Use of a combination of amylase, xylanase and yeast in alcohol fermentation with at least part of the raw material being wheat

Abstract

The present application relates to the field of alcohol fermentation, and particularly relates to application of a combination of amylase, xylanase and yeast in alcohol fermentation using wheat as at least part of raw materials, and further provides a method for mixed alcohol fermentation using wheat and cassava, which comprises the following steps: (1) contacting mixed powder slurry containing cassava and wheat with amylase and xylanase to liquefy the mixed powder slurry, and obtaining a liquefied liquid; (2) inoculating yeast into the liquefied liquid to perform alcohol fermentation. The present application can reduce the viscosity of the liquefied liquid, reduce the total residual sugar content after fermentation, reduce the viscosity of the fermentation liquid, and reduce the dry matter content of the waste mash liquid.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of alcohol fermentation, and in particular to the application of a combination of amylase, xylanase and yeast in alcohol fermentation using wheat as at least part of raw materials. BACKGROUND

[0002] The main raw materials for fuel ethanol fermentation include grain raw materials, potato raw materials, sugar raw materials and cellulose raw materials, among which the starch raw materials mainly include corn, wheat, rice and cassava, etc. With the rapid development of the fuel ethanol industry, the raw materials are gradually changing from single raw materials to mixed raw materials. In recent years, with the increase in grain output, the inventory of aged rice and wheat in China is huge, and a large amount of aged grain needs to be processed. The co-fermentation of multiple starch raw materials for the production of fuel ethanol can realize the diversification of raw materials and effectively solve the accumulation of aged grain.

[0003] Aged wheat is grain stored for a long time (more than 3 years), which is generally used for fuel ethanol production due to problems such as mold, mycotoxins and excessive heavy metals. The mixed use of wheat and cassava raw materials can achieve flexible production of diversified raw materials, avoid excessive dependence of enterprises on corn raw materials, effectively alleviate the problem of tight raw material supply, form a pattern of replacement of multiple raw materials, enhance the vitality and market competitiveness of enterprises, and promote the healthy and rapid development of enterprises. Due to the low price of wheat and cassava with excessive content, the production cost of enterprises can be reduced by optimizing the treatment, good economic benefits can be achieved, and the safety of grain can be ensured.

[0004] Since wheat is cheap, the use of mixed fermentation of cassava and wheat and the increase in the addition ratio of wheat helps to reduce the cost of ethanol production. However, the protein content in wheat is high, and the bran content is large, which leads to high viscosity of liquefaction, high total residual sugar at the end of fermentation, high viscosity of fermentation broth and high dry matter content of waste beer clear liquid. SUMMARY

[0005] The purpose of the present application is to overcome the above-mentioned problems existing in the prior art, and to provide the application of a combination of amylase, xylanase and saccharifying yeast in alcohol fermentation using wheat as at least part of raw materials. The present application can reduce the viscosity of liquefaction liquid, the total residual sugar content after fermentation, the viscosity of fermentation broth and the dry matter content of waste beer clear liquid.

[0006] In order to achieve the above-mentioned purpose, the first aspect of the present application provides the application of a combination of amylase, xylanase and yeast in reducing at least one of the viscosity of liquefaction liquid, the total residual sugar content after fermentation, the viscosity of fermentation broth and the dry matter content of waste beer clear liquid in the process of alcohol fermentation using wheat as at least part of alcohol fermentation raw materials.

[0007] The second aspect of the present application provides a method for mixed alcohol fermentation using wheat and cassava, which comprises:

[0008] (1) contacting a mixed powder slurry containing cassava and wheat with amylase and xylanase to liquefy the mixed powder slurry, to obtain a liquefied liquid;

[0009] (2) inoculating the liquefied liquid with yeast to perform alcohol fermentation.

[0010] The present application, on the basis of the original process, boldly tries various fermentation accessories, and finally optimizes a set of process suitable for alcohol fermentation using wheat as at least part of the alcohol fermentation raw material, especially the process of mixing cassava and wheat, i.e. using xylanase in the liquefaction stage, so that the viscosity of the fermentation liquid, the viscosity of the waste mash liquid, the total residual sugar, and the dry matter content of the waste mash clear liquid are all reduced. DETAILED DESCRIPTION

[0011] The endpoints of the ranges and any values disclosed herein are not limited to the precise values stated. The endpoints of the ranges and any values are understood to be approximate values. The endpoints of the ranges and any values are understood and are to be construed as specifically incorporating the language "about". For numeric values, the endpoints of the ranges, the endpoints of the ranges and individual point values, and individual point values can be combined with each other to generate one or more new numeric ranges, which are to be considered as specifically disclosed herein.

[0012] Unless otherwise indicated herein, temperatures are set forth in degrees Celsius.

[0013] In a first aspect, the present application provides the use of a combination of amylase, xylanase and yeast in the process of alcohol fermentation using wheat as at least part of the alcohol fermentation raw material to reduce at least one of the viscosity of the liquefied liquid, the total residual sugar content after fermentation, the viscosity of the fermentation liquid and the dry matter content of the waste mash clear liquid.

[0014] According to the present application, the alcohol fermentation raw material can be pure wheat, or a mixed raw material of wheat and other raw materials, which can be conventional raw materials for alcohol fermentation in the art, such as corn, cassava, etc.

[0015] According to the present application, the wheat is preferably aged wheat.

[0016] Unless otherwise specified herein, "aging" herein refers to the storage time of grain being more than 3 years.

[0017] According to the present application, the wheat can be hulled or unhulled wheat, preferably hulled but not peeled wheat.

[0018] According to the present invention, the xylanase can be any of the xylanases conventionally used in the art. In terms of temperature resistance, it can be a thermostable xylanase or a common xylanase, preferably a thermostable xylanase. The term "thermostable xylanase" refers to a xylanase with an enzyme activity of not less than 20,000 U / g at a temperature not lower than 70°C. In terms of acidity or alkalinity, it can be an acidic xylanase or a basic xylanase, preferably an acidic xylanase. According to a preferred embodiment of the present invention, the xylanase is a thermostable acidic xylanase, which is commercially available, for example, purchased from Xiasheng Industrial Group Co., Ltd., product number FDG-2234.

[0019] According to the present invention, preferably, to further improve the fermentation effect, the amylase contains thermostable α-amylase and protease, and the enzyme activity, calculated as α-amylase, is not less than 140,000 U / mL. The thermostable α-amylase refers to an α-amylase with an enzyme activity of not less than 140,000 U / mL at a temperature not lower than 70°C. The protease is preferably an acidic protease. According to a preferred embodiment of the present invention, the amylase is commercially available, specifically a super-strong compound amylase (Avantec CN) (containing thermostable amylase and acidic protease) purchased from Novozymes (China) Biotechnology Co., Ltd., with an enzyme activity of 140,000 U / mL.

[0020] Fermentation refers to the process by which microorganisms, under aerobic or anaerobic conditions, produce microbial cells, direct metabolites, or secondary metabolites. Fermentation can be carried out using methods conventionally used in the art. However, the inventors of this invention have discovered that using saccharifying yeast in conjunction with xylanase and amylase as described above during fermentation can reduce the total residual sugar content after fermentation, decrease the viscosity of the fermentation broth, reduce the dry matter content of the fermentation waste mash, and increase the alcohol content. Therefore, preferably, the yeast is saccharifying yeast, which is commercially available, for example, from Angel Yeast Co., Ltd., with a total yeast count ≥ 2 × 10⁻⁶. 10 per g.

[0021] Secondly, the present invention provides a method for mixed alcoholic fermentation of wheat and cassava, the method comprising:

[0022] (1) The mixed slurry containing cassava and wheat is contacted with amylase and xylanase to liquefy the mixed slurry and obtain a liquefied liquid;

[0023] (2) Inoculate the liquefied liquid with saccharified yeast for alcoholic fermentation.

[0024] According to the present invention, the cassava may be peeled or unpeeled, preferably unpeeled cassava.

[0025] According to the present application, the selection of the wheat has been described in detail in the first aspect above, which will not be repeated here.

[0026] According to the present application, the dry content of the mixed powder slurry can be a conventional content, preferably 30-40 wt%, for example, can be 30 wt%, 31 wt%, 32 wt%, 33 wt%, 34 wt%, 35 wt%, 36 wt%, 37 wt%, 38 wt%, 39 wt%, 40 wt%.

[0027] According to the present application, the pH value of the mixed powder slurry is preferably acidic, more preferably 5-6, for example, can be 5, 5.2, 5.4, 5.6, 5.8, 6.

[0028] Among them, the pH value of the mixed powder slurry can be adjusted by using conventional acid-base regulators, for example, dilute hydrochloric acid, dilute sulfuric acid, phosphoric acid, citric acid, ammonia, sodium hydroxide, etc.

[0029] According to the present application, the amount of wheat can be adjusted within a wide range, but the inventors of the present application have found that in order to further improve the fermentation performance of the present application, it is preferred that the blending ratio of wheat is 20-60% by dry weight, for example, can be 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%; more preferably 20-40%.

[0030] According to the present application, the preparation method of the mixed powder slurry comprises: mixing cassava crushing product and wheat crushing product mixture with mixing liquid to obtain the mixed powder slurry.

[0031] Among them, the mixing of the starch raw material with the mixing liquid to obtain the powder slurry is a step, and the mixing liquid can be various mixing liquids commonly used, for example, can include but not limited to tap water, process water, for example, waste water generated in the fermentation process to produce ethanol, such as condensate, clear liquid, etc.

[0032] Among them, the specific selection of the xylanase has been described in detail in the first aspect above, which will not be repeated here.

[0033] The amount of xylanase can be selected within a wide range, in order to further improve the fermentation performance of the present application, it is preferred that the amount of xylanase is 0.05-0.5 kg per ton of mixed powder slurry by dry weight, for example, can be 0.05 kg, 0.1 kg, 0.2 kg, 0.3 kg, 0.4 kg, 0.5 kg, preferably 0.2-0.4 kg.

[0034] Among them, the specific selection of the amylase has been described in detail in the first aspect above, which will not be repeated here.

[0035] The amount of the amylase can be selected in a wide range, and preferably, 0.1-0.5 kg of the amylase is used per ton of the mixed powder slurry in dry weight, for example, 0.1 kg, 0.2 kg, 0.3 kg, 0.4 kg, 0.5 kg, preferably 0.2-0.4 kg.

[0036] In the present application, liquefaction refers to a process of hydrolyzing the starchy raw material into small molecular substances (dextrin, oligosaccharide, etc.), and the liquefaction can be performed using a method commonly used in the art. However, in order to further improve the fermentation performance, preferably, the conditions of the liquefaction include: maintaining at a temperature of 80-100°C (for example, 80°C, 82°C, 84°C, 86°C, 88°C, 90°C, 92°C, 94°C, 96°C, 98°C, 100°C) for 1-3 h (for example, 1 h, 1.3 h, 1.5 h, 1.8 h, 2 h, 2.2 h, 2.5 h, 2.8 h, 3 h, 3.3 h, 3.5 h, 3.8 h, 4 h) to obtain the liquefied liquid.

[0037] Further preferably, the liquefaction is performed under stirring, and the stirring speed can be 150-250 rpm.

[0038] The method of the present application is particularly suitable for fermenting to prepare ethanol using unpeeled cassava and unpeeled and unhusked aged wheat as raw materials.

[0039] The specific selection of the yeast is described in detail in the first aspect above, and will not be repeated here.

[0040] According to the present application, the yeast can be commercially available, for example, saccharifying yeast available from Angel Yeast Co., Ltd., with a total number of yeast ≥ 2 x 10 10 individual / g.

[0041] According to the present application, preferably, the inoculation ratio of the saccharifying yeast is 0.05-0.2% by weight, more preferably 0.05-0.1% by weight, based on dry powder.

[0042] According to the present application, the fermentation conditions can be performed according to the conventional method in the art, but in order to further improve the fermentation performance of the present application, preferably, the present application is fermented by the following two stages:

[0043] The first stage fermentation is from the start of fermentation to 6-10 hours of fermentation, and the temperature of the first stage fermentation is 25-33°C;

[0044] The second stage fermentation is from 6-10 hours of fermentation to the end of fermentation, and the temperature of the second stage fermentation is 33-40°C.

[0045] The temperature of the first stage fermentation is lower than the temperature of the second stage fermentation, preferably 1-5℃, for example, 3℃.

[0046] According to the present application, the fermentation time can be 60-80 hours.

[0047] According to the present application, in order to further improve the fermentation performance of the present application, preferably, the method further comprises adding at least one of ammonium sulfate, saccharifying enzyme, acid protease and bactericide to the liquefied liquid.

[0048] Preferably, the method further comprises adding ammonium sulfate, saccharifying enzyme, acid protease and bactericide to the liquefied liquid.

[0049] The amount of the ammonium sulfate added is preferably 5-10 kg, more preferably 8-9 kg, per ton of the liquefied liquid in dry weight.

[0050] The amount of the saccharifying enzyme added is preferably 0.1-1 kg, more preferably 0.5-0.8 kg, per ton of the liquefied liquid in dry weight. It can be commercially available, for example, from Shandong Longda Biological Engineering Co., Ltd., with an enzyme activity of 260000 U / g. The definition of enzyme activity can be found in the instruction manual thereof.

[0051] The amount of the acid protease added is preferably 0.01-0.1 kg, more preferably 0.05-0.07 kg, per ton of the liquefied liquid in dry weight. It can be commercially available, for example, from Jeneke (China) Biological Engineering Co., Ltd., with an enzyme activity of 260000 U / g. The definition of enzyme activity can be found in the instruction manual thereof.

[0052] The amount of the bactericide added is preferably such that the final concentration thereof is 1-10 ppm, more preferably 4-6 ppm.

[0053] The bactericide can be any bactericide commonly used in the process of alcohol fermentation, for example, can include but is not limited to, Anquimycin, penicillin, Bacteriostatic Kang, Anquinnin, preferably, the bactericide is Anquimycin.

[0054] According to the present application, preferably, the method further comprises distilling the fermentation liquid obtained by fermentation to obtain alcohol and waste mash; and then performing solid-liquid separation on the waste mash to obtain clear liquid and wet grains.

[0055] The distillation refers to the step of separating ethanol from the fermentation mash, which can be separated by using the separation method commonly used in the art. The waste mash after distillation can be filtered by using plate and frame filter cloth to obtain clear liquid.

[0056] According to a particularly preferred embodiment of the present application, a method for mixed alcohol fermentation using cassava and wheat

[0057] (1) The cassava and aged wheat are crushed and mixed, wherein the wheat doping ratio is 30-40% by weight, then a mixing liquid (25-35% by weight of clear liquid, 65-75% by weight of process water) is added, so that the dry matter of the slurry is 31-33% by weight, and the pH value of the slurry is adjusted to 5-6.

[0058] (2) The obtained slurry is added with super compounded amylase (added amount is 0.3-0.4 kg / t of grain) and xylanase (added amount is 0.3-0.4 kg / t of grain), and heated to 88-92℃ for 2-3h, and the rotation speed is maintained at 150-250rpm, to obtain a liquefied mash.

[0059] (3) The obtained liquefied mash is added with ammonium sulfate (added amount is 8.2-8.6 kg / t of raw materials), saccharifying enzyme (added amount is 0.65-0.7 kg / t of raw materials), acid protease (added amount is 0.055-0.065 kg / t of raw materials), and anfungtai (final concentration is 5-5.5ppm) to prepare an ethanol fermentation medium, and the inoculation amount of dry saccharifying yeast powder is 0.05-0.1% by weight. From the start of fermentation to 7.5-8.5 hours of fermentation, the fermentation temperature is controlled at 30-32℃; from 8.5-9.5 hours of fermentation to the end of fermentation, the fermentation temperature is controlled at 33-35℃, and the total fermentation time is 68-72h, to obtain a fermentation mash.

[0060] (4) The fermentation mash obtained in step (3) is evaporated and condensed by a distillation device to obtain a distillation liquid containing alcohol and a waste mash liquid, and the waste mash liquid is filtered by a plate and frame cloth to obtain a clear liquid.

[0061] The present application will be described in detail below through examples.

[0062] The α-amylase is purchased from Genencor (China) Bioengineering Co., Ltd., and the enzyme activity is 140000 U / mL;

[0063] The super compounded amylase is purchased from Novozymes (China) Biotechnology Co., Ltd., and the enzyme activity is 140000 U / mL calculated based on α-amylase;

[0064] The xylanase is a high-temperature resistant acid xylanase, and is purchased from Xiasen Industrial Group Co., Ltd., with the product number FDG-2234, and the enzyme activity is 20000 U / g;

[0065] The saccharifying enzyme is purchased from Shandong Longda Bioengineering Co., Ltd., and the enzyme activity is 260000 U / g;

[0066] The protease is an acid protease, and is purchased from Genencor (China) Bioengineering Co., Ltd., and the enzyme activity is 260000 U / g;

[0067] Aged wheat is the aged wheat stored for 3 years, the starch content is 57.2% (w / w), and the moisture content is 12% (w / w);

[0068] Cassava is the newly purchased cassava powder, the starch content is 59% (w / w), and the moisture content is 13% (w / w);

[0069] Anjuntai is purchased from Liuzhou Longtai Technology Co., Ltd., and the bactericidal rate is 85% of the commercially available product;

[0070] The dry powder of fermenting yeast and the dry powder of saccharifying yeast are both purchased from Angel Yeast Co., Ltd., and the total number of yeasts is ≥2×10 10

[0071] Reducing sugar refers to the reducing sugar in the liquefied mash obtained after liquefaction, and the determination method is the Fehling reagent method;

[0072] Total sugar refers to the total sugar (including reducing sugar, disaccharide, starch, etc.) in the liquefied mash obtained after liquefaction, and the determination method is the Fehling reagent method;

[0073] DE value refers to the percentage of reducing sugar (calculated as glucose) in the dry matter of sugar syrup, and the determination method of reducing sugar and sugar syrup is the Fehling reagent method;

[0074] Residual reducing sugar refers to the residual reducing sugar in the fermentation mash obtained after the end of ethanol fermentation, and the determination method is the Fehling reagent method;

[0075] Residual total sugar refers to the residual sugar (including reducing sugar, disaccharide, starch, etc.) in the fermentation mash obtained after the end of ethanol fermentation, and the determination method is the Fehling reagent method;

[0076] Residual starch refers to the residual starch in the fermentation mash obtained after the end of ethanol fermentation, which is obtained by subtracting the total sugar after filtration from the total sugar;

[0077] Alcohol content refers to the degree of alcohol distilled from the fermentation mash obtained after the end of ethanol fermentation by a distillation device;

[0078] Acidity refers to the molar mass of NaOH consumed for neutralizing the acidic substances (including inorganic acid, organic acid, strong acid weak base salt, etc.) in per liter of liquefied mash, and the determination method is the acid-base neutralization method;

[0079] Waste mash liquid filtration rate refers to the rate of waste mash liquid passing through the plate and frame cloth filtration after distillation;

[0080] Filter dry matter refers to the solid content of the filtrate of the waste mash liquid after plate and frame cloth filtration, which is determined by a rapid moisture meter.

[0081] Process water is the water used for cooling in the alcohol process

[0082] ​The supernatant is supernatant after solid-liquid separation of waste mash after alcohol evaporation in the alcohol process, and COD in the supernatant is less than or equal to 60000 mg / L.

[0083] Example 1

[0084] This example is used to illustrate the method for mixed alcohol fermentation using cassava and wheat provided by the present application

[0085] (1) The cassava and aged wheat were crushed and mixed, wherein the wheat doping ratio was 30% by weight, then a mixing liquid (30% by weight of supernatant and 70% by weight of process water) was added, so that the dry matter of the powder slurry was 32% by weight, and the pH value of the powder slurry was adjusted to 5.6.

[0086] (2) The obtained powder slurry was added with super compounded amylase (added amount was 0.3 kg / t of grain) and xylanase (added amount was 0.3 kg / t of grain), maintained at 60℃ for 30 min, then the temperature was increased to 90℃ and maintained for 2 h, and the rotation speed was maintained at 200 rpm, to obtain a liquefied mash. The viscosity, reducing sugar, total sugar and DE value of the liquefied mash were determined, and the results are shown in Table 1.

[0087] (3) The obtained liquefied mash was added with ammonium sulfate (added amount was 8.4 kg / t of raw materials), saccharifying enzyme (added amount was 0.68 kg / t of raw materials), acid protease (added amount was 0.06 kg / t of raw materials) and anfungtai (final concentration was 5 ppm) to prepare an ethanol fermentation medium, and the inoculation amount of dry saccharifying yeast powder was 0.05% by weight. From the start of fermentation to the 8th hour of fermentation, the fermentation temperature was controlled at 30℃; from the 9th hour of fermentation to the end of fermentation, the fermentation temperature was controlled at 33℃, and the total fermentation time was 70 h, to obtain a fermented mash. The residual reducing sugar, residual total sugar, residual starch and viscosity in the fermented mash were determined, and the results are shown in Table 2.

[0088] (4) The obtained fermented mash in step (3) was evaporated and condensed by a distillation device to obtain a distillation liquid containing alcohol and a waste mash liquid, and the waste mash liquid was filtered by a plate and frame cloth to obtain a supernatant. The alcohol content, acidity, filtration speed of the waste mash liquid and dry matter of the supernatant were determined, and the results are shown in Table 2.

[0089] Example 2

[0090] This example is used to illustrate the method for mixed alcohol fermentation using cassava and wheat provided by the present application

[0091] (1) The cassava and aged wheat were crushed and mixed, wherein the wheat doping ratio was 20% by weight, then a mixing liquid (30% by weight of supernatant and 70% by weight of process water) was added, so that the dry matter of the powder slurry was 34% by weight, and the pH value of the powder slurry was adjusted to 5.

[0092] (2) To the obtained powder slurry, super strong compound amylase (0.4 kg / t of grains) and xylanase (0.4 kg / t of grains) were added, and the temperature was maintained at 55°C for 40 min, and then the temperature was raised to 95°C for 1.5 h, and the rotation speed was maintained at 150 rpm to obtain a liquefied mash. The viscosity, reducing sugar, total sugar and DE value of the liquefied mash were determined, and the results are shown in Table 1.

[0093] (3) To the obtained liquefied mash, ammonium sulfate (8 kg / t of raw materials), saccharifying enzyme (0.8 kg / t of raw materials), acid protease (0.05 kg / t of raw materials) and Anjuntai (6 ppm) were added to prepare an ethanol fermentation medium, and the inoculation amount of dry saccharifying yeast powder was 0.08% by weight. From the start of fermentation to the 7th hour of fermentation, the fermentation temperature was controlled at 28°C, and from the 8th hour of fermentation to the end of fermentation, the fermentation temperature was controlled at 33°C, and the total fermentation time was 70 h to obtain a fermented mash. The residual reducing sugar, residual total sugar, residual starch and viscosity in the fermented mash were determined, and the results are shown in Table 2.

[0094] (4) The fermented mash obtained in step (3) was evaporated and condensed by a distillation device to obtain a distillation liquid containing wine and a waste mash liquid, and the waste mash liquid was filtered by a plate and frame cloth to obtain a clear liquid. The alcohol content, acidity, filtration speed of the waste mash liquid and dry matter of the clear liquid were determined, and the results are shown in Table 2.

[0095] Example 3

[0096] This example is used to illustrate the method for mixed alcohol fermentation using cassava and wheat provided by the present application

[0097] (1) Cassava and aged wheat were crushed and mixed, wherein the wheat adulteration ratio was 40% by weight, and then a mixing liquid (30% by weight of clear liquid and 70% by weight of process water) was added so that the dry matter of the powder slurry was 36% by weight, and the pH value of the powder slurry was adjusted to 6.

[0098] (2) To the obtained powder slurry, super strong compound amylase (0.25 kg / t of grains) and xylanase (0.25 kg / t of grains) were added, and the temperature was maintained at 65°C for 20 min, and then the temperature was raised to 80°C for 3 h, and the rotation speed was maintained at 250 rpm to obtain a liquefied mash. The viscosity, reducing sugar, total sugar and DE value of the liquefied mash were determined, and the results are shown in Table 1.

[0099] (3) The obtained liquefied mash was added with ammonium sulfate (added amount 9 kg / t raw material), saccharifying enzyme (added amount 0.5 kg / t raw material), acid protease (added amount 0.07 kg / t raw material), and Anfengtai (final concentration 4 ppm) to prepare an ethanol fermentation medium, and the fermentation was inoculated with 0.1 wt% of dry saccharifying yeast. The fermentation temperature was controlled at 32°C from the start of the fermentation to the 9th hour of the fermentation, and the fermentation temperature was controlled at 34°C from the 10th hour of the fermentation to the end of the fermentation, and the total fermentation time was 70 h, to obtain a fermentation mash. The residual reducing sugar, residual total sugar, residual starch, and viscosity in the fermentation mash were determined, and the results are shown in Table 2.

[0100] (4) The fermentation mash obtained in step (3) was evaporated and condensed by a distillation device to obtain a distillation liquid containing alcohol and a waste mash liquid, and the waste mash liquid was filtered by a plate and frame cloth filter to obtain a clear liquid. The alcohol content, acidity, filtration rate of the waste mash liquid, and dry matter of the clear liquid were determined, and the results are shown in Table 2.

[0101] Example 4

[0102] This example is used to illustrate the method for mixed alcohol fermentation using cassava and wheat provided by the present application

[0103] The fermentation was carried out according to the method of Example 1, except that the liquefaction did not include the step of maintaining at 60°C for 30 min, but directly maintained at 90°C for 2.5 h, and the results are shown in Tables 1 and 2.

[0104] Example 5

[0105] This example is used to illustrate the method for mixed alcohol fermentation using cassava and wheat provided by the present application

[0106] The fermentation was carried out according to the method of Example 1, except that in step (2), the super compounded amylase was replaced with an equal amount of α-amylase (Genencor amylase was purchased from Genencor (China) Bioengineering Co., Ltd., with an enzyme activity of 140000 U / mL), and no xylanase was added; and in step (3), the saccharifying yeast was replaced with an equal amount of fermenting yeast. The results are shown in Tables 1 and 2.

[0107] Comparative Example 1

[0108] This comparative example is used to illustrate a reference method for mixed alcohol fermentation using cassava and wheat

[0109] The fermentation was carried out according to the method of Example 1, except that in step (2), the super compounded amylase was replaced with an equal amount of α-amylase (Genencor amylase was purchased from Genencor (China) Bioengineering Co., Ltd., with an enzyme activity of 140000 U / mL), and no xylanase was added; and in step (3), the saccharifying yeast was replaced with an equal amount of fermenting yeast. The results are shown in Tables 1 and 2.

[0110] Comparative Example 2

[0111] This comparative example is used to illustrate a method for mixed alcoholic fermentation using cassava and wheat as a reference.

[0112] Fermentation was carried out according to the method of Example 1, except that xylanase was not added in step (2); and in step (3), the saccharifying yeast was replaced with an equal amount of Ferment Lysate. The results are shown in Tables 1 and 2.

[0113] Comparative Example 3

[0114] This comparative example is used to illustrate a method for mixed alcoholic fermentation using cassava and wheat as a reference.

[0115] Fermentation was carried out according to the method of Example 1, except that in step (2), the super-strong compound amylase was replaced with an equal amount of α-amylase (Genentech amylase was purchased from Genentech (China) Biotechnology Co., Ltd., with an enzyme activity of 140,000 U / mL), and xylanase was not added. The results are shown in Tables 1 and 2.

[0116] Comparative Example 4

[0117] This comparative example is used to illustrate a method of mixed alcoholic fermentation using wheat as a reference.

[0118] Fermentation was carried out using the same method as in Comparative Example 1, except that 100% wheat was used instead of cassava. The results are shown in Tables 1 and 2.

[0119] Comparative Example 5

[0120] This comparative example is used to illustrate the method of mixed alcoholic fermentation using cassava as a reference.

[0121] Fermentation was carried out using the method described in Comparative Example 1, except that 100% cassava was used instead of aged wheat. The results are shown in Tables 1 and 2.

[0122] Table 1

[0123] Viscosity (mPa.s) Reducing sugars (%) Total sugars (%) DE value Example 1 125.09 9.21 24.70 28.78 Example 2 137.00 9.80 26.30 28.82 Example 3 160.64 9.98 27.50 27.72 Example 4 120.77 9.39 24.72 29.30 Example 5 133.10 8.61 24.65 26.90 Comparative Example 1 341.48 8.53 24.67 26.67 Comparative Example 2 301.79 9.15 24.71 28.59 Comparative Example 3 341.48 8.53 24.67 26.67 Comparative Example 4 1022.01 8.32 24.00 26.00 Comparative Example 5 201.05 8.70 25.10 27.19

[0124] Table 2

[0125]

[0126] As can be seen from the results in Tables 1 and 2, the addition of xylanase in this invention reduces the viscosity of the liquefied liquid, ultimately resulting in a decrease in the viscosity of the fermentation liquid, total residual sugar, and dry matter content of the waste mash. The preferred combination of "using xylanase and amylase with an enzyme activity of not less than 140,000 U / mL (calculated as α-amylase) during the liquefaction stage, and using saccharifying yeast during the fermentation stage" further reduces the viscosity of the liquefied liquid and fermentation liquid, total residual sugar, and dry matter content of the waste mash, while significantly increasing the alcohol content.

[0127] The preferred embodiments of the present application are described in detail above, but the present application is not limited thereto. Various simple modifications can be made to the technical solutions of the present application within the technical concept of the present application, including the combination of various technical features in any other suitable manner, and these simple modifications and combinations should also be considered as disclosed by the present application and fall within the protection scope of the present application.

Claims

1. A method for reducing the viscosity of liquefied liquid, the total residual sugar content after fermentation, the viscosity of fermentation broth, and the dry matter content of waste mash in mixed alcoholic fermentation of wheat and cassava, characterized in that, The method includes: (1) The slurry of cassava and wheat is contacted with amylase and xylanase to liquefy the slurry and obtain a liquefied liquid. (2) Inoculate yeast into the liquefied liquid for alcoholic fermentation; The amylase is a thermostable α-amylase and a protease, and the enzyme activity, calculated as α-amylase, is not less than 140,000 U / mL. The yeast in question is a saccharified yeast; In step (1), the liquefaction conditions include maintaining the temperature at 80-100℃ for 1-4 hours to obtain the liquefied liquid; The fermentation conditions include: the first stage of fermentation, from the start of fermentation to 6-10 hours, is at a temperature of 25-33℃; the second stage of fermentation, from 6-10 hours to the end of fermentation, is at a temperature of 33-40℃. The temperature of the first stage of fermentation is 1-5°C lower than that of the second stage of fermentation.

2. The method according to claim 1, wherein, In step (1), the dry matter content of the mixed slurry is 30-40% by weight; and / or The pH value of the mixed slurry is 5-6; and / or The blending ratio of wheat is 20-60% by dry weight; and / or The wheat in question is aged wheat.

3. The method according to claim 1 or 2, wherein, In step (1), the amount of xylanase used is 0.05-0.5 kg per ton of mixed slurry on a dry weight basis; and / or The dosage of amylase is 0.1-0.5 kg per ton of mixed slurry by dry weight.

4. The method according to claim 1 or 2, wherein, In step (2), the amount of yeast inoculated as dry powder is 0.05-0.2g relative to 100g of the liquefied liquid.

5. The method according to claim 1 or 2, wherein, The method further includes adding at least one of ammonium sulfate, saccharifying enzyme, acidic protease, and bactericide to the liquefied liquid.

6. The method according to claim 1 or 2, wherein, The method also includes distilling the fermentation broth obtained from fermentation to obtain alcohol and waste mash; then separating the solid and liquid components of the waste mash to obtain clear liquid and wet lees.

7. A method for reducing the viscosity, total residual sugar, and dry matter content of liquefied liquid and fermentation broth in mixed alcoholic fermentation using cassava and wheat, characterized in that, The method includes: (1) Crush and mix cassava with aged wheat, wherein the wheat content is 30-40% by weight, then add mixing liquid to make the dry matter of the slurry 31-33% by weight, and adjust the pH of the slurry to 5-6. The mixing liquid comprises 25-35% by weight of clear liquid and 65-75% by weight of process water; (2) Add super-strong compound amylase and xylanase to the obtained slurry, heat to 88-92℃ and maintain for 2-3 hours, and maintain the rotation speed at 150-250 rpm to obtain liquefied liquid; The amount of super-strong compound amylase added is 0.3-0.4 kg per ton of mixed slurry on a dry weight basis, and the amount of xylanase added is 0.3-0.4 kg. (3) Add ammonium sulfate, saccharifying enzyme, acidic protease, and anthraquinone to the obtained liquefied liquid to prepare an ethanol fermentation medium, and inoculate with saccharifying yeast; control the fermentation temperature at 30-32℃ from the start of fermentation to 7.5-8.5 hours of fermentation; control the fermentation temperature at 33-35℃ from 8.5-9.5 hours of fermentation to the end of fermentation, and the total fermentation time is 68-72 hours to obtain fermented mash; Of which, relative to each ton of liquefied liquid by dry weight, the amount of ammonium sulfate added is 8.2-8.6 kg, the amount of saccharifying enzyme added is 0.65-0.7 kg, and the amount of acidic protease added is 0.055-0.065 kg; The amount of Anjuntai added is such that its final concentration is 5-5.5 ppm; The amount of saccharified yeast inoculated as dry powder was 0.05-0.1% by weight. (4) The obtained fermented mash is evaporated and condensed by a distillation device to obtain alcoholic distillate and waste mash. The waste mash is filtered through a plate and frame cloth to obtain clear liquid.

Images