CN101914512B - Method for fermenting chymosin by using nisin fermentation wastewater as partial nitrogen source - Google Patents
Method for fermenting chymosin by using nisin fermentation wastewater as partial nitrogen source Download PDFInfo
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- IJGRMHOSHXDMSA-UHFFFAOYSA-N nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 title claims abstract description 76
- NVNLLIYOARQCIX-MSHCCFNRSA-N Nisin Chemical compound N1C(=O)[C@@H](CC(C)C)NC(=O)C(=C)NC(=O)[C@@H]([C@H](C)CC)NC(=O)[C@@H](NC(=O)C(=C/C)/NC(=O)[C@H](N)[C@H](C)CC)CSC[C@@H]1C(=O)N[C@@H]1C(=O)N2CCC[C@@H]2C(=O)NCC(=O)N[C@@H](C(=O)N[C@H](CCCCN)C(=O)N[C@@H]2C(NCC(=O)N[C@H](C)C(=O)N[C@H](CC(C)C)C(=O)N[C@H](CCSC)C(=O)NCC(=O)N[C@H](CS[C@@H]2C)C(=O)N[C@H](CC(N)=O)C(=O)N[C@H](CCSC)C(=O)N[C@H](CCCCN)C(=O)N[C@@H]2C(N[C@H](C)C(=O)N[C@@H]3C(=O)N[C@@H](C(N[C@H](CC=4NC=NC=4)C(=O)N[C@H](CS[C@@H]3C)C(=O)N[C@H](CO)C(=O)N[C@H]([C@H](C)CC)C(=O)N[C@H](CC=3NC=NC=3)C(=O)N[C@H](C(C)C)C(=O)NC(=C)C(=O)N[C@H](CCCCN)C(O)=O)=O)CS[C@@H]2C)=O)=O)CS[C@@H]1C NVNLLIYOARQCIX-MSHCCFNRSA-N 0.000 title claims abstract description 50
- 108010053775 Nisin Proteins 0.000 title claims abstract description 50
- 239000004309 nisin Substances 0.000 title claims abstract description 50
- 235000010297 nisin Nutrition 0.000 title claims abstract description 50
- 229910052757 nitrogen Inorganic materials 0.000 title claims abstract description 38
- 108090000746 Chymosin Proteins 0.000 title abstract description 32
- 238000000855 fermentation Methods 0.000 title abstract description 11
- 230000004151 fermentation Effects 0.000 title abstract description 11
- 229940080701 Chymosin Drugs 0.000 title abstract 6
- 239000002351 wastewater Substances 0.000 title abstract 4
- 239000001963 growth media Substances 0.000 claims abstract description 30
- 241000235403 Rhizomucor miehei Species 0.000 claims abstract description 5
- 239000002921 fermentation waste Substances 0.000 claims description 43
- 241000235395 Mucor Species 0.000 claims description 30
- 241000209094 Oryza Species 0.000 claims description 30
- 235000007164 Oryza sativa Nutrition 0.000 claims description 30
- 241001622468 Pterocaulon Species 0.000 claims description 30
- 235000009566 rice Nutrition 0.000 claims description 30
- 108010073771 Soybean Proteins Proteins 0.000 claims description 28
- 235000019710 soybean protein Nutrition 0.000 claims description 28
- 210000000582 Semen Anatomy 0.000 claims description 27
- 239000000843 powder Substances 0.000 claims description 27
- HEMHJVSKTPXQMS-UHFFFAOYSA-M sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 24
- WQZGKKKJIJFFOK-GASJEMHNSA-N D-Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 claims description 16
- 102000004139 alpha-Amylases Human genes 0.000 claims description 16
- 108090000637 alpha-Amylases Proteins 0.000 claims description 16
- 229940024171 alpha-amylase Drugs 0.000 claims description 16
- 239000008103 glucose Substances 0.000 claims description 16
- 235000010482 polyoxyethylene sorbitan monooleate Nutrition 0.000 claims description 16
- 229920000053 polysorbate 80 Polymers 0.000 claims description 16
- 239000011734 sodium Substances 0.000 claims description 16
- BZKBCQXYZZXSCO-UHFFFAOYSA-N sodium hydride Inorganic materials [H-].[Na+] BZKBCQXYZZXSCO-UHFFFAOYSA-N 0.000 claims description 16
- 239000002054 inoculum Substances 0.000 claims description 14
- 238000002360 preparation method Methods 0.000 claims description 12
- 229920000591 gum Polymers 0.000 claims description 11
- 241000233866 Fungi Species 0.000 claims description 8
- 230000001954 sterilising Effects 0.000 claims description 8
- 238000004659 sterilization and disinfection Methods 0.000 claims description 8
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 8
- 238000011081 inoculation Methods 0.000 claims description 7
- 238000011177 media preparation Methods 0.000 claims description 7
- 244000005700 microbiome Species 0.000 claims description 2
- 230000000813 microbial Effects 0.000 abstract description 9
- 239000002699 waste material Substances 0.000 abstract description 5
- 238000004519 manufacturing process Methods 0.000 abstract description 4
- 238000007599 discharging Methods 0.000 abstract description 2
- 238000001514 detection method Methods 0.000 description 4
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- 239000012530 fluid Substances 0.000 description 3
- 238000005516 engineering process Methods 0.000 description 2
- QJGQUHMNIGDVPM-UHFFFAOYSA-N nitrogen group Chemical group [N] QJGQUHMNIGDVPM-UHFFFAOYSA-N 0.000 description 2
- 230000002797 proteolythic Effects 0.000 description 2
- 229940108461 rennet Drugs 0.000 description 2
- 108010058314 rennet Proteins 0.000 description 2
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- 240000003440 Asimina triloba Species 0.000 description 1
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- 108091005540 Aspartic proteases Proteins 0.000 description 1
- 102000035336 Aspartic proteases Human genes 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- 235000009467 Carica papaya Nutrition 0.000 description 1
- 235000011201 Ginkgo Nutrition 0.000 description 1
- 240000002883 Ginkgo biloba Species 0.000 description 1
- 240000007472 Leucaena leucocephala Species 0.000 description 1
- 235000010643 Leucaena leucocephala Nutrition 0.000 description 1
- 210000004080 Milk Anatomy 0.000 description 1
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Abstract
The invention provides a method for fermenting chymosin by using nisin fermentation wastewater as a partial nitrogen source, relating to the method for producing the chymosin by microbial fermentation. The invention solves the problems that at current, the method for producing the chymosin by the microbial fermentation has high cost, and the direct discharge of the fermentation wastewater of the nisin results in environment pollution. The method comprises the following steps: 1, preparing a culture medium; 2, preparing rhizomucor miehei seed liquor; 3, inoculating the rhizomucor miehei seed liquor to a triangular flask; and 4, culturing the inoculated seed liquor for 96h at the temperature of 37-38 DEG C to obtain the chymosin. The invention can utilize waste, reduce the environment pollution caused by directly discharging the fermentation wastewater of the nisin, and lower the cost for fermenting the chymosin. The invention can be applied to the field of microbial fermentation.
Description
Technical field
The present invention relates to a kind of microbial fermentation and produce the method for rennin.
Background technology
Rennin belongs to aspartic protease, claims aspartate protease again, is to produce the indispensable preparation of cheese.The tradition source of rennin is the abomasum of phase lactation calf; The ratios of animal rennet and curdled milk vigor and proteolytic activity becomes makes caseic first-selected enzyme; But growth of animal slowly and cost an arm and a leg; Increase easily enterprise cost, and from animal, extract complicacy of enzyme liquid program and technology.All contain the proteolytic enzyme that can make curdling solid in the plants such as pawpaw, Fructus Fici, pineapple, difficulty, acacia, ginkgo; But the rennin of plant origin is because of there being too high proteolyze vigor or own poisonous, and therefore a lot of plant rennets do not obtain the large-scale commercial applications application.
Microbial rennet wide material sources, but kind of mikrobe fermenting and producing rennin surplus finding have 40 at present, if these microbial host actinomycetes, bacterium and fungi etc., because the control of the method for microbial fermentation easily, cost is low, and safety non-toxic, and is widely used.The nitrogenous source of the rennin of microbial fermentation production at present is mainly from textured soybean protein and Semen Maydis powder, and production cost is higher.
Contain abundant nitrogenous source in the fermented waste fluid of nisin.With the fermented waste fluid of nisin directly the discharging meeting environment is polluted, enter the difficulty that treatment tank also can increase processing.
Summary of the invention
The present invention produces the cost height of rennin in order to solve present microbial fermentation, and the direct exhaust emission problem of environment of the fermented waste fluid of nisin provides a kind of method of utilizing nisin fermentation waste solution to make the partial nitrogen source fermenting and producing rennin.
The present invention utilizes nisin fermentation waste solution to make the method for partial nitrogen source fermenting and producing rennin, carries out according to the following steps: one, culture medium preparation: with NaH
2PO
4, Na
2HPO
4, glucose, alpha-amylase, tween 80, Semen Maydis powder, W-Gum, nisin fermentation waste solution and textured soybean protein be dissolved in the water preparing culture medium, wherein NaH
2PO
4: 3.25g/L, Na
2HPO
4: 3.11g/L, glucose: 2.14g/L, alpha-amylase: 20g/L, tween 80: 0.13g/L, Semen Maydis powder: 30～80g/L, W-Gum: 0～30g/L, nisin fermentation waste solution: 150～700g/L, textured soybean protein: 0～20g/L; The use mass concentration is 5%～6% sodium hydroxide accent pH to 6.7～6.8, sterilization 30min under 0.14MPa pressure; Two, the preparation of rice black root Mucor seed liquor: with rice black root Mucor by volume 4% inoculum size be inoculated in the seed culture medium, shaking table is cultivated 20～24h under 37～38 ℃ of conditions, the rotating speed of shaking table is 260rpm; Three, inoculation: prepared culture medium in the step 1 is packed in the triangular flask of 250ml, with rice black root Mucor seed liquor by volume 2.5% inoculum size be inoculated in the triangular flask; Four, cultivate: triangular flask is placed down on the shaking table in 37～38 ℃ of conditions cultivates, the rotating speed of shaking table is 260～280rpm, cultivates 96 hours, promptly obtains rennin; The nitrogen content of Semen Maydis powder is 1.075% (quality) in the step 1, and the nitrogen content of nisin fermentation waste solution is 0.24% (quality), and the nitrogen content of textured soybean protein is 8% (quality); The fungi count of the rice black root Mucor seed liquor that makes in the step 2 is 2～6 * 10
6Cfu/ml; The Intake Quantity of substratum is 20% of a triangular flask volume in the step 3.
The present invention utilizes nisin fermentation waste solution to make partial nitrogen source producing lab ferment in next life, has accomplished utilization of waste material, has reduced the direct environmental pollution caused by discharge of nisin fermentation waste solution, has reduced the cost of fermenting and producing rennin.
Embodiment
Technical scheme of the present invention is not limited to following cited embodiment, also comprises the arbitrary combination between each embodiment.
Embodiment one: this embodiment utilizes nisin fermentation waste solution to make the method for partial nitrogen source fermenting and producing rennin, carries out according to the following steps: one, culture medium preparation: with NaH
2PO
4, Na
2HPO
4, glucose, alpha-amylase, tween 80, Semen Maydis powder, W-Gum, nisin fermentation waste solution and textured soybean protein be dissolved in the water preparing culture medium, wherein NaH
2PO
4: 3.25g/L, Na
2HPO
4: 3.11g/L, glucose: 2.14g/L, alpha-amylase: 20g/L, tween 80: 0.13g/L, Semen Maydis powder: 30～80g/L, W-Gum: 0～30g/L, nisin fermentation waste solution: 150～700g/L, textured soybean protein: 0～20g/L; The use mass concentration is 5%～6% sodium hydroxide accent pH to 6.7～6.8, sterilization 30min under 0.14MPa pressure; Two, the preparation of rice black root Mucor seed liquor: with rice black root Mucor by volume 4% inoculum size be inoculated in the seed culture medium, shaking table is cultivated 20～24h under 37～38 ℃ of conditions, the rotating speed of shaking table is 260rpm; Three, inoculation: prepared culture medium in the step 1 is packed in the triangular flask of 250ml, with rice black root Mucor seed liquor by volume 2.5% inoculum size be inoculated in the triangular flask; Four, cultivate: triangular flask is placed down on the shaking table in 37～38 ℃ of conditions cultivates, the rotating speed of shaking table is 260～280rpm, cultivates 96 hours, promptly obtains rennin; The nitrogen content of Semen Maydis powder is 1.075% (quality) in the step 1, and the nitrogen content of nisin fermentation waste solution is 0.24% (quality), and the nitrogen content of textured soybean protein is 8% (quality); The fungi count of the rice black root Mucor seed liquor that makes in the step 2 is 2～6 * 10
6Cfu/ml; The Intake Quantity of substratum is 20% of a triangular flask volume in the step 3.
Seed culture medium in this embodiment step 2 is by NaH
2PO
4, Na
2HPO
4, glucose, alpha-amylase, tween 80, Semen Maydis powder, textured soybean protein and water forms; NaH wherein
2PO
4: 3.25g/L, Na
2HPO
4: 3.11g/L, glucose: 2.14g/L, alpha-amylase: 20g/L, tween 80: 0.13g/L, Semen Maydis powder: 70g/L, textured soybean protein: 18g/L; The use mass concentration is 5%～6% sodium hydroxide accent pH to 6.7～6.8, sterilization 30min under 0.14MPa pressure.
Rice black root Mucor (Rhizomucor miehei) in this embodiment step 2 is preserved in Chinese common micro-organisms culture presevation administrative center, and culture presevation is numbered AS 3.4960.
Embodiment two: what this embodiment and embodiment one were different is: the Semen Maydis powder concentration that adds in the step 1 is 35～70g/L.Other is identical with embodiment one.
Embodiment three: what this embodiment was different with embodiment one or two is: the Semen Maydis powder concentration that adds in the step 1 is 50～60g/L.Other is identical with embodiment one or two.
Embodiment four: what this embodiment was different with one of embodiment one to three is: the W-Gum concentration that adds in the step 1 is 5～25g/L.Other is identical with one of embodiment one to three.
Embodiment five: what this embodiment was different with one of embodiment one to four is: the W-Gum concentration that adds in the step 1 is 10～20g/L.Other is identical with one of embodiment one to four.
Embodiment six: what this embodiment was different with one of embodiment one to five is: the W-Gum concentration that adds in the step 1 is 26.25g/L.Other is identical with one of embodiment one to five.
Embodiment seven: what this embodiment was different with one of embodiment one to six is: the nisin fermentation waste solution concentration that adds in the step 1 is 200～600g/L.Other is identical with one of embodiment one to six.
Embodiment eight: what this embodiment was different with one of embodiment one to seven is: the nisin fermentation waste solution concentration that adds in the step 1 is 300～500g/L.Other is identical with one of embodiment one to seven.
Embodiment nine: what this embodiment was different with one of embodiment one to eight is: the nisin fermentation waste solution concentration that adds in the step 1 is 160g/L.Other is identical with one of embodiment one to eight.
Embodiment ten: what this embodiment was different with one of embodiment one to nine is: the nisin fermentation waste solution concentration that adds in the step 1 is 480g/L.Other is identical with one of embodiment one to nine.
Embodiment 11: what this embodiment was different with one of embodiment one to ten is: the textured soybean protein's concentration that adds in the step 1 is 5～15g/L.Other is identical with one of embodiment one to ten.
Embodiment 12: what this embodiment was different with one of embodiment one to 11 is: the textured soybean protein's concentration that adds in the step 1 is 10g/L.Other is identical with one of embodiment one to 11.
Embodiment 13: what this embodiment was different with one of embodiment one to 12 is: the textured soybean protein's concentration that adds in the step 1 is 3.6g/L.Other is identical with one of embodiment one to 12.
Embodiment 14: what this embodiment was different with one of embodiment one to 13 is: the textured soybean protein's concentration that adds in the step 1 is 9g/L.Other is identical with one of embodiment one to 13.
Embodiment 15: what this embodiment was different with one of embodiment one to 14 is: the textured soybean protein's concentration that adds in the step 1 is 18g/L.Other is identical with one of embodiment one to 14.
Embodiment 16: what this embodiment was different with one of embodiment one to 15 is: the rotating speed of shaking table is 270rpm in the step 4.Other is identical with one of embodiment one to 15.
Embodiment 17: this embodiment utilizes nisin fermentation waste solution to make the method for partial nitrogen source fermenting and producing rennin, carries out according to the following steps: one, culture medium preparation: with NaH
2PO
4, Na
2HPO
4, glucose, alpha-amylase, tween 80, Semen Maydis powder and nisin fermentation waste solution be dissolved in the water preparing culture medium, wherein NaH
2PO
4: 3.25g/L, Na
2HPO
4: 3.11g/L, glucose: 2.14g/L, alpha-amylase: 20g/L, tween 80: 0.13g/L, Semen Maydis powder: 70g/L, nisin fermentation waste solution: 600g/L; The use mass concentration is 5%～6% sodium hydroxide accent pH to 6.7～6.8, sterilization 30min under 0.14MPa pressure; Two, the preparation of rice black root Mucor seed liquor: with rice black root Mucor by volume 4% inoculum size be inoculated in the seed culture medium, shaking table is cultivated 20～24h under 37～38 ℃ of conditions, the rotating speed of shaking table is 260rpm; Three, inoculation: prepared culture medium in the step 1 is packed in the triangular flask of 250ml, with rice black root Mucor seed liquor by volume 2.5% inoculum size be inoculated in the triangular flask; Four, cultivate: triangular flask is placed down on the shaking table in 37～38 ℃ of conditions cultivates, the rotating speed of shaking table is 260rpm, cultivates 96 hours, promptly obtains rennin; The nitrogen content of Semen Maydis powder is 1.075% (quality) in the step 1, and the nitrogen content of nisin fermentation waste solution is 0.24% (quality), and the nitrogen content of textured soybean protein is 8% (quality); The fungi count of the rice black root Mucor seed liquor that makes in the step 2 is 2～6 * 10
6Cfu/ml; The Intake Quantity of substratum is 20% of a triangular flask volume in the step 3.
The rennin of this embodiment preparation is 53IMCU/ml through the detection vigor.
Embodiment 18: this embodiment utilizes nisin fermentation waste solution to make the method for partial nitrogen source fermenting and producing rennin, carries out according to the following steps: one, culture medium preparation: with NaH
2PO
4, Na
2HPO
4, glucose, alpha-amylase, tween 80, Semen Maydis powder, nisin fermentation waste solution and textured soybean protein be dissolved in the water preparing culture medium, wherein NaH
2PO
4: 3.25g/L, Na
2HPO
4: 3.11g/L, glucose: 2.14g/L, alpha-amylase: 20g/L, tween 80: 0.13g/L, Semen Maydis powder: 70g/L, nisin fermentation waste solution: 480g/L, textured soybean protein: 3.6g/L; The use mass concentration is 5%～6% sodium hydroxide accent pH to 6.7～6.8, sterilization 30min under 0.14MPa pressure; Two, the preparation of rice black root Mucor seed liquor: with rice black root Mucor by volume 4% inoculum size be inoculated in the seed culture medium, shaking table is cultivated 20～24h under 37～38 ℃ of conditions, the rotating speed of shaking table is 260rpm; Three, inoculation: prepared culture medium in the step 1 is packed in the triangular flask of 250ml, with rice black root Mucor seed liquor by volume 2.5% inoculum size be inoculated in the triangular flask; Four, cultivate: triangular flask is placed down on the shaking table in 37～38 ℃ of conditions cultivates, the rotating speed of shaking table is 270rpm, cultivates 96 hours, promptly obtains rennin; The nitrogen content of Semen Maydis powder is 1.075% (quality) in the step 1, and the nitrogen content of nisin fermentation waste solution is 0.24% (quality), and the nitrogen content of textured soybean protein is 8% (quality); The fungi count of the rice black root Mucor seed liquor that makes in the step 2 is 2～6 * 10
6Cfu/ml; The Intake Quantity of substratum is 20% of a triangular flask volume in the step 3.
The rennin of this embodiment preparation is 75IMCU/ml through the detection vigor.
Embodiment 19: this embodiment utilizes nisin fermentation waste solution to make the method for partial nitrogen source fermenting and producing rennin, carries out according to the following steps: one, culture medium preparation: with NaH
2PO
4, Na
2HPO
4, glucose, alpha-amylase, tween 80, Semen Maydis powder, nisin fermentation waste solution and textured soybean protein be dissolved in the water preparing culture medium, wherein NaH
2PO
4: 3.25g/L, Na
2HPO
4: 3.11g/L, glucose: 2.14g/L, alpha-amylase: 20g/L, tween 80: 0.13g/L, Semen Maydis powder: 70g/L, nisin fermentation waste solution: 300g/L, textured soybean protein: 9g/L; The use mass concentration is 5%～6% sodium hydroxide accent pH to 6.7～6.8, sterilization 30min under 0.14MPa pressure; Two, the preparation of rice black root Mucor seed liquor: with rice black root Mucor by volume 4% inoculum size be inoculated in the seed culture medium, shaking table is cultivated 20～24h under 37～38 ℃ of conditions, the rotating speed of shaking table is 260rpm; Three, inoculation: prepared culture medium in the step 1 is packed in the triangular flask of 250ml, with rice black root Mucor seed liquor by volume 2.5% inoculum size be inoculated in the triangular flask; Four, cultivate: triangular flask is placed down on the shaking table in 37～38 ℃ of conditions cultivates, the rotating speed of shaking table is 260rpm, cultivates 96 hours, promptly obtains rennin; The nitrogen content of Semen Maydis powder is 1.075% (quality) in the step 1, and the nitrogen content of nisin fermentation waste solution is 0.24% (quality), and the nitrogen content of textured soybean protein is 8% (quality); The fungi count of the rice black root Mucor seed liquor that makes in the step 2 is 2～6 * 10
6Cfu/ml; The Intake Quantity of substratum is 20% of a triangular flask volume in the step 3.
The rennin of this embodiment preparation is 93IMCU/ml through the detection vigor.
Embodiment 20: this embodiment utilizes nisin fermentation waste solution to make the method for partial nitrogen source fermenting and producing rennin, carries out according to the following steps: one, culture medium preparation: with NaH
2PO
4, Na
2HPO
4, glucose, alpha-amylase, tween 80, Semen Maydis powder, W-Gum, nisin fermentation waste solution and textured soybean protein be dissolved in the water preparing culture medium, wherein NaH
2PO
4: 3.25g/L, Na
2HPO
4: 3.11g/L, glucose: 2.14g/L, alpha-amylase: 20g/L, tween 80: 0.13g/L, Semen Maydis powder: 35g/L, W-Gum: 26.25g/L, nisin fermentation waste solution: 160g/L, textured soybean protein: 18g/L; The use mass concentration is 5%～6% sodium hydroxide accent pH to 6.7～6.8, sterilization 30min under 0.14MPa pressure; Two, the preparation of rice black root Mucor seed liquor: with rice black root Mucor by volume 4% inoculum size be inoculated in the seed culture medium, shaking table is cultivated 20～24h under 37～38 ℃ of conditions, the rotating speed of shaking table is 260rpm; Three, inoculation: prepared culture medium in the step 1 is packed in the triangular flask of 250ml, with rice black root Mucor seed liquor by volume 2.5% inoculum size be inoculated in the triangular flask; Four, cultivate: triangular flask is placed down on the shaking table in 37～38 ℃ of conditions cultivates, the rotating speed of shaking table is 280rpm, cultivates 96 hours, promptly obtains rennin; The nitrogen content of Semen Maydis powder is 1.075% (quality) in the step 1, and the nitrogen content of nisin fermentation waste solution is 0.24% (quality), and the nitrogen content of textured soybean protein is 8% (quality); The fungi count of the rice black root Mucor seed liquor that makes in the step 2 is 2～6 * 10
6Cfu/ml; The Intake Quantity of substratum is 20% of a triangular flask volume in the step 3.
The rennin of this embodiment preparation is 97.5IMCU/ml through the detection vigor.
Claims (5)
1. utilize nisin fermentation waste solution to make the method for partial nitrogen source fermenting and producing rennin; It is characterized in that utilizing nisin fermentation waste solution to make the method for partial nitrogen source fermenting and producing rennin, carry out according to the following steps: one, culture medium preparation: NaH
2PO
4, Na
2HPO
4, glucose, alpha-amylase, tween 80, Semen Maydis powder, W-Gum, nisin fermentation waste solution and textured soybean protein be dissolved in the water preparing culture medium, wherein NaH
2PO
4: 3.25g/L, Na
2HPO
4: 3.11g/L, glucose: 2.14g/L, alpha-amylase: 20g/L, tween 80: 0.13g/L, Semen Maydis powder: 30～80g/L, W-Gum: 0～30g/L, nisin fermentation waste solution: 150～700g/L, textured soybean protein: 0～20g/L; The use mass concentration is 5%～6% sodium hydroxide accent pH to 6.7～6.8, sterilization 30min under 0.14MPa pressure; Two, rice black root Mucor (
Rhizomucor miehei) preparation of seed liquor: with rice black root Mucor by volume 4% inoculum size be inoculated in the seed culture medium, shaking table is cultivated 20～24h under 37～38 ℃ of conditions, the rotating speed of shaking table is 260rpm; Three, inoculation: prepared culture medium in the step 1 is packed in the triangular flask of 250ml, with rice black root Mucor seed liquor by volume 2.5% inoculum size be inoculated in the triangular flask; Four, cultivate: triangular flask is placed down on the shaking table in 37～38 ℃ of conditions cultivates, the rotating speed of shaking table is 260～280rpm, cultivates 96 hours, promptly obtains rennin; The mass percentage content of the nitrogen content of Semen Maydis powder is 1.075% in the step 1, and the mass percentage content of the nitrogen content of nisin fermentation waste solution is 0.24%, and the mass percentage content of the nitrogen content of textured soybean protein is 8%; The fungi count of the rice black root Mucor seed liquor that makes in the step 2 is 2～6 * 10
6Cfu/ml; The Intake Quantity of substratum is 20% of a triangular flask volume in the step 3; Rice black root Mucor in the step 2 (
Rhizomucor miehei) being preserved in Chinese common micro-organisms culture presevation administrative center, culture presevation is numbered AS 3.4960.
2. the method for utilizing nisin fermentation waste solution to make the partial nitrogen source fermenting and producing rennin according to claim 1 is characterized in that the Semen Maydis powder concentration that adds in the step 1 is 35～70g/L.
3. the method for utilizing nisin fermentation waste solution to make the partial nitrogen source fermenting and producing rennin according to claim 2 is characterized in that the nisin fermentation waste solution concentration that adds in the step 1 is 200～600g/L.
4. according to claim 1, the 2 or 3 described methods of utilizing nisin fermentation waste solution to make the partial nitrogen source fermenting and producing rennin, it is characterized in that the nisin fermentation waste solution concentration that adds in the step 1 is 300～500g/L.
5. the method for utilizing nisin fermentation waste solution to make the partial nitrogen source fermenting and producing rennin according to claim 4 is characterized in that the textured soybean protein's concentration that adds in the step 1 is 5～15g/L.
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| 吴进菊等.中国曲中凝乳酶高产菌株的筛选及产酶条件的研究.《中国食品学报》.2009,(第01期), * |
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