CN101591682A - Processing method with preparing pyruvate by biological fermentation method - Google Patents
Processing method with preparing pyruvate by biological fermentation method Download PDFInfo
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Abstract
The present invention relates to a kind of processing method with preparing pyruvate by biological fermentation method.It is characterized in that mainly comprising following processing step: fermention medium batching, fermented bacterium cultivation, fermentation, deactivation, after precipitation, filter, sterilization promptly gets pyruvate salt solution.The present invention utilizes the production of pyruvic acid by biological fermentation method product salt, avoids directly using pyruvic acid in the production process, has saved production cost greatly, in the process that adopts production of pyruvic acid by biological fermentation method salt, do not have pollutent to discharge, do not pollute, belong to green production process for environment.Compare with chemical synthesis, the advantage that biological fermentation process provided by the present invention has that raw materials cost is low, wide material sources, production product purity height, quality are good, produce the reaction conditions gentleness is to have the market competitiveness, the good processing method of development prospect at present.
Description
Technical field:
The present invention relates to a kind of processing method with preparing pyruvate by biological fermentation method.
Background technology:
Pyruvic acid and salt is in the existing certain production history of China, but mostly is to adopt the method for chemosynthesis to produce the pyruvic acid of a spot of SILVER REAGENT, industrial raw material and salt, production technique backwardness at present.As: domestic CALCIUM PYRUVIC by pyruvic acid and calcium ion in conjunction with generation, main technique forms through high temperature, evaporation, crystallization with pyruvic acid and milk of lime.Producing of pyruvate salt will be raw material with the pyruvic acid all generally, so the preparation of pyruvic acid is very crucial.The production method of raw material pyruvic acid generally adopts chemical synthesis, chemical synthesis produces pyruvic acid cost height, it is low, seriously polluted to produce effects, be difficult to reach country to environment requirement, simultaneously, pyruvate salt is mostly as the raw material of food or series cosmetics product, directly utilized by human body, the security of the pyruvic acid that chemical synthesis is produced remains to be considered.On the other hand, the pyruvic acid international price is higher, and China is from external import in the employed pyruvate salt series products and the raw material overwhelming majority aspect biological medicine, the fat-reducing, has improved the production cost of pyruvate salt series products.
Summary of the invention:
Technical problem to be solved by this invention be to provide a kind of low cost of manufacture, quality product height, can effective antipollution processing method with preparing pyruvate by biological fermentation method.
A kind of processing method with preparing pyruvate by biological fermentation method is characterized in that mainly comprising following processing step:
(1) fermention medium batching:
In the water of every 1000mL, put into glucose 90g～120g, peptone 3～15g, lime carbonate or sodium hydroxide or sodium bicarbonate 30g～40g, with the above-mentioned raw materials thorough mixing, the pH value of mixture is 5.0～5.6, after carrying out sterilization in 10 minutes～15 minutes under 108 ℃～120 ℃ temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
False pompon pseudoyeast or torulopsis glabrata are carried out the seed enlarged culturing after cultivating 16～24 hours under 30～32 ℃, spawn culture 20～24 hours when bacterial classification concentration weight ratio reaches 0.8%～1.2%, changes over to and carries out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 28 ℃～32 ℃ in the fermenting process, pressure is 0.03Mpa～0.05Mpa, time is 48～56 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains pyruvate salt and bacterial classification is continued deactivation 10 minutes～20 minutes under 80 ℃～95 ℃ temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get pyruvate salt solution.
The preservation of above-mentioned false pompon pseudoyeast, Torulopsis glabrata kind: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in 0.5～1 year is upgraded once.
The activation of above-mentioned bacterial classification: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, cultivated 16～24 hours down at 30 ℃～32 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone is 10～15g; Glucose is 20～30g; Agar is 15～20g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.0～5.6.
The composition of liquid seed culture medium: peptone is 10～15g; Glucose is 20～30g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.0～5.6.
The present invention utilizes the production of pyruvic acid by biological fermentation method product salt, avoids directly using pyruvic acid in the production process, has saved production cost greatly, in the process that adopts production of pyruvic acid by biological fermentation method salt, do not have pollutent to discharge, do not pollute, belong to green production process for environment.Compare with chemical synthesis, the advantage that biological fermentation process provided by the present invention has that raw materials cost is low, wide material sources, production product purity height, quality are good, produce the reaction conditions gentleness is to have the market competitiveness, the good processing method of development prospect at present.
Embodiment:
Embodiment 1:
Present embodiment is produced the processing method of CALCIUM PYRUVIC with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into glucose 90g, peptone 3g, lime carbonate 30g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.0, after carrying out sterilization in 10 minutes under 108 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
False pompon pseudoyeast bacterial classification is carried out shake-flask culture, cultivated 16 hours down at 30 ℃, entering seeding tank when reaching the maximum growth logarithmic phase of bacterial classification carries out the seed enlarged culturing, and it is 20 little that bacterial classification is cultivated in seeding tank
When bacterial classification concentration reaches 0.8%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 28 ℃ in the fermenting process, pressure is 0.03Mpa, time is 48 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains CALCIUM PYRUVIC and bacterial classification is continued deactivation 10 minutes under 80 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get CALCIUM PYRUVIC solution.
The preservation of above-mentioned false pompon pseudoyeast bacterial classification: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, half a year, activation was upgraded once.
The activation of above-mentioned false pompon pseudoyeast bacterial classification: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 16 hours down at 30 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone is 10g; Glucose is 20g; Agar is 15g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.0.
The composition of liquid seed culture medium: peptone is 10g; Glucose is 20g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.0.
Embodiment 2:
Present embodiment is produced the processing method of CALCIUM PYRUVIC with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into glucose 95g, peptone 7g, lime carbonate 33g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.1, after carrying out sterilization in 11 minutes under 110 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
False pompon pseudoyeast bacterial classification is carried out shake-flask culture, cultivated 18 hours down at 30.5 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 21 hours, when bacterial classification concentration reaches 0.9%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 29 ℃ in the fermenting process, pressure is 0.035Mpa, time is 49 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains CALCIUM PYRUVIC and bacterial classification is continued deactivation 12 minutes under 82 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get CALCIUM PYRUVIC solution.
The preservation of above-mentioned false pompon pseudoyeast bacterial classification: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in eight months is upgraded once.
The activation of above-mentioned false pompon pseudoyeast bacterial classification: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 17 hours down at 30.5 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone is 11g; Glucose is 22g; Agar is 16g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.2.
The composition of liquid seed culture medium: peptone is 11g; Glucose is 22g; Distilled water is 1000mL, and the pH value of above-mentioned substratum is 5.2.
Embodiment 3:
Present embodiment is produced the processing method of CALCIUM PYRUVIC with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into glucose 100g, peptone 9g, lime carbonate 35g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.2, after carrying out sterilization in 12 minutes under 113 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
False pompon pseudoyeast bacterial classification is carried out shake-flask culture, cultivated 20 hours down at 31 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 22 hours, when bacterial classification concentration reaches 1.0%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 30 ℃ in the fermenting process, pressure is 0.04Mpa, time is 51 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains CALCIUM PYRUVIC and bacterial classification is continued deactivation 14 minutes under 84 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get CALCIUM PYRUVIC solution.
The preservation of above-mentioned false pompon pseudoyeast bacterial classification: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in nine months is upgraded once.
The activation of above-mentioned false pompon pseudoyeast bacterial classification: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 20 hours down at 31 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone is 12g; Glucose is 24g; Agar is 17g; Distilled water: 1000mL, the pH value of above-mentioned substratum are 5.3.
The composition of liquid seed culture medium: peptone is 12g; Glucose is 24g; Distilled water is 1000mL, and the pH value of above-mentioned substratum is 5.3.
Embodiment 4:
Present embodiment is produced the processing method of CALCIUM PYRUVIC with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into glucose 110g, peptone 12g, lime carbonate 38g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.4, after carrying out sterilization in 14 minutes under 118 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
False pompon pseudoyeast bacterial classification is carried out shake-flask culture, cultivated 22 hours down at 31.5 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 23 hours, when bacterial classification concentration reaches 1.1%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 31 ℃ in the fermenting process, pressure is 0.045Mpa, time is 54 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains CALCIUM PYRUVIC and bacterial classification is continued deactivation 18 minutes under 90 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get CALCIUM PYRUVIC solution.
The preservation of above-mentioned false pompon pseudoyeast bacterial classification: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in ten months is upgraded once.
The activation of above-mentioned false pompon pseudoyeast bacterial classification: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 22 hours down at 31.5 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone: 14g; Glucose: 28g; Agar: 19g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.5.
The composition of liquid seed culture medium: peptone: 14g; Glucose: 28g; Distilled water: 1000mL, the pH value of above-mentioned substratum are 5.5.
Embodiment 5:
Present embodiment is produced the processing method of CALCIUM PYRUVIC with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into glucose 120g, peptone 15g, lime carbonate 40g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.6, after carrying out sterilization in 15 minutes under 120 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
False pompon pseudoyeast bacterial classification is carried out shake-flask culture, cultivated 24 hours down at 32 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 24 hours, when bacterial classification concentration reaches 1.2%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 32 ℃ in the fermenting process, pressure is 0.05Mpa, time is 56 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains CALCIUM PYRUVIC and bacterial classification is continued deactivation 20 minutes under 95 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get CALCIUM PYRUVIC solution.
The preservation of above-mentioned false pompon pseudoyeast bacterial classification: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in 1 year is upgraded once.
The activation of above-mentioned false pompon pseudoyeast bacterial classification: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 24 hours down at 32 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone: 15g; Glucose: 30g; Agar: 20g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.6.
The composition of liquid seed culture medium: peptone: 15g; Glucose: 30g; Distilled water 1000mL, the pH value of above-mentioned substratum is: 5.6.
Embodiment 6:
Present embodiment is produced the processing method of CALCIUM PYRUVIC with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 90g, food-grade albumen peptone 3g, food grade lime carbonate 30g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.0, after carrying out sterilization in 10 minutes under 108 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
The Torulopsis glabrata kind is carried out shake-flask culture, cultivated 16 hours down at 30 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 20 hours, when bacterial classification concentration reaches 0.8%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 28 ℃ in the fermenting process, pressure is 0.03Mpa, time is 48 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains CALCIUM PYRUVIC and bacterial classification is continued deactivation 10 minutes under 80 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get CALCIUM PYRUVIC solution.
The preservation of above-mentioned Torulopsis glabrata kind: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, half a year, activation was upgraded once.
The activation of above-mentioned Torulopsis glabrata kind: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 16 hours down at 30 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone is 10g; Glucose is 20g; Agar is 15g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.0.
The composition of liquid seed culture medium: peptone is 10g; Glucose is 20g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.0.
Embodiment 7:
Present embodiment is produced the processing method of CALCIUM PYRUVIC with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 95g, food-grade albumen peptone 7g, food grade lime carbonate 33g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.1, after carrying out sterilization in 11 minutes under 110 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
The Torulopsis glabrata kind is carried out shake-flask culture, cultivated 18 hours down at 30.5 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 21 hours, when bacterial classification concentration reaches 0.9%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 29 ℃ in the fermenting process, pressure is 0.035Mpa, time is 49 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains CALCIUM PYRUVIC and bacterial classification is continued deactivation 12 minutes under 82 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get CALCIUM PYRUVIC solution.
The preservation of above-mentioned Torulopsis glabrata kind: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in eight months is upgraded once.
The activation of above-mentioned Torulopsis glabrata kind: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 17 hours down at 30.5 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone is 11g; Glucose is 22g; Agar is 16g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.2.
The composition of liquid seed culture medium: peptone is 11g; Glucose is 22g; Distilled water is 1000mL, and the pH value of above-mentioned substratum is 5.2.
Embodiment 8:
Present embodiment is produced the processing method of CALCIUM PYRUVIC with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 100g, food-grade albumen peptone 9g, food grade lime carbonate 35g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.2, after carrying out sterilization in 12 minutes under 113 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
The Torulopsis glabrata kind is carried out shake-flask culture, cultivated 20 hours down at 31 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 22 hours, when bacterial classification concentration reaches 1.0%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 30 ℃ in the fermenting process, pressure is 0.04Mpa, time is 51 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains CALCIUM PYRUVIC and bacterial classification is continued deactivation 14 minutes under 84 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get CALCIUM PYRUVIC solution.
The preservation of above-mentioned Torulopsis glabrata kind: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in nine months is upgraded once.
The activation of above-mentioned Torulopsis glabrata kind: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 20 hours down at 31 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone is 12g; Glucose is 24g; Agar is 17g; Distilled water: 1000mL, the pH value of above-mentioned substratum are 5.3.
The composition of liquid seed culture medium: peptone is 12g; Glucose is 24g; Distilled water is 1000mL, and the pH value of above-mentioned substratum is 5.3.
Embodiment 9:
Present embodiment is produced the processing method of CALCIUM PYRUVIC with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 110g, food-grade albumen peptone 12g, food grade lime carbonate 38g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.4, after carrying out sterilization in 14 minutes under 118 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
The Torulopsis glabrata kind is carried out shake-flask culture, cultivated 22 hours down at 31.5 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 23 hours, when bacterial classification concentration reaches 1.1%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 31 ℃ in the fermenting process, pressure is 0.045Mpa, time is 54 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains CALCIUM PYRUVIC and bacterial classification is continued deactivation 18 minutes under 90 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get CALCIUM PYRUVIC solution.
The preservation of above-mentioned Torulopsis glabrata kind: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in ten months is upgraded once.
The activation of above-mentioned Torulopsis glabrata kind: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 22 hours down at 31.5 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone: 14g; Glucose: 28g; Agar: 19g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.5.
The composition of liquid seed culture medium: peptone: 14g; Glucose: 28g; Distilled water: 1000mL, the pH value of above-mentioned substratum are 5.5.
Embodiment 10:
Present embodiment is produced the processing method of CALCIUM PYRUVIC with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 120g, food-grade albumen peptone 15g, food grade lime carbonate 40g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.6, after carrying out sterilization in 15 minutes under 120 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
The Torulopsis glabrata kind is carried out shake-flask culture, cultivated 24 hours down at 32 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 24 hours, when bacterial classification concentration reaches 1.2%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 32 ℃ in the fermenting process, pressure is 0.05Mpa, time is 56 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains CALCIUM PYRUVIC and bacterial classification is continued deactivation 20 minutes under 95 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get CALCIUM PYRUVIC solution.
The preservation of above-mentioned Torulopsis glabrata kind: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in 1 year is upgraded once.
The activation of above-mentioned Torulopsis glabrata kind: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 24 hours down at 32 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone: 15g; Glucose: 30g; Agar: 20g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.6.
The composition of liquid seed culture medium: peptone: 15g; Glucose: 30g; Distilled water 1000mL, the pH value of above-mentioned substratum is: 5.6.
Embodiment 11:
Present embodiment is produced the processing method of Sodium.alpha.-ketopropionate with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 90g, food-grade albumen peptone 3g, sodium hydroxide 30g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.0, after carrying out sterilization in 10 minutes under 108 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
False pompon pseudoyeast bacterial classification is carried out shake-flask culture, cultivated 16 hours down at 30 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 20 hours, when bacterial classification concentration reaches 0.8%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 28 ℃ in the fermenting process, pressure is 0.03Mpa, time is 48 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains Sodium.alpha.-ketopropionate and bacterial classification is continued deactivation 10 minutes under 80 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get Sodium.alpha.-ketopropionate solution.
The preservation of above-mentioned false pompon pseudoyeast bacterial classification: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, half a year, activation was upgraded once.
The activation of above-mentioned false pompon pseudoyeast bacterial classification: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 16 hours down at 30 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone is 10g; Glucose is 20g; Agar is 15g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.0.
The composition of liquid seed culture medium: peptone is 10g; Glucose is 20g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.0.
Embodiment 12:
Present embodiment is produced the processing method of Sodium.alpha.-ketopropionate with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 95g, food-grade albumen peptone 7g, sodium hydroxide 33g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.1, after carrying out sterilization in 11 minutes under 110 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
False pompon pseudoyeast bacterial classification is carried out shake-flask culture, cultivated 18 hours down at 30.5 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 21 hours, when bacterial classification concentration reaches 0.9%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 29 ℃ in the fermenting process, pressure is 0.035Mpa, time is 49 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains Sodium.alpha.-ketopropionate and bacterial classification is continued deactivation 12 minutes under 82 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get Sodium.alpha.-ketopropionate solution.
The preservation of above-mentioned false pompon pseudoyeast bacterial classification: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in eight months is upgraded once.
The activation of above-mentioned false pompon pseudoyeast bacterial classification: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 17 hours down at 30.5 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone is 11g; Glucose is 22g; Agar is 16g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.2.
The composition of liquid seed culture medium: peptone is 11g; Glucose is 22g; Distilled water is 1000mL, and the pH value of above-mentioned substratum is 5.2.
Embodiment 13:
Present embodiment is produced the processing method of Sodium.alpha.-ketopropionate with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 100g, food-grade albumen peptone 9g, sodium hydroxide 35g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.2, after carrying out sterilization in 12 minutes under 113 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
False pompon pseudoyeast bacterial classification is carried out shake-flask culture, cultivated 20 hours down at 31 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 22 hours, when bacterial classification concentration reaches 1.0%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 30 ℃ in the fermenting process, pressure is 0.04Mpa, time is 51 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains Sodium.alpha.-ketopropionate and bacterial classification is continued deactivation 14 minutes under 84 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get Sodium.alpha.-ketopropionate solution.
The preservation of above-mentioned false pompon pseudoyeast bacterial classification: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in nine months is upgraded once.
The activation of above-mentioned false pompon pseudoyeast bacterial classification: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 20 hours down at 31 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone is 12g; Glucose is 24g; Agar is 17g; Distilled water: 1000mL, the pH value of above-mentioned substratum are 5.3.
The composition of liquid seed culture medium: peptone is 12g; Glucose is 24g; Distilled water is 1000mL, and the pH value of above-mentioned substratum is 5.3.
Embodiment 14:
Present embodiment is produced the processing method of Sodium.alpha.-ketopropionate with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 110g, food-grade albumen peptone 12g, sodium hydroxide 38g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.4, after carrying out sterilization in 14 minutes under 118 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
False pompon pseudoyeast bacterial classification is carried out shake-flask culture, cultivated 22 hours down at 31.5 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 23 hours, when bacterial classification concentration reaches 1.1%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 31 ℃ in the fermenting process, pressure is 0.045Mpa, time is 54 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains Sodium.alpha.-ketopropionate and bacterial classification is continued deactivation 18 minutes under 90 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get Sodium.alpha.-ketopropionate solution.
The preservation of above-mentioned false pompon pseudoyeast bacterial classification: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in ten months is upgraded once.
The activation of above-mentioned false pompon pseudoyeast bacterial classification: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 22 hours down at 31.5 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone: 14g; Glucose: 28g; Agar: 19g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.5.
The composition of liquid seed culture medium: peptone: 14g; Glucose: 28g; Distilled water: 1000mL, the pH value of above-mentioned substratum are 5.5.
Embodiment 15:
Present embodiment is produced the processing method of Sodium.alpha.-ketopropionate with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 120g, food-grade albumen peptone 15g, sodium hydroxide 40g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.6, after carrying out sterilization in 15 minutes under 120 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
False pompon pseudoyeast bacterial classification is carried out shake-flask culture, cultivated 24 hours down at 32 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 24 hours, when bacterial classification concentration reaches 1.2%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 32 ℃ in the fermenting process, pressure is 0.05Mpa, time is 56 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains Sodium.alpha.-ketopropionate and bacterial classification is continued deactivation 20 minutes under 95 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get Sodium.alpha.-ketopropionate solution.
The preservation of above-mentioned false pompon pseudoyeast bacterial classification: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in 1 year is upgraded once.
The activation of above-mentioned false pompon pseudoyeast bacterial classification: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 24 hours down at 32 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone: 15g; Glucose: 30g; Agar: 20g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.6.
The composition of liquid seed culture medium: peptone: 15g; Glucose: 30g; Distilled water 1000mL, the pH value of above-mentioned substratum is: 5.6.
Embodiment 16:
Present embodiment is produced the processing method of Sodium.alpha.-ketopropionate with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 90g, food-grade albumen peptone 3g, sodium hydroxide 30g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.0, after carrying out sterilization in 10 minutes under 108 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
The Torulopsis glabrata kind is carried out shake-flask culture, cultivated 16 hours down at 30 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 20 hours, when bacterial classification concentration reaches 0.8%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 28 ℃ in the fermenting process, pressure is 0.03Mpa, time is 48 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains Sodium.alpha.-ketopropionate and bacterial classification is continued deactivation 10 minutes under 80 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get Sodium.alpha.-ketopropionate solution.
The preservation of above-mentioned Torulopsis glabrata kind: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, half a year, activation was upgraded once.
The activation of above-mentioned Torulopsis glabrata kind: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 16 hours down at 30 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone is 10g; Glucose is 20g; Agar is 15g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.0.
The composition of liquid seed culture medium: peptone is 10g; Glucose is 20g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.0.
Embodiment 17:
Present embodiment is produced the processing method of Sodium.alpha.-ketopropionate with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 95g, food-grade albumen peptone 7g, sodium hydroxide 33g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.1, after carrying out sterilization in 11 minutes under 110 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
The Torulopsis glabrata kind is carried out shake-flask culture, cultivated 18 hours down at 30.5 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 21 hours, when bacterial classification concentration reaches 0.9%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 29 ℃ in the fermenting process, pressure is 0.035Mpa, time is 49 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains Sodium.alpha.-ketopropionate and bacterial classification is continued deactivation 12 minutes under 82 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get Sodium.alpha.-ketopropionate solution.
The preservation of above-mentioned Torulopsis glabrata kind: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in eight months is upgraded once.
The activation of above-mentioned Torulopsis glabrata kind: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 17 hours down at 30.5 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone is 11g; Glucose is 22g; Agar is 16g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.2.
The composition of liquid seed culture medium: peptone is 11g; Glucose is 22g; Distilled water is 1000mL, and the pH value of above-mentioned substratum is 5.2.
Embodiment 18:
Present embodiment is produced the processing method of Sodium.alpha.-ketopropionate with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 100g, food-grade albumen peptone 9g, sodium hydroxide 35g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.2, after carrying out sterilization in 12 minutes under 113 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
The Torulopsis glabrata kind is carried out shake-flask culture, cultivated 20 hours down at 31 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 22 hours, when bacterial classification concentration reaches 1.0%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 30 ℃ in the fermenting process, pressure is 0.04Mpa, time is 51 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains Sodium.alpha.-ketopropionate and bacterial classification is continued deactivation 14 minutes under 84 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get Sodium.alpha.-ketopropionate solution.
The preservation of above-mentioned Torulopsis glabrata kind: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in nine months is upgraded once.
The activation of above-mentioned Torulopsis glabrata kind: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 20 hours down at 31 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone is 12g; Glucose is 24g; Agar is 17g; Distilled water: 1000mL, the pH value of above-mentioned substratum are 5.3.
The composition of liquid seed culture medium: peptone is 12g; Glucose is 24g; Distilled water is 1000mL, and the pH value of above-mentioned substratum is 5.3.
Embodiment 19:
Present embodiment is produced the processing method of Sodium.alpha.-ketopropionate with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 110g, food-grade albumen peptone 12g, sodium hydroxide 38g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.4, after carrying out sterilization in 14 minutes under 118 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
The Torulopsis glabrata kind is carried out shake-flask culture, cultivated 22 hours down at 31.5 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 23 hours, when bacterial classification concentration reaches 1.1%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 31 ℃ in the fermenting process, pressure is 0.045Mpa, time is 54 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains Sodium.alpha.-ketopropionate and bacterial classification is continued deactivation 18 minutes under 90 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get Sodium.alpha.-ketopropionate solution.
The preservation of above-mentioned Torulopsis glabrata kind: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in ten months is upgraded once.
The activation of above-mentioned Torulopsis glabrata kind: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 22 hours down at 31.5 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone: 14g; Glucose: 28g; Agar: 19g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.5.
The composition of liquid seed culture medium: peptone: 14g; Glucose: 28g; Distilled water: 1000mL, the pH value of above-mentioned substratum are 5.5.
Embodiment 20:
Present embodiment is produced the processing method of Sodium.alpha.-ketopropionate with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 120g, food-grade albumen peptone 15g, sodium hydroxide 40g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.6, after carrying out sterilization in 15 minutes under 120 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
The Torulopsis glabrata kind is carried out shake-flask culture, cultivated 24 hours down at 32 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 24 hours, when bacterial classification concentration reaches 1.2%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 32 ℃ in the fermenting process, pressure is 0.05Mpa, time is 56 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains Sodium.alpha.-ketopropionate and bacterial classification is continued deactivation 20 minutes under 95 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get Sodium.alpha.-ketopropionate solution.
The preservation of above-mentioned Torulopsis glabrata kind: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in 1 year is upgraded once.
The activation of above-mentioned Torulopsis glabrata kind: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 24 hours down at 32 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone: 15g; Glucose: 30g; Agar: 20g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.6.
The composition of liquid seed culture medium: peptone: 15g; Glucose: 30g; Distilled water 1000mL, the pH value of above-mentioned substratum is: 5.6.
Embodiment 21:
Present embodiment is produced the processing method of Sodium.alpha.-ketopropionate with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 90g, food-grade albumen peptone 3g, sodium bicarbonate 30g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.0, after carrying out sterilization in 10 minutes under 108 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
False pompon pseudoyeast bacterial classification is carried out shake-flask culture, cultivated 16 hours down at 30 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 20 hours, when bacterial classification concentration reaches 0.8%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 28 ℃ in the fermenting process, pressure is 0.03Mpa, time is 48 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains Sodium.alpha.-ketopropionate and bacterial classification is continued deactivation 10 minutes under 80 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get Sodium.alpha.-ketopropionate solution.
The preservation of above-mentioned false pompon pseudoyeast bacterial classification: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, half a year, activation was upgraded once.
The activation of above-mentioned false pompon pseudoyeast bacterial classification: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 16 hours down at 30 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone is 10g; Glucose is 20g; Agar is 15g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.0.
The composition of liquid seed culture medium: peptone is 10g; Glucose is 20g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.0.
Embodiment 22:
Present embodiment is produced the processing method of Sodium.alpha.-ketopropionate with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 95g, food-grade albumen peptone 7g, sodium bicarbonate 33g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.1, after carrying out sterilization in 11 minutes under 110 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
False pompon pseudoyeast bacterial classification is carried out shake-flask culture, cultivated 18 hours down at 30.5 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 21 hours, when bacterial classification concentration reaches 0.9%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 29 ℃ in the fermenting process, pressure is 0.035Mpa, time is 49 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains Sodium.alpha.-ketopropionate and bacterial classification is continued deactivation 12 minutes under 82 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get Sodium.alpha.-ketopropionate solution.
The preservation of above-mentioned false pompon pseudoyeast bacterial classification: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in eight months is upgraded once.
The activation of above-mentioned false pompon pseudoyeast bacterial classification: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 17 hours down at 30.5 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone is 11g; Glucose is 22g; Agar is 16g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.2.
The composition of liquid seed culture medium: peptone is 11g; Glucose is 22g; Distilled water is 1000mL, and the pH value of above-mentioned substratum is 5.2.
Embodiment 23:
Present embodiment is produced the processing method of Sodium.alpha.-ketopropionate with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 100g, food-grade albumen peptone 9g, sodium bicarbonate 35g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.2, after carrying out sterilization in 12 minutes under 113 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
False pompon pseudoyeast bacterial classification is carried out shake-flask culture, cultivated 20 hours down at 31 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 22 hours, when bacterial classification concentration reaches 1.0%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 30 ℃ in the fermenting process, pressure is 0.04Mpa, time is 51 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains Sodium.alpha.-ketopropionate and bacterial classification is continued deactivation 14 minutes under 84 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get Sodium.alpha.-ketopropionate solution.
The preservation of above-mentioned false pompon pseudoyeast bacterial classification: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in nine months is upgraded once.
The activation of above-mentioned false pompon pseudoyeast bacterial classification: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 20 hours down at 31 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone is 12g; Glucose is 24g; Agar is 17g; Distilled water: 1000mL, the pH value of above-mentioned substratum are 5.3.
The composition of liquid seed culture medium: peptone is 12g; Glucose is 24g; Distilled water is 1000mL, and the pH value of above-mentioned substratum is 5.3.
Embodiment 24:
Present embodiment is produced the processing method of Sodium.alpha.-ketopropionate with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 110g, food-grade albumen peptone 12g, sodium bicarbonate 38g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.4, after carrying out sterilization in 14 minutes under 118 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
False pompon pseudoyeast bacterial classification is carried out shake-flask culture, cultivated 22 hours down at 31.5 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 23 hours, when bacterial classification concentration reaches 1.1%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 31 ℃ in the fermenting process, pressure is 0.045Mpa, time is 54 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains Sodium.alpha.-ketopropionate and bacterial classification is continued deactivation 18 minutes under 90 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get Sodium.alpha.-ketopropionate solution.
The preservation of above-mentioned false pompon pseudoyeast bacterial classification: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in ten months is upgraded once.
The activation of above-mentioned false pompon pseudoyeast bacterial classification: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 22 hours down at 31.5 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone: 14g; Glucose: 28g; Agar: 19g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.5.
The composition of liquid seed culture medium: peptone: 14g; Glucose: 28g; Distilled water: 1000mL, the pH value of above-mentioned substratum are 5.5.
Embodiment 25:
Present embodiment is produced the processing method of Sodium.alpha.-ketopropionate with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 120g, food-grade albumen peptone 15g, sodium bicarbonate 40g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.6, after carrying out sterilization in 15 minutes under 120 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
False pompon pseudoyeast bacterial classification is carried out shake-flask culture, cultivated 24 hours down at 32 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 24 hours, when bacterial classification concentration reaches 1.2%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 32 ℃ in the fermenting process, pressure is 0.05Mpa, time is 56 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains Sodium.alpha.-ketopropionate and bacterial classification is continued deactivation 20 minutes under 95 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get Sodium.alpha.-ketopropionate solution.
The preservation of above-mentioned false pompon pseudoyeast bacterial classification: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in 1 year is upgraded once.
The activation of above-mentioned false pompon pseudoyeast bacterial classification: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 24 hours down at 32 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone: 15g; Glucose: 30g; Agar: 20g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.6.
The composition of liquid seed culture medium: peptone: 15g; Glucose: 30g; Distilled water 1000mL, the pH value of above-mentioned substratum is: 5.6.
Embodiment 26:
Present embodiment is produced the processing method of Sodium.alpha.-ketopropionate with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 90g, food-grade albumen peptone 3g, sodium bicarbonate 30g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.0, after carrying out sterilization in 10 minutes under 108 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
The Torulopsis glabrata kind is carried out shake-flask culture, cultivated 16 hours down at 30 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 20 hours, when bacterial classification concentration reaches 0.8%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 28 ℃ in the fermenting process, pressure is 0.03Mpa, time is 48 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains Sodium.alpha.-ketopropionate and bacterial classification is continued deactivation 10 minutes under 80 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get Sodium.alpha.-ketopropionate solution.
The preservation of above-mentioned Torulopsis glabrata kind: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, half a year, activation was upgraded once.
The activation of above-mentioned Torulopsis glabrata kind: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 16 hours down at 30 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone is 10g; Glucose is 20g; Agar is 15g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.0.
The composition of liquid seed culture medium: peptone is 10g; Glucose is 20g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.0.
Embodiment 27:
Present embodiment is produced the processing method of Sodium.alpha.-ketopropionate with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 95g, food-grade albumen peptone 7g, sodium bicarbonate 33g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.1, after carrying out sterilization in 11 minutes under 110 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
The Torulopsis glabrata kind is carried out shake-flask culture, cultivated 18 hours down at 30.5 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 21 hours, when bacterial classification concentration reaches 0.9%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 29 ℃ in the fermenting process, pressure is 0.035Mpa, time is 49 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains Sodium.alpha.-ketopropionate and bacterial classification is continued deactivation 12 minutes under 82 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get Sodium.alpha.-ketopropionate solution.
The preservation of above-mentioned Torulopsis glabrata kind: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in eight months is upgraded once.
The activation of above-mentioned Torulopsis glabrata kind: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 17 hours down at 30.5 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone is 11g; Glucose is 22g; Agar is 16g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.2.
The composition of liquid seed culture medium: peptone is 11g; Glucose is 22g; Distilled water is 1000mL, and the pH value of above-mentioned substratum is 5.2.
Embodiment 28:
Present embodiment is produced the processing method of Sodium.alpha.-ketopropionate with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 100g, food-grade albumen peptone 9g, sodium bicarbonate 35g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.2, after carrying out sterilization in 12 minutes under 113 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
The Torulopsis glabrata kind is carried out shake-flask culture, cultivated 20 hours down at 31 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 22 hours, when bacterial classification concentration reaches 1.0%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 30 ℃ in the fermenting process, pressure is 0.04Mpa, time is 51 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains Sodium.alpha.-ketopropionate and bacterial classification is continued deactivation 14 minutes under 84 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get Sodium.alpha.-ketopropionate solution.
The preservation of above-mentioned Torulopsis glabrata kind: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in nine months is upgraded once.
The activation of above-mentioned Torulopsis glabrata kind: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 20 hours down at 31 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone is 12g; Glucose is 24g; Agar is 17g; Distilled water: 1000mL, the pH value of above-mentioned substratum are 5.3.
The composition of liquid seed culture medium: peptone is 12g; Glucose is 24g; Distilled water is 1000mL, and the pH value of above-mentioned substratum is 5.3.
Embodiment 29:
Present embodiment is produced the processing method of Sodium.alpha.-ketopropionate with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 110g, food-grade albumen peptone 12g, sodium bicarbonate 38g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.4, after carrying out sterilization in 14 minutes under 118 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
The Torulopsis glabrata kind is carried out shake-flask culture, cultivated 22 hours down at 31.5 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 23 hours, when bacterial classification concentration reaches 1.1%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 31 ℃ in the fermenting process, pressure is 0.045Mpa, time is 54 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains Sodium.alpha.-ketopropionate and bacterial classification is continued deactivation 18 minutes under 90 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get Sodium.alpha.-ketopropionate solution.
The preservation of above-mentioned Torulopsis glabrata kind: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in ten months is upgraded once.
The activation of above-mentioned Torulopsis glabrata kind: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 22 hours down at 31.5 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone: 14g; Glucose: 28g; Agar: 19g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.5.
The composition of liquid seed culture medium: peptone: 14g; Glucose: 28g; Distilled water: 1000mL, the pH value of above-mentioned substratum are 5.5.
Embodiment 30:
Present embodiment is produced the processing method of Sodium.alpha.-ketopropionate with biological fermentation process, mainly comprises following processing step:
(1) fermention medium batching:
At normal temperatures, put into oral glucose 120g, food-grade albumen peptone 15g, sodium bicarbonate 40g in the distilled water of 1000mL, with the said mixture thorough mixing, the pH value of mixture is 5.6, after carrying out sterilization in 15 minutes under 120 ℃ of temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
The Torulopsis glabrata kind is carried out shake-flask culture, cultivated 24 hours down at 32 ℃, when reaching the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing, bacterial classification was cultivated in seeding tank 24 hours, when bacterial classification concentration reaches 1.2%, change over to and carry out fermentation culture in the above-mentioned fermentor tank;
(3) fermentation:
Temperature is controlled at 32 ℃ in the fermenting process, pressure is 0.05Mpa, time is 56 hours, during the fermentation, contain the glucose of high density in the fermentor tank, the glucose of this moment, except the normal growth that guarantees bacterial classification, also supply raw materials, reduce to 1% when following, stop fermenting when glucose residual sugar in the fermented liquid for producing pyruvic acid;
(4) the above-mentioned fermented liquid that contains Sodium.alpha.-ketopropionate and bacterial classification is continued deactivation 20 minutes under 95 ℃ of temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get Sodium.alpha.-ketopropionate solution.
The preservation of above-mentioned Torulopsis glabrata kind: above-mentioned bacterial classification is placed inclined-plane solid seed culture medium, be kept in 4 ℃ of refrigerators, activation in 1 year is upgraded once.
The activation of above-mentioned Torulopsis glabrata kind: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, liquid amount 100/500mL, cultivated 24 hours down at 32 ℃, reach the maximum growth logarithmic phase of bacterial classification, enter seeding tank and carry out the seed enlarged culturing.
The composition of inclined-plane solid seed culture medium: peptone: 15g; Glucose: 30g; Agar: 20g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.6.
The composition of liquid seed culture medium: peptone: 15g; Glucose: 30g; Distilled water 1000mL, the pH value of above-mentioned substratum is: 5.6.
Claims (4)
1, a kind of processing method with preparing pyruvate by biological fermentation method is characterized in that mainly comprising following processing step:
(1) fermention medium batching:
In the water of every 1000mL, put into glucose 90g～120g, peptone 3～15g, lime carbonate or sodium hydroxid or sodium bicarbonate 30g～40g, with the above-mentioned raw materials thorough mixing, the pH value of mixture is 5.0～5.6, after carrying out sterilization in 10 minutes～15 minutes under 108 ℃～120 ℃ temperature, it is standby to put into fermentor tank with said mixture;
(2) fermented bacterium is cultivated:
False pompon pseudoyeast or torulopsis glabrata are carried out the seed enlarged culturing again after cultivating 16～24 hours under 30～32 ℃,, when bacterial classification concentration weight ratio reaches 0.8%～1.2%, change over to and carry out fermentation culture in the above-mentioned fermentor tank through cultivating 20～24 hours;
(3) fermentation:
Temperature is controlled at 28 ℃～32 ℃ in the fermenting process, and pressure is 0.03Mpa～0.05Mpa, and the time is 48～56 hours, reduces to 1% when following when glucose residual sugar in the fermented liquid, stops fermentation;
(4) the above-mentioned fermented liquid that contains pyruvate salt and bacterial classification is continued deactivation 10 minutes～20 minutes under 80 ℃～95 ℃ temperature, bacterial classification is lost vigor, after after precipitating, filter, sterilizing, promptly get pyruvate salt solution.
2, the processing method with preparing pyruvate by biological fermentation method as claimed in claim 1, it is characterized in that the activation of described false pompon pseudoyeast or Torulopsis glabrata kind: on inclined-plane solid seed culture medium, get a ring bacterial classification and place liquid seed culture medium, carry out shake-flask culture, after cultivating 16～24 hours under 30 ℃～32 ℃, enter seeding tank and carry out the seed enlarged culturing.
3, the processing method with preparing pyruvate by biological fermentation method as claimed in claim 2, it is characterized in that the composition of described inclined-plane solid seed culture medium: peptone is 10～15g; Glucose is 20～30g; Agar is 15～20g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.0～5.6.
4, the processing method with preparing pyruvate by biological fermentation method as claimed in claim 2, it is characterized in that the composition of described liquid seed culture medium: peptone is 10～15g; Glucose is 20～30g; Distilled water 1000mL, the pH value of above-mentioned substratum are 5.0～5.6.
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| CN102864178A (en) * | 2012-08-29 | 2013-01-09 | 太仓市茂通化建有限公司 | Method for producing pyruvic acid by utilizing torulopsis glabrata |
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