethanol |
| US5231017A (en) * | 1991-05-17 | 1993-07-27 | Solvay Enzymes, Inc. | Process for producing ethanol |
| US7244597B2 (en) | 2000-11-10 | 2007-07-17 | Novozymes A/S | Secondary liquefaction in ethanol production |
| CN100564534C (en) * | 2002-02-08 | 2009-12-02 | 金克克国际有限公司 | Produce the alcoholic acid method with carbon substrate |
| US8409640B2 (en) | 2003-03-10 | 2013-04-02 | Poet Research, Inc. | Methods and systems for producing ethanol using raw starch and fractionation |
| US8409639B2 (en) | 2003-03-10 | 2013-04-02 | Poet Research, Inc. | Methods and systems for producing ethanol using raw starch and fractionation |
| US8679793B2 (en) | 2003-03-10 | 2014-03-25 | Poet Research, Inc. | Method for producing ethanol using raw starch |
| US8497082B2 (en) | 2003-03-10 | 2013-07-30 | Poet Research, Inc. | Composition comprising corn flour and saccharification enzymes |
| US8470550B2 (en) | 2003-03-10 | 2013-06-25 | Poet Research, Inc. | Composition comprising raw starch for the production of ethanol |
| US7820419B2 (en) | 2005-02-07 | 2010-10-26 | Novozymes North America Inc. | Fermentation product production processes |
| WO2007035730A2 (en) | 2005-09-20 | 2007-03-29 | Novozymes North America, Inc. | Process of producing a fermentation product |
| US8216817B2 (en) | 2005-09-20 | 2012-07-10 | Novozymes North America, Inc. | Process of producing a fermentation product |
| US8597919B2 (en) | 2005-10-10 | 2013-12-03 | Poet Research, Inc. | Methods and systems for producing ethanol using raw starch and selecting plant material |
| US20110124070A1 (en) * | 2008-05-29 | 2011-05-26 | Gang Duan | Process for alcohol and co-product production from grain sorghum |
| WO2010009515A1 (en) * | 2008-07-25 | 2010-01-28 | Applimex Systems Pty Ltd | Enzyme hydrolysis method |
| US8450094B1 (en) | 2009-03-03 | 2013-05-28 | Poet Research, Inc. | System for management of yeast to facilitate the production of ethanol |
| US8815552B2 (en) | 2009-03-03 | 2014-08-26 | Poet Research, Inc. | System for fermentation of biomass for the production of ethanol |
| US9234167B2 (en) | 2009-03-03 | 2016-01-12 | Poet Research, Inc. | System for management of yeast to facilitate the production of ethanol |
| US9416376B2 (en) | 2009-03-03 | 2016-08-16 | Poet Research, Inc. | System for management of yeast to facilitate the production of ethanol |
| US11987831B2 (en) | 2012-03-30 | 2024-05-21 | Novozymes A/S | Processes for producing a fermentation product |
| CN113403347A (en) * | 2021-08-05 | 2021-09-17 | 苏州迈博汇生物科技有限公司 | Alcohol production process with high alcohol yield |
| CN113403347B (en) * | 2021-08-05 | 2023-08-15 | 苏州迈博汇生物科技有限公司 | Alcohol production process with high alcohol yield |
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