CN1361278A - Detersile proteinase and its production process and usage and microbe producing the said proteinase - Google Patents
Detersile proteinase and its production process and usage and microbe producing the said proteinase Download PDFInfo
- Publication number
- CN1361278A CN1361278A CN 00120641 CN00120641A CN1361278A CN 1361278 A CN1361278 A CN 1361278A CN 00120641 CN00120641 CN 00120641 CN 00120641 A CN00120641 A CN 00120641A CN 1361278 A CN1361278 A CN 1361278A
- Authority
- CN
- China
- Prior art keywords
- enzyme
- sumizyme
- skin
- under
- immersion
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
- 108091005804 Peptidases Proteins 0.000 title claims abstract description 80
- 102000035195 Peptidases Human genes 0.000 title claims abstract description 80
- 235000019833 protease Nutrition 0.000 title abstract description 7
- 238000004519 manufacturing process Methods 0.000 title description 8
- 230000000694 effects Effects 0.000 claims abstract description 56
- 238000000034 method Methods 0.000 claims abstract description 52
- 230000008569 process Effects 0.000 claims abstract description 26
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 10
- 241000193830 Bacillus <bacterium> Species 0.000 claims abstract description 7
- 102000004169 proteins and genes Human genes 0.000 claims abstract description 6
- 108090000623 proteins and genes Proteins 0.000 claims abstract description 6
- 108090000765 processed proteins & peptides Proteins 0.000 claims abstract 3
- 102000004190 Enzymes Human genes 0.000 claims description 118
- 108090000790 Enzymes Proteins 0.000 claims description 118
- 230000035617 depilation Effects 0.000 claims description 60
- 238000007654 immersion Methods 0.000 claims description 30
- 239000010985 leather Substances 0.000 claims description 29
- CDBYLPFSWZWCQE-UHFFFAOYSA-L Sodium Carbonate Chemical compound [Na+].[Na+].[O-]C([O-])=O CDBYLPFSWZWCQE-UHFFFAOYSA-L 0.000 claims description 18
- 239000007788 liquid Substances 0.000 claims description 15
- 244000005700 microbiome Species 0.000 claims description 13
- 241000194103 Bacillus pumilus Species 0.000 claims description 12
- 238000010790 dilution Methods 0.000 claims description 9
- 239000012895 dilution Substances 0.000 claims description 9
- 238000002360 preparation method Methods 0.000 claims description 9
- 235000017550 sodium carbonate Nutrition 0.000 claims description 9
- 229910000029 sodium carbonate Inorganic materials 0.000 claims description 9
- 239000000243 solution Substances 0.000 claims description 8
- 238000002791 soaking Methods 0.000 claims description 7
- 238000005516 engineering process Methods 0.000 claims description 6
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 claims description 3
- DEFVIWRASFVYLL-UHFFFAOYSA-N ethylene glycol bis(2-aminoethyl)tetraacetic acid Chemical compound OC(=O)CN(CC(O)=O)CCOCCOCCN(CC(O)=O)CC(O)=O DEFVIWRASFVYLL-UHFFFAOYSA-N 0.000 claims description 3
- 238000002415 sodium dodecyl sulfate polyacrylamide gel electrophoresis Methods 0.000 claims description 3
- 108010009736 Protein Hydrolysates Proteins 0.000 claims description 2
- QWVGKYWNOKOFNN-UHFFFAOYSA-N o-cresol Chemical compound CC1=CC=CC=C1O QWVGKYWNOKOFNN-UHFFFAOYSA-N 0.000 claims description 2
- 239000003531 protein hydrolysate Substances 0.000 claims description 2
- 210000003491 skin Anatomy 0.000 claims 4
- 230000002951 depilatory effect Effects 0.000 claims 2
- 206010040914 Skin reaction Diseases 0.000 claims 1
- WWPPGEJAMWSVJJ-UHFFFAOYSA-N [Na].[K].[K] Chemical compound [Na].[K].[K] WWPPGEJAMWSVJJ-UHFFFAOYSA-N 0.000 claims 1
- 210000003780 hair follicle Anatomy 0.000 claims 1
- 230000035772 mutation Effects 0.000 claims 1
- 229920001184 polypeptide Polymers 0.000 claims 1
- 102000004196 processed proteins & peptides Human genes 0.000 claims 1
- 230000035483 skin reaction Effects 0.000 claims 1
- 231100000430 skin reaction Toxicity 0.000 claims 1
- 229920001436 collagen Polymers 0.000 abstract description 14
- 102000008186 Collagen Human genes 0.000 abstract description 13
- 108010035532 Collagen Proteins 0.000 abstract description 13
- 238000006243 chemical reaction Methods 0.000 abstract description 10
- 241001494479 Pecora Species 0.000 abstract description 2
- 230000002255 enzymatic effect Effects 0.000 abstract 2
- 229940088598 enzyme Drugs 0.000 description 107
- 210000004209 hair Anatomy 0.000 description 29
- 239000004365 Protease Substances 0.000 description 23
- 235000019419 proteases Nutrition 0.000 description 23
- 238000012360 testing method Methods 0.000 description 18
- 108010019160 Pancreatin Proteins 0.000 description 15
- 229940055695 pancreatin Drugs 0.000 description 15
- 210000003746 feather Anatomy 0.000 description 10
- BDAGIHXWWSANSR-UHFFFAOYSA-N methanoic acid Natural products OC=O BDAGIHXWWSANSR-UHFFFAOYSA-N 0.000 description 10
- 238000005406 washing Methods 0.000 description 10
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 9
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 description 8
- UIIMBOGNXHQVGW-UHFFFAOYSA-M Sodium bicarbonate Chemical compound [Na+].OC([O-])=O UIIMBOGNXHQVGW-UHFFFAOYSA-M 0.000 description 8
- 230000008859 change Effects 0.000 description 8
- 239000003112 inhibitor Substances 0.000 description 7
- VYZAMTAEIAYCRO-UHFFFAOYSA-N Chromium Chemical compound [Cr] VYZAMTAEIAYCRO-UHFFFAOYSA-N 0.000 description 6
- OSWFIVFLDKOXQC-UHFFFAOYSA-N 4-(3-methoxyphenyl)aniline Chemical compound COC1=CC=CC(C=2C=CC(N)=CC=2)=C1 OSWFIVFLDKOXQC-UHFFFAOYSA-N 0.000 description 5
- 235000019687 Lamb Nutrition 0.000 description 5
- 230000001580 bacterial effect Effects 0.000 description 5
- 210000001217 buttock Anatomy 0.000 description 5
- 239000003795 chemical substances by application Substances 0.000 description 5
- 235000019253 formic acid Nutrition 0.000 description 5
- 150000003839 salts Chemical class 0.000 description 5
- 239000006228 supernatant Substances 0.000 description 5
- NLXLAEXVIDQMFP-UHFFFAOYSA-N Ammonia chloride Chemical compound [NH4+].[Cl-] NLXLAEXVIDQMFP-UHFFFAOYSA-N 0.000 description 4
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 4
- 108010076119 Caseins Proteins 0.000 description 4
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 description 4
- 229910000147 aluminium phosphate Inorganic materials 0.000 description 4
- 239000000872 buffer Substances 0.000 description 4
- 229910052799 carbon Inorganic materials 0.000 description 4
- 239000005018 casein Substances 0.000 description 4
- BECPQYXYKAMYBN-UHFFFAOYSA-N casein, tech. Chemical compound NCCCCC(C(O)=O)N=C(O)C(CC(O)=O)N=C(O)C(CCC(O)=N)N=C(O)C(CC(C)C)N=C(O)C(CCC(O)=O)N=C(O)C(CC(O)=O)N=C(O)C(CCC(O)=O)N=C(O)C(C(C)O)N=C(O)C(CCC(O)=N)N=C(O)C(CCC(O)=N)N=C(O)C(CCC(O)=N)N=C(O)C(CCC(O)=O)N=C(O)C(CCC(O)=O)N=C(O)C(COP(O)(O)=O)N=C(O)C(CCC(O)=N)N=C(O)C(N)CC1=CC=CC=C1 BECPQYXYKAMYBN-UHFFFAOYSA-N 0.000 description 4
- 235000021240 caseins Nutrition 0.000 description 4
- 238000002474 experimental method Methods 0.000 description 4
- YBYRMVIVWMBXKQ-UHFFFAOYSA-N phenylmethanesulfonyl fluoride Chemical compound FS(=O)(=O)CC1=CC=CC=C1 YBYRMVIVWMBXKQ-UHFFFAOYSA-N 0.000 description 4
- 238000005554 pickling Methods 0.000 description 4
- FGIUAXJPYTZDNR-UHFFFAOYSA-N potassium nitrate Chemical compound [K+].[O-][N+]([O-])=O FGIUAXJPYTZDNR-UHFFFAOYSA-N 0.000 description 4
- 235000018102 proteins Nutrition 0.000 description 4
- 235000017557 sodium bicarbonate Nutrition 0.000 description 4
- 229910000030 sodium bicarbonate Inorganic materials 0.000 description 4
- 239000000126 substance Substances 0.000 description 4
- 239000000758 substrate Substances 0.000 description 4
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 3
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 3
- UCKMPCXJQFINFW-UHFFFAOYSA-N Sulphide Chemical compound [S-2] UCKMPCXJQFINFW-UHFFFAOYSA-N 0.000 description 3
- BFNBIHQBYMNNAN-UHFFFAOYSA-N ammonium sulfate Chemical compound N.N.OS(O)(=O)=O BFNBIHQBYMNNAN-UHFFFAOYSA-N 0.000 description 3
- 229910052921 ammonium sulfate Inorganic materials 0.000 description 3
- 235000011130 ammonium sulphate Nutrition 0.000 description 3
- 239000007853 buffer solution Substances 0.000 description 3
- 239000008103 glucose Substances 0.000 description 3
- 230000007062 hydrolysis Effects 0.000 description 3
- 238000006460 hydrolysis reaction Methods 0.000 description 3
- 230000003301 hydrolyzing effect Effects 0.000 description 3
- 239000000203 mixture Substances 0.000 description 3
- 239000012466 permeate Substances 0.000 description 3
- 238000000746 purification Methods 0.000 description 3
- 239000002994 raw material Substances 0.000 description 3
- 235000011121 sodium hydroxide Nutrition 0.000 description 3
- GRVFOGOEDUUMBP-UHFFFAOYSA-N sodium sulfide (anhydrous) Chemical compound [Na+].[Na+].[S-2] GRVFOGOEDUUMBP-UHFFFAOYSA-N 0.000 description 3
- OWEGMIWEEQEYGQ-UHFFFAOYSA-N 100676-05-9 Natural products OC1C(O)C(O)C(CO)OC1OCC1C(O)C(O)C(O)C(OC2C(OC(O)C(O)C2O)CO)O1 OWEGMIWEEQEYGQ-UHFFFAOYSA-N 0.000 description 2
- 241000894006 Bacteria Species 0.000 description 2
- 102000029816 Collagenase Human genes 0.000 description 2
- 108060005980 Collagenase Proteins 0.000 description 2
- BWGNESOTFCXPMA-UHFFFAOYSA-N Dihydrogen disulfide Chemical compound SS BWGNESOTFCXPMA-UHFFFAOYSA-N 0.000 description 2
- 101000925662 Enterobacteria phage PRD1 Endolysin Proteins 0.000 description 2
- 229930091371 Fructose Natural products 0.000 description 2
- 239000005715 Fructose Substances 0.000 description 2
- RFSUNEUAIZKAJO-ARQDHWQXSA-N Fructose Chemical compound OC[C@H]1O[C@](O)(CO)[C@@H](O)[C@@H]1O RFSUNEUAIZKAJO-ARQDHWQXSA-N 0.000 description 2
- 244000068988 Glycine max Species 0.000 description 2
- 235000010469 Glycine max Nutrition 0.000 description 2
- GUBGYTABKSRVRQ-PICCSMPSSA-N Maltose Natural products O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@@H](CO)OC(O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-PICCSMPSSA-N 0.000 description 2
- 101150071716 PCSK1 gene Proteins 0.000 description 2
- 102000012479 Serine Proteases Human genes 0.000 description 2
- 108010022999 Serine Proteases Proteins 0.000 description 2
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 description 2
- 229930006000 Sucrose Natural products 0.000 description 2
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 2
- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 description 2
- 239000003513 alkali Substances 0.000 description 2
- 235000019270 ammonium chloride Nutrition 0.000 description 2
- VBIXEXWLHSRNKB-UHFFFAOYSA-N ammonium oxalate Chemical compound [NH4+].[NH4+].[O-]C(=O)C([O-])=O VBIXEXWLHSRNKB-UHFFFAOYSA-N 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- 239000011575 calcium Substances 0.000 description 2
- 150000001768 cations Chemical class 0.000 description 2
- 229960002424 collagenase Drugs 0.000 description 2
- 239000006071 cream Substances 0.000 description 2
- 238000013016 damping Methods 0.000 description 2
- 238000005238 degreasing Methods 0.000 description 2
- MNNHAPBLZZVQHP-UHFFFAOYSA-N diammonium hydrogen phosphate Chemical compound [NH4+].[NH4+].OP([O-])([O-])=O MNNHAPBLZZVQHP-UHFFFAOYSA-N 0.000 description 2
- 238000004043 dyeing Methods 0.000 description 2
- 238000011156 evaluation Methods 0.000 description 2
- 239000000706 filtrate Substances 0.000 description 2
- 239000012467 final product Substances 0.000 description 2
- 235000021323 fish oil Nutrition 0.000 description 2
- 239000012530 fluid Substances 0.000 description 2
- 238000007689 inspection Methods 0.000 description 2
- 239000006210 lotion Substances 0.000 description 2
- 230000000813 microbial effect Effects 0.000 description 2
- 231100000350 mutagenesis Toxicity 0.000 description 2
- 235000019198 oils Nutrition 0.000 description 2
- 239000004323 potassium nitrate Substances 0.000 description 2
- 235000010333 potassium nitrate Nutrition 0.000 description 2
- 239000000843 powder Substances 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- 230000002797 proteolythic effect Effects 0.000 description 2
- 239000002265 redox agent Substances 0.000 description 2
- 238000000926 separation method Methods 0.000 description 2
- 239000010865 sewage Substances 0.000 description 2
- 239000001632 sodium acetate Substances 0.000 description 2
- 229960004249 sodium acetate Drugs 0.000 description 2
- 235000017281 sodium acetate Nutrition 0.000 description 2
- 239000005720 sucrose Substances 0.000 description 2
- 239000002699 waste material Substances 0.000 description 2
- PAWQVTBBRAZDMG-UHFFFAOYSA-N 2-(3-bromo-2-fluorophenyl)acetic acid Chemical compound OC(=O)CC1=CC=CC(Br)=C1F PAWQVTBBRAZDMG-UHFFFAOYSA-N 0.000 description 1
- 206010000372 Accident at work Diseases 0.000 description 1
- GUBGYTABKSRVRQ-XLOQQCSPSA-N Alpha-Lactose Chemical compound O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@@H](CO)O[C@H](O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-XLOQQCSPSA-N 0.000 description 1
- 239000004382 Amylase Substances 0.000 description 1
- 102000013142 Amylases Human genes 0.000 description 1
- 108010065511 Amylases Proteins 0.000 description 1
- 239000000120 Artificial Saliva Substances 0.000 description 1
- BTBUEUYNUDRHOZ-UHFFFAOYSA-N Borate Chemical compound [O-]B([O-])[O-] BTBUEUYNUDRHOZ-UHFFFAOYSA-N 0.000 description 1
- 241000283690 Bos taurus Species 0.000 description 1
- 241000283707 Capra Species 0.000 description 1
- FBPFZTCFMRRESA-KVTDHHQDSA-N D-Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 description 1
- 239000004375 Dextrin Substances 0.000 description 1
- 229920001353 Dextrin Polymers 0.000 description 1
- 108010010803 Gelatin Proteins 0.000 description 1
- 108090000288 Glycoproteins Proteins 0.000 description 1
- 102000003886 Glycoproteins Human genes 0.000 description 1
- 238000003794 Gram staining Methods 0.000 description 1
- 101001091385 Homo sapiens Kallikrein-6 Proteins 0.000 description 1
- PMMYEEVYMWASQN-DMTCNVIQSA-N Hydroxyproline Chemical compound O[C@H]1CN[C@H](C(O)=O)C1 PMMYEEVYMWASQN-DMTCNVIQSA-N 0.000 description 1
- DGAQECJNVWCQMB-PUAWFVPOSA-M Ilexoside XXIX Chemical compound C[C@@H]1CC[C@@]2(CC[C@@]3(C(=CC[C@H]4[C@]3(CC[C@@H]5[C@@]4(CC[C@@H](C5(C)C)OS(=O)(=O)[O-])C)C)[C@@H]2[C@]1(C)O)C)C(=O)O[C@H]6[C@@H]([C@H]([C@@H]([C@H](O6)CO)O)O)O.[Na+] DGAQECJNVWCQMB-PUAWFVPOSA-M 0.000 description 1
- 102100034866 Kallikrein-6 Human genes 0.000 description 1
- LKDRXBCSQODPBY-AMVSKUEXSA-N L-(-)-Sorbose Chemical compound OCC1(O)OC[C@H](O)[C@@H](O)[C@@H]1O LKDRXBCSQODPBY-AMVSKUEXSA-N 0.000 description 1
- OUYCCCASQSFEME-QMMMGPOBSA-N L-tyrosine Chemical compound OC(=O)[C@@H](N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-QMMMGPOBSA-N 0.000 description 1
- GUBGYTABKSRVRQ-QKKXKWKRSA-N Lactose Natural products OC[C@H]1O[C@@H](O[C@H]2[C@H](O)[C@@H](O)C(O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@H]1O GUBGYTABKSRVRQ-QKKXKWKRSA-N 0.000 description 1
- 239000004367 Lipase Substances 0.000 description 1
- 102000004882 Lipase Human genes 0.000 description 1
- 108090001060 Lipase Proteins 0.000 description 1
- 229930195725 Mannitol Natural products 0.000 description 1
- 102000005741 Metalloproteases Human genes 0.000 description 1
- 108010006035 Metalloproteases Proteins 0.000 description 1
- VZUNGTLZRAYYDE-UHFFFAOYSA-N N-methyl-N'-nitro-N-nitrosoguanidine Chemical compound O=NN(C)C(=N)N[N+]([O-])=O VZUNGTLZRAYYDE-UHFFFAOYSA-N 0.000 description 1
- 102000035092 Neutral proteases Human genes 0.000 description 1
- 108091005507 Neutral proteases Proteins 0.000 description 1
- WCUXLLCKKVVCTQ-UHFFFAOYSA-M Potassium chloride Chemical compound [Cl-].[K+] WCUXLLCKKVVCTQ-UHFFFAOYSA-M 0.000 description 1
- 238000003457 Shi epoxidation reaction Methods 0.000 description 1
- 229920002472 Starch Polymers 0.000 description 1
- 240000008042 Zea mays Species 0.000 description 1
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 description 1
- 235000002017 Zea mays subsp mays Nutrition 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 239000013543 active substance Substances 0.000 description 1
- SRBFZHDQGSBBOR-QMKXCQHVSA-N alpha-L-arabinopyranose Chemical compound O[C@H]1CO[C@@H](O)[C@H](O)[C@H]1O SRBFZHDQGSBBOR-QMKXCQHVSA-N 0.000 description 1
- 235000019418 amylase Nutrition 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 1
- GUBGYTABKSRVRQ-QUYVBRFLSA-N beta-maltose Chemical compound OC[C@H]1O[C@H](O[C@H]2[C@H](O)[C@@H](O)[C@H](O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@@H]1O GUBGYTABKSRVRQ-QUYVBRFLSA-N 0.000 description 1
- 229910021538 borax Inorganic materials 0.000 description 1
- 238000005282 brightening Methods 0.000 description 1
- 229910052791 calcium Inorganic materials 0.000 description 1
- 159000000007 calcium salts Chemical class 0.000 description 1
- 229940041514 candida albicans extract Drugs 0.000 description 1
- 239000004202 carbamide Substances 0.000 description 1
- 235000013877 carbamide Nutrition 0.000 description 1
- 108010079058 casein hydrolysate Proteins 0.000 description 1
- 239000003518 caustics Substances 0.000 description 1
- 238000004140 cleaning Methods 0.000 description 1
- 230000000536 complexating effect Effects 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 235000005822 corn Nutrition 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 230000000593 degrading effect Effects 0.000 description 1
- 235000019425 dextrin Nutrition 0.000 description 1
- -1 dipotassium arsenic Chemical compound 0.000 description 1
- PMMYEEVYMWASQN-UHFFFAOYSA-N dl-hydroxyproline Natural products OC1C[NH2+]C(C([O-])=O)C1 PMMYEEVYMWASQN-UHFFFAOYSA-N 0.000 description 1
- 230000002500 effect on skin Effects 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 238000003912 environmental pollution Methods 0.000 description 1
- 238000006911 enzymatic reaction Methods 0.000 description 1
- 238000011049 filling Methods 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- 238000004108 freeze drying Methods 0.000 description 1
- 238000005227 gel permeation chromatography Methods 0.000 description 1
- 239000008273 gelatin Substances 0.000 description 1
- 229920000159 gelatin Polymers 0.000 description 1
- 235000019322 gelatine Nutrition 0.000 description 1
- 235000011852 gelatine desserts Nutrition 0.000 description 1
- 230000005484 gravity Effects 0.000 description 1
- 229960002591 hydroxyproline Drugs 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 230000000266 injurious effect Effects 0.000 description 1
- 229910052500 inorganic mineral Inorganic materials 0.000 description 1
- 238000004255 ion exchange chromatography Methods 0.000 description 1
- 239000008101 lactose Substances 0.000 description 1
- 235000019421 lipase Nutrition 0.000 description 1
- 230000002101 lytic effect Effects 0.000 description 1
- 159000000003 magnesium salts Chemical class 0.000 description 1
- 239000000594 mannitol Substances 0.000 description 1
- 235000010355 mannitol Nutrition 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 239000012092 media component Substances 0.000 description 1
- 238000009992 mercerising Methods 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- 229910021645 metal ion Inorganic materials 0.000 description 1
- 238000001465 metallisation Methods 0.000 description 1
- 235000010755 mineral Nutrition 0.000 description 1
- 239000011707 mineral Substances 0.000 description 1
- 231100000219 mutagenic Toxicity 0.000 description 1
- 230000003505 mutagenic effect Effects 0.000 description 1
- 230000007935 neutral effect Effects 0.000 description 1
- 238000006386 neutralization reaction Methods 0.000 description 1
- 229910017464 nitrogen compound Inorganic materials 0.000 description 1
- 150000002830 nitrogen compounds Chemical class 0.000 description 1
- QJGQUHMNIGDVPM-UHFFFAOYSA-N nitrogen group Chemical group [N] QJGQUHMNIGDVPM-UHFFFAOYSA-N 0.000 description 1
- 239000006916 nutrient agar Substances 0.000 description 1
- 235000015097 nutrients Nutrition 0.000 description 1
- 150000002897 organic nitrogen compounds Chemical class 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 150000003016 phosphoric acids Chemical class 0.000 description 1
- 229920000191 poly(N-vinyl pyrrolidone) Polymers 0.000 description 1
- 239000011148 porous material Substances 0.000 description 1
- 239000001103 potassium chloride Substances 0.000 description 1
- 235000011164 potassium chloride Nutrition 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- 238000001556 precipitation Methods 0.000 description 1
- 238000002203 pretreatment Methods 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 238000002975 protease activity determination Methods 0.000 description 1
- 230000017854 proteolysis Effects 0.000 description 1
- 238000012797 qualification Methods 0.000 description 1
- 230000035484 reaction time Effects 0.000 description 1
- 230000000630 rising effect Effects 0.000 description 1
- 238000005096 rolling process Methods 0.000 description 1
- 108010028349 saliva Orthana Proteins 0.000 description 1
- 238000013341 scale-up Methods 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 230000011218 segmentation Effects 0.000 description 1
- 210000000697 sensory organ Anatomy 0.000 description 1
- 238000012807 shake-flask culturing Methods 0.000 description 1
- 239000011734 sodium Substances 0.000 description 1
- 229910052708 sodium Inorganic materials 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
- HLBBKKJFGFRGMU-UHFFFAOYSA-M sodium formate Chemical compound [Na+].[O-]C=O HLBBKKJFGFRGMU-UHFFFAOYSA-M 0.000 description 1
- 239000004328 sodium tetraborate Substances 0.000 description 1
- 235000010339 sodium tetraborate Nutrition 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 210000001082 somatic cell Anatomy 0.000 description 1
- 230000002269 spontaneous effect Effects 0.000 description 1
- 238000001694 spray drying Methods 0.000 description 1
- 238000010186 staining Methods 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 230000009897 systematic effect Effects 0.000 description 1
- FGMPLJWBKKVCDB-UHFFFAOYSA-N trans-L-hydroxy-proline Natural products ON1CCCC1C(O)=O FGMPLJWBKKVCDB-UHFFFAOYSA-N 0.000 description 1
- OUYCCCASQSFEME-UHFFFAOYSA-N tyrosine Natural products OC(=O)C(N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-UHFFFAOYSA-N 0.000 description 1
- 210000002268 wool Anatomy 0.000 description 1
- 239000012138 yeast extract Substances 0.000 description 1
Images
Landscapes
- Treatment And Processing Of Natural Fur Or Leather (AREA)
Abstract
The present invention is one unknown strain of bacillus (in the storage number of CGMCC No.0518) and obtain the proteinase with said strain or its mutant. The said proteinase can hydrolyze protein and peptide; has its highest enzymatic activity at 50 deg.c and pH 9.6; stable enzymatic activity at 50 deg.c and pH 6-11, optimal reaction temperature of 50 deg.c; and molecular weight of 40,000 Daltons. The proteinase of the present invention has excellent detersile effect on pig skin, sheep skin and ox skin without damage to skin collagen. It may be also used in the water soaked and softening process is tanning.
Description
The present invention relates to a kind of Sumizyme MP and production method thereof, its in soaking of hide or skin, enzyme unhairing and soft and soggy metallization processes application and produce the microorganism of this proteolytic enzyme.
In past 15 years, the center of gravity of world's tanning industry has been shifted to the Asia gradually.Chinese leather output has increased octuple, and wherein the pigskin annual production reaches 100,000,000, occupy first place in the world, and 5,000 ten thousand on ox-hide, sheepskin is more than 7,000 ten thousand.Leather in 1998 and goods foreign exchange earning reach 9,700,000,000 dollars, are China's light industry first place of earning foreign exchange.China has also produced the serious environmental pollution when obtaining economic benefit.Along with developing rapidly of tanning industry, the degree of this pollution will be serious day by day.
The most serious source of pollution of tanning industry are mainly from the sulfide in traditional dehairing process.In order to solve sulphide staining, the researchist had once done many-sided effort.Be to increase facility investment on the one hand, sulfide sewage is handled, make it to reach the emission standard (pollution, aftertreatment earlier) of regulation.This method is expensive big, has increased cost.Administer the sewage except existing more than 200 fairly large tame tanneries have " condition ", all the other country-wide thousands of little factories can't handle sour water at all.Facts have proved that efficient ways replaces grey alkaline process depilation with enzyme unhairing exactly, is about to source of pollution and eliminates among production process.
China begins to test the enzyme process dehairing process from nineteen sixty-eight, popularizes in an all-round way in China by 1973, because some practical problemss that occur in promoting are not are not conscientiously researched and solved, makes enzyme unhairing fail to persevere.The most important reason that this situation occurs is that loose side appears in finished leather easily behind the enzyme unhairing, and is with low quality because of microbial proteinous zymin cost height, and safety coefficient is low, if any also causing mashed skin accidentally, leads to industrial accident.Trace it to its cause, mainly contain following two aspects:
1, the unhairing protease preparation variety is few, selects suitable zymin to come into operation and is restricted.Protease preparation commonly used has 166, AS1398,3942 neutral proteases and 209,2709 Sumizyme MPs.Wherein, have only 166 and 209 to be the special screenings of losing hair or feathers that are, other are several all to be the industry introduced variety.Therefore, the zymin kind is few, is difficult to satisfy the special requirement of process hides depilation.Add and respectively use the difference of factory at aspects such as technical qualification, management, equipment and raw materials, the kind of enzyme is single, causes no suitable choice.Technically, also have only the pigskin depilation to reach a standard substantially.This is one of enzyme unhairing reason of being difficult to promote the use of for a long time.
2, depilation zymin composition complexity, unstable properties, poor specificity.When depilation, unhairing protease energy protein hydrolysate, yet a lot of because of the proteolytic enzyme kind in the preparation, each proteolytic enzyme respectively has its different specificity again.They are very different to the hydrolysis ability that various structures effect substrate far from it shows.Present used protease preparation only is the mixture of the various active substances that produced in its metabolic process of microorganism.This crude zyme preparation contains other multiple protein enzymes and various assorted enzyme (lipase, saccharifying enzyme, amylase) simultaneously again often based on a kind of proteolytic enzyme.They not only have the glycoprotein lytic enzyme of the energy hydrolysis mucoid relevant with depilation and Saliva Orthana and so on, and also have the elastoser relevant with soft and soggyization.In addition, also contain the energy loss collagen lytic enzyme of collagen of being injurious to life, thereby the main raw material(s)-collagen that makes leather produce usually suffers to damage in various degree, influence resultant leather quality.
The enzyme unhairing of carrying out at present is actually the multiple enzymatic reaction that the multiple protein enzyme acts on multiple proteins substrate in the rawhide, but, owing to contain a large amount of collagen lytic enzymes in the thick enzyme (during the depilation terminal point, hydroxyproline content has remarkable increase in the solution when depilation begins), make the collagen protein that keep be subjected to very big loss.The proteolysis result of this " comprehensive ", the as easy as rolling off a log defect such as finished leather loose side that occur cause resultant leather quality inferior.Obviously, this expense height, quality product depilating method low, inconvenient operation are difficult to adhere to.
Make enzyme unhairing replace the depilation of grey alkaline process and make it to be applied, can the key variety of problems that exists with zymin that is to lose hair or feathers be resolved.
(application number: 95197015.1) relate to the method for carrying out dehairing of hides with enzyme, it may further comprise the steps Chinese patent: 1. rawhide is soaked; 2. will lead immersion through the rawhide that soaks; 3. by adding entry, under the effect of mechanical effect and at least a proteolytic enzyme, the rawhide through above-mentioned processing being lost hair or feathers.1. in 3., rawhide has passed through the effect of once at least a disulfide protein redox agent at least in step, and this method has following shortcoming:
1, this patent is lost hair or feathers with any proteolytic ferment or their mixture, and the Collagenase of damage rawhide collagen also belongs to proteolytic ferment, if (application number: method 95197015.1) is carried out dehairing of hides with Collagenase by Chinese patent, main raw material(s)-collagen that leather is produced is damaged, even caused mashed skin accident.
2, operation easier is bigger.Example: if the optimum pH that these two kinds of enzymes of disulfide protein redox agent and proteolytic enzyme show respectively is very different, they just must add in succession, and will carry out pH regulator.
Chinese patent (application number: 95193076.1) also relate to enzyme unhairing, but use the hair removal effect of Sumizyme MP of its patent very bad; Even perhaps depilation is fully, also rawhide is become the lump of block rubber, lose productive value.
The purpose of this invention is to provide a kind of proteolytic enzyme that can be used for immersion technology, dehairing process and the softening process of tanning industry, the method for making this proteolytic enzyme, this proteolytic enzyme in tanning industry application and produce the microorganism of this proteolytic enzyme, to increase the specificity and the unicity of proteolytic enzyme, do not damage rawhide collagen when making enzyme unhairing, simplify the operation, guarantee resultant leather quality.
Invention is described
The contriver finds that the proteolytic enzyme BP-Ap42 by the bacterial strain UN-31-C-42 generation of the bacillus pumilus (Bacillus pumilus) of bacillus is a kind of Sumizyme MP.The enzyme unhairing of only using this enzyme to carry out shows that this enzyme has extraordinary hair removal effect, and does not damage collagen, and short and hair removal effect is better than AS1398 with depilation time of enzyme AS1398 proteolytic enzyme than routine.Simultaneously, this enzyme has good bating effect to leatherware, is better than the pancreatin that present softening process is used.Immersion technology before this enzyme also can be used for losing hair or feathers.
Therefore, the invention provides this new proteolytic enzyme, its microorganism of production method, purposes and this proteolytic enzyme of generation.Produce the microorganism of enzyme
The microorganism that produces enzyme of the present invention is a kind of and bacillus pumilus (Bacillus pumilus) closely-related bacterial strain UN-31-C-42 on bacteriology of bacillus.Bacteriology characteristic
Bacillus strain UN-31-C-42 related to the present invention has following bacteriology characteristic:
(a) form: genus bacillus
(b) gramstaining: feminine gender
(c) generation of spore: the positive
(d) shape of spore: ellipse garden
(e) movability: the positive
(f) to the reaction of oxygen: aerophil
(g) catalatic generation: the positive
(h) growth under anaerobic: feminine gender
(i) volt one general Er Shi reaction (VP) reaction: the positive
(j) Suan generation:
Glucose: feminine gender
Pectinose: the positive
Mannitol: feminine gender
(k) by the glucose aerogenesis: the positive
(l) liquification of gelatin: the positive
(m) degraded of starch: the positive
(n) can utilize ammonium oxalate, urea, ammonium chloride, ammonium sulfate, saltpetre and Secondary ammonium phosphate, and do not utilize ammonium nitrate.
(p) utilization of carbon source:
Can be carbon source with maltose, semi-lactosi, fructose and sucrose, can not be carbon source with lactose and sorbose.
According to the systematic bacteriology handbook (Vo1.2, William and Wilkins, 1986) of Bergey, preliminary evaluation is a bacillus pumilus, names UN-31-C-42.
In addition, the proteolytic enzyme improvements in productivity mutant (getting by spontaneous or induced mutation) that belongs to said bacterial strain UN-31-C-42 can be used as the bacterium that produces proteolytic enzyme of the present invention.In order to prepare these mutant, can use conventional method, for example by uviolizing or mutagenic compound such as N-methyl-N '-nitro-N-nitrosoguanidine (NTG) to the original strain induced mutation after, culture is inoculated into contains in the caseic nutrient agar.The mutant that selection has excellent productivity from the bacterium colony that forms bigger clear endless belt in periphery of bacterial colonies; Cooperation is measured protease activity by shake flat experiment, and the experiment etc. of losing hair or feathers, and obtains producing the microorganism strains of proteolytic enzyme of the present invention.Produce the method for proteolytic enzyme
Any substratum that can make the microorganism growth that produces proteolytic enzyme of the present invention and produce said proteolytic enzyme may be used to produce proteolytic enzyme of the present invention.
For example, can use glucose, fructose, maltose, sucrose, dextrin as carbon source, use inorganic nitrogen compound (as ammonium oxalate, ammonium chloride, saltpetre, Secondary ammonium phosphate etc.) or organic nitrogen compound (as casein, soybean cake, corn immersion liquid, analysis for soybean powder etc.) as nitrogenous source, to add mineral substance in addition, as phosphoric acid salt, sylvite, magnesium salts and calcium salt.The initial pH value of substratum is 7.5-10.In the present invention, culture is cultivated under aerobic condition, culture temperature is 37 ℃, and incubation time is 36 hours to 48 hours, can obtain the enzyme of maximum production.
Can carry out purifying to the culture that obtains by above process according to the separation and the purification process of routine.By salt precipitation protein, spray drying process, lyophilization etc., soluble salt is joined in the supernatant liquor or filtrate (said supernatant liquor and filtrate are to obtain by solid residue centrifugal or filtration somatic cells and substratum) of gained, can obtain proteolytic enzyme of the present invention.The purification process (as ion exchange chromatography and gel permeation chromatography) that can be used in combination other is further purified said enzyme.The contriver is the proteolytic enzyme called after BP-Ap42 of the present invention that is obtained by the way.The depilation activity of measuring protease activities and testing it by following method.
1, the mensuration of proteinase activity
1ml enzyme liquid is measured under the pH9.6 condition at 50 ℃, and the enzyme amount that the per minute caseinhydrolysate produces 1 μ g tyrosine is an enzyme activity unit (U).
2, the active test of losing hair or feathers
With borax one sodium hydrate buffer solution (pH9.6) enzyme liquid is transferred to 400U/ml, dress enzyme liquid 50mL in the 250mL triangular flask, (10 * 15cm) 3,37 ℃ of constant temperature shaking tables (rotating speed 160r/min) are gone up depilation, regularly observe the depilation situation to put into pig buttocks salt brined hide after the cleaning.The characteristic of proteolytic enzyme
The physics and the chemical property of the proteolytic enzyme that the present invention obtained are as follows:
1, substrate specificity
This proteolytic enzyme casein of degrading.
2, optimum pH:
With the damping fluid dilution enzyme liquid of different pH values, measure enzymic activity down for 50 ℃, the result is as shown in Figure 1.In the scope of pH6-9.6, enzyme work raises with pH, and is wherein more smooth at pH7-pH8.5 scope inner curve; In the pH10-11 scope, descend with the pH rising.Therefore optimum pH is 9.6.
3, optimal reactive temperature
With borax-sodium hydrate buffer solution (pH9.6) dilution enzyme liquid, under the differential responses temperature, measure enzyme and live.The result shows that the optimal reactive temperature of this enzyme is 50 ℃, surpasses 50 ℃ of enzymes reduce rapidly (seeing accompanying drawing 2) alive.
4, the influence of inhibitor
Used 6 kinds of different inhibitor, measured their influences this proteolytic enzyme.As 100%, living for the relative enzyme of residue in the enzyme work that adds the sample behind the inhibitor back of converting, the results are shown in table 1 with the sample enzyme work that do not add inhibitor.
Table 1 inhibitor is for the influence of fermented liquid neutral and alkali protease activity
Inhibitor dipotassium arsenic
EDTA EGTA PMSF DFP methyl-phenoxide
(working concentration is 1mM/L) sour sodium remains relative enzyme (%) 14.7 12.7 2.1 2.1 90.3 93.4 alive
The result shows that specificity suppresses the inhibitor PMSF of serine protease, and DFP can suppress this protease activities fully, shows that this proteolytic enzyme is a kind of serine protease.EDTA, EGTA also can suppress this enzyme, shows that this protease activities needs the participation of metal ion.
5, molecular weight
The molecular weight of this enzyme that records by the SDS-polyacrylamide gel electrophoresis is 40,000 dalton.
6, stability
At 4 ℃ of good stabilities.The using method of Sumizyme MP
1, the usage of enzyme during soaking of hide or skin: use wet salted hide, carry out skin and ease back.Enzyme dosage 50U/g during the ox-hide immersion, goatskin, lamb skin, pigskin enzyme dosage are 30U/g.Temperature is 18 ℃ in the drum, and liquor ratio 3 adds yellow soda ash and regulates pH10, and time 15h eases back.
2, the bank up usage of enzyme unhairing: wet salted hide eases back by the condition immersion of above-mentioned immersion; Be used to the enzyme unhairing of banking up behind the drip-dry water.Ox-hide, goatskin, lamb skin, the pigskin condition that is adopted in the enzyme unhairing process of banking up all is: 18 ℃ of temperature, and with the sodium carbonate solution dilution enzyme liquid of pH10, enzyme dosage is 100U/g, the 16h that banks up can reach good hair removal effect.
3, Sumizyme MP is to skin remollescent usage: through immersion, the skin behind depilation, liming, multiple ash, the deliming in drum, 38 ℃ of temperature, pH10, softening 2h, liquor ratio 2, ox-hide, goatskin, lamb skin remollescent enzyme dosage are 100U/g, and pigskin remollescent enzyme dosage is 150U/g.
Description of drawings
The optimal reaction pH value of Fig. 1, Sumizyme MP BP-Ap42;
Fig. 2, Sumizyme MP BP-Ap42 optimal reactive temperature;
The cultivation of embodiment 1, bacillus pumilus UN-31-C-42
Bacterial strain UN-31-C-42 is being contained casein 0.5%, yeast extract paste 0.3%, KH
2PO
40.1%, K
2HPO
40.2%, CaCl
20.1%, MgSO
4.7H
2In the substratum of O 0.02%,, nutrient solution 10000rpm is got supernatant liquor after centrifugal 10 minutes in 37 ℃ of shake-flask culture 48 hours.The enzyme of supernatant liquor is lived and is 496U/ml.The purifying of embodiment 2, Sumizyme MP BP-Ap42
The culture 10000rpm that will obtain in embodiment 1 gets supernatant liquor after centrifugal 10 minutes, with the ammonium sulfate of 20-70% saturation ratio this solution is carried out segmentation and saltouts.With the resolution of precipitate that obtains phosphoric acid buffer (pH6.8), and dialyse with this damping fluid at 20mM.Under pH6.8, this solution is joined in the CM-cellulofine C-52 post (Whatman company) then, use 20mM phosphoric acid buffer (pH6.8) wash-out that contains 0.1M NaCl.Enzyme peak alive is carried out lyophilize, and the dry powder that obtains is dissolved in the 20mM phosphoric acid buffer (pH6.8) again.This solution is joined in the Sepheadex G-75 post (Pharmacia company), use 20mM phosphoric acid buffer (pH6.8) wash-out that contains 0.1M Nacl, collect enzyme peak alive.This component is carried out the analysis of SDS-polyacrylamide gel electrophoresis, show that this enzyme is a single band.The water immersion test of embodiment 3, Sumizyme MP BP-Ap42
(1) at first, Sumizyme MP BP-Ap42 is measured with the AS1398 protease activity that is used to contrast, reaction substrate is casein (preparing with borate buffer solution), 40 ℃ of temperature of reaction, and reaction times 10min, its measurement result is shown in table 2.
-: do not survey enzymic activity
Obviously, the AS1398 protease activity that produce in Yunnan is higher than the Wuxi and produces proteolytic enzyme, thereby the AS1398 proteolytic enzyme of selecting for use Yunnan to produce during controlled trial.Enzyme concn determined when (2) skin soaked
Use wet salted hide, carry out the skin test that eases back, its condition of easing back is: protease concentration 10,30,50, and 70U/mL, temperature is 18 ℃ in the drum, and liquor ratio 3 adds yellow soda ash and regulates pH10, and time 15h eases back.
Protease concentration sees Table 3 to the influence of skin immersion effect.
Table 3 protease concentration is to the influence of skin immersion effect
Skin immersion effect+poor, ++ general, +++better
Table 3 is the result show, ox-hide 50U/mL, and all the other 3 kinds of skins just can make skin reach the requirement of quick immersion with the proteolytic enzyme of 30U/mL.
After the enzyme-added immersion, the water-filling degree of skin is higher, and the skin effect that eases back is more satisfactory.Wherein, the weightening finish that eases back of goatskin is than the highest (table 4).
Weightening finish is than (kilogram) before and after the immersion of table 4 wet salted hide
Optimum pH determined when enzyme unhairing test (1) the Sumizyme MP BP-Ap42 that banks up of embodiment 4, Sumizyme MP BP-Ap42 was used to bank up enzyme unhairing
In the present invention, adopt the enzyme unhairing of banking up.Depilation eases back by above-mentioned definite condition immersion with skin, and is stand-by behind the drip-dry water.
The condition that is adopted in the enzyme unhairing process of banking up is: 18 ℃ of temperature, use pH9, and 10,11 sodium carbonate solution is by the enzyme dilution enzyme liquid of every gram skin with 200U.
Different pH values see Table 5 to the influence of the enzyme unhairing effect of banking up.Under the pH10 condition, the hair removal effect of pigskin is poor slightly, and the hair removal effect of ox-hide, lamb skin and goatskin is all fine.
The depilation ability: ++ ++ touch promptly; +++heavily pushes away and can fall; ++ pull out and can fall; + pull at and can fall
-La does not fall (the following symbol of respectively showing is herewith shown) the determining of the suitableeest enzyme amount when (2) Sumizyme MP BP-Ap42 is used to bank up enzyme unhairing
The optimum amount of enzyme in order to obtain to bank up enzyme unhairing, the depilation condition of employing is: 18 ℃ of temperature, pH10, the 12-16h that banks up, enzyme working concentration are 50,100,200U/g.Enzyme concn sees Table 6 to the influence of the enzyme unhairing effect of banking up.
From the hair removal effect of 4 kinds of skins, with the 200U/g enzyme concn 16h that banks up, hair removal effect is fine.(3) BP-Ap42 and the comparison of AS1398 proteolytic enzyme in the enzyme unhairing of banking up
BP-Ap42 all uses 200U/g with the AS1398 proteolytic enzyme that is used for control experiment.The experiment with skin from the ridge line to cuing open.The depilation condition: 20 ℃ of room temperatures, 15-24h banks up; AS1398 reacts pH8; BP-Ap42 reacts pH10.
Table 7 BP-Ap42 and AS1398 proteolytic enzyme depilation simultaneous test
Revision test result shows: the depilation speed of BP-Ap42 is fast, and 15h gets final product; AS1398 generally wants about 24h.The pigskin hair removal effect of BP-Ap42 proteolytic enzyme is slightly poorer than the AS1398 enzyme; Both are similar to the depilation of lamb skin; To goatskin and ox-hide, the BP-Ap42 hair removal effect is good, and especially the hair removal effect of ox-hide is better.The pelt grain effect of this proteolytic enzyme depilation is pure white, careful than the skin of AS1398 depilation, softness, and the mercerising sense is also stronger.From every index of sense organ and feel evaluation, the BP-Ap42 hair removal effect is better than AS1398.
The scale-up of whole skin, can repeat the result of lab scale preferably by each 50.(4) depilation of buttocks part of pighide and goatskin neck test
Because the Maonans of buttocks part of pighide and goatskin neck is most in taking off, thereby before the enzyme unhairing of banking up, use earlier the BP-Ap42 pre-treatment.With the BP-Ap42 enzyme liquid bag buttocks 4h of 400U/g, and then lose hair or feathers with the BP-Ap42 proteolytic enzyme of 200U/g after the pigskin degreasing, it is poor so both to have reduced the position, helps the depilation of buttocks again.
After the goatskin immersion, be coated with the neck hair on the neck 3h of portion with the BP-Ap42 enzyme liquid of 400U/g, with the BP-Ap42 proteolytic enzyme depilation of 200U/g, the hair of whole skin takes off to the greatest extent fully behind the 15h again, and neck hair on the neck portion does not stay surplus mao, and the ridge line is not obvious.Embodiment 5, Sumizyme MP BP-Ap42 the determining of enzyme concn during to soft and soggyization of softening test (1) of skin
Through the skin behind immersion, depilation, liming, multiple ash, the deliming, temperature is 38 ℃ in the drum, reaction pH10, and softening 2h, enzyme dosage is 50,100,150,200U/g, liquor ratio 2.
Table 8 protease concentration influences the skin remollescent
Soft and soggyization effect :+relatively poor, ++ it is general, +++better test-results shows, the softening 150U/g that uses of pigskin, and ox-hide and sheepskin are softening to get final product with 100U/g.(2) comparison of the softening bark effect of BP-Ap42 and pancreatin
Pancreatin is identical with BP-Ap42 proteolytic enzyme consumption: pigskin 150U/g, cattle and sheep skin 100U/g; 38 ℃ of temperature, softening 2-3h (commentaries on classics stops combination), liquor ratio 2, pancreatin reaction pH8, BP-Ap42 reacts pH10.Comparing result is shown in table 9.
The soft and soggyization simultaneous test of table 9 BP-Ap42 proteolytic enzyme and pancreatin
The rerum natura of embodiment 6, enzyme unhairing and softening finished leather detects test
Carry out the physical and chemical inspection of leather finished product by kid shoes upper leather and pigskin shoes upper leather that different tests obtained, the results are shown in table 10 and table 11.
Table 10 sheepskin finished leather rerum natura detects
* the softening 4.BP-Ap42 depilation/BP-Ap42 of 1.AS1398 depilation/BP-Ap42 is softening
2.AS1398 the softening 5.BP-Ap42 depilation/pancreatin of depilation/pancreatin is softening
3.AS1398 depilation/not softening 6.BP-Ap42 depilation/not softening sheepskin test result shows:
The Pi Junneng of (1) two kind of enzyme unhairing has reached the GB standard, but the skin (1,2 of AS1398 depilation, 3) skin (4,5,6) of strength ratio BP-Ap42 depilation will hang down, illustrate that AS1398 is eager to excel to the hydrolytic action of collagen, comparatively speaking, the effect of BP-Ap42 will relax manyly;
(2) behind the enzyme unhairing of the same race with different enzyme batings, remollescent skin (3,6) intensity is not higher as can be seen, pancreatin remollescent skin (2,5) is lower, BP-Ap42 remollescent skin (1,4) is high slightly.Illustrate that pancreatin is better than BP-Ap42 to the hydrolytic action of collagen.Table 11 pigskin finished leather rerum natura
* the softening 4.BP-Ap42 depilation/BP-Ap42 of 1.AS1398 depilation/BP-Ap42 is softening
2.AS1398 the softening 5.BP-Ap42 depilation/pancreatin of depilation/pancreatin is softening
3.AS1398 depilation/not softening 6.BP-Ap42 depilation/not softening
The test-results and the sheepskin of pigskin are roughly the same, illustrate that BP-Ap42 proteolytic enzyme depilation is better than AS1398; Bating effect is better than pancreatin.
For relatively the depilation of bacillus pumilus Sumizyme MP and grey alkaline process is to the influence of leather rerum natura, we have carried out softening simultaneous test to the sheepskin that 2 kinds of methods obtain, and the results are shown in table 12.
Table 12 softens simultaneous test
* the softening 4. Sodium Sulphide depilation/pancreatin of 1.BP-Ap42 depilation/pancreatin are softening
2.BP-Ap42 depilation/BP-Ap42 softens 5. Sodium Sulphide depilation/BP-Ap42 enzyme batings
3.BP-Ap42 depilation/the softening simultaneous test finished leather rerum natura of softening two kinds of depilating methods does not detect and shows:
(1) strength of leather of Sodium Sulphide depilation generally is higher than the leather of BP-Ap42 enzyme unhairing.Illustrate that enzyme is better than alkali to the hydrolytic action of collagen, at the different leather kinds such as the production of soft leather, when formulating technology, can consider to use enzyme to handle, to reach the purpose of making soft leather;
(2) the different enzyme batings of enzyme unhairing of the same race (1,2,3), the skin intensity of BP-Ap42 enzyme bating are still than pancreatin remollescent height, this and table 11, and 12 conclusion is consistent.
It can be said that brightly, BP-Ap42 proteolytic enzyme also is a kind of good tenderizer really.
Conclusion
(1) the Sumizyme MP BP-Ap42 that bacillus pumilus produced can be used for the immersion in the process hides process, depilation and softening.Soaking time shortens to 15 hours from 24 hours of routine; Hair removal effect is thorough, and pore is clear, and grain is complete; Surface when being used for softening is pure whiter, softness, and grain is more careful, is a kind of good novel enzyme preparation.
(2) this Sumizyme MP and AS1398 proteolytic enzyme are in depilation, and softening going up compared, and it is more thorough to lose hair or feathers, more weak to the collagen hydrolysis, and softening skin is more soft.
Embodiment 7, kid shoes upper leather process program
1, immersion: liquor ratio 3.0, enzyme 30U/g, temperature is a normal temperature in the drum, time 12-15h, pH10;
2, the enzyme unhairing of banking up: enzyme dosage 200U/g, pH10, normal temperature changes 15min, goes out the drum 15-24h that banks up, and treats that hair has taken off after washing;
3, expand: NaOH 1%, Ca (OH)
23%, liquor ratio 3.0, normal temperature changes 60min, stops drum and spends the night;
4, multiple ash: Ca (OH)
25%, change 60min, washing 10min;
5, deliming: 2.5,32 ℃ of liquor ratios, (NH
4)
2SO
42% changes 60min;
6, softening: 1.2,38 ℃ of liquor ratios, pH10.0, BP-Ap42 enzyme 100U/g (body lotion) changes 120min;
7, pickling: liquor ratio 0.5, normal temperature, salt 6% changes 10min, and formic acid 1% (dilution in 1: 10) changes 10min, and sulfuric acid 1.2% (dilution in 1: 10,4 * 1/4) changes 60min, and pH2.5 stops drum and spends the night, and changes 30min next day, and discharge opeing remains 1/3 pickling waste liquid approximately;
8, chrome tanning: cation oil 1%, change 20min, chromium powder 6%, NaAc 1%, changes 60min, checks full impregnated.Sodium bicarbonate 1.2% divides 4 addings, changes 15min at every turn, to pH3.8, adds 50 ℃ of hot water to liquor ratio 2.5, regulates in the drum temperature to 38 ℃, changes 60min, stops drum and spends the night, and changes 30min (pH3.8) next day, goes out drum and takes horse 24h;
9, shaving behind the split, deburring then;
10, ease back: liquor ratio 3.0,40 ℃ of temperature, permeate agent 0.3% changes 120min, stops drum and spends the night;
11, retanning: liquor ratio 2.0,20 ℃ of normal temperature, AA 3.0%, change 30min, regulate pH4.5, MB-14.0%, 201 1.0%, change 60min, chromium powder 4.0%, change 90min, regulate pH4.0, washing 10min, sodium bicarbonate 0.6%, sodium formiate 1.0% changes 60min, pH6.5, whether the tetrabromo-mcresolsulfonphthalein inspection neutralizes, washing 20min;
12, dyeing and stuffing: 1.5,40 ℃ of liquor ratios, erie black ATT 1%, direct fast black 2% changes 30min, mends and adds water to liquor ratio 2.0, regulate 55 ℃ of the interior temperature of drum, 201 12%, 801 2%, L-3 3%, and ST 2%, sulfited fish oil 1%, change 60min, formic acid 0.5% * 2 changes 20min at every turn, pH3.8, washing goes out drum and takes horse;
13, primary coat: black cream 1%, BS-100 1.5%, 302 2%, wax 0.2%;
14, middle level: black cream 1%, complexing deceives 0.1%, and PUC-9702 0.8%, and PUC-9703 1.5%, and PUC-333 1.5%, wax 0.2%;
15, top layer: PUC-333 1.5%, wax 0.5%, hand feeling agent 0.01%, brightening agent 0.03%;
Annotate: immersion, depilation, the softening middle pH that regulates all uses yellow soda ash.
Embodiment 8, pigskin shoes upper leather process program
1, immersion: liquor ratio 3.0, normal temperature, pH10, BP-Ap42 enzyme 30U/g (body lotion) changes 30min, stops drum and spends the night;
2, degreasing: 2.5,32 ℃ of liquor ratios, soda ash 4%, peregal 4% changes 60min;
3, caustic dip: liquor ratio 2.5, NaOH 1%, changes 30min, stops drum and spends the night;
4, dealkalize: washing 10min, 1.5,32 ℃ of liquor ratios, ammonium sulfate 1% changes 60min;
5, depilation: BP-Ap42 enzyme 200U/g, pH10 rolls enzyme 30min, and 15h banks up;
6, softening: 1.2,38 ℃ of liquor ratios, BP-Ap42 enzyme 100U/mL changes 30min, washing 10min;
7, pickling: liquor ratio 0.5, normal temperature, salt 6% changes 10min, formic acid 1% (dilution in 1: 10), sulfuric acid 1.2% (dilution in 1: 10,4 * 1/4) changes 60min, and pH2.5 stops drum and spends the night, and changes 30min next day;
8, chrome tanning: remove pickling waste liquid (staying 1/3), cation oil 1% changes 20min, chromium powder 6%, and sodium-acetate 1% changes 30min, checks full impregnated.Sodium bicarbonate 1.2% divides 4 addings, changes 15min at every turn, and pH3.8 adds 50 ℃ of hot water to liquor ratio 2.5, regulates 38 ℃ of the interior temperature of drum, changes 60min, stops drum and spends the night, and changes 30min (pH3.8) next day, goes out drum and takes horse 24h;
9, shaving behind the split, deburring then;
10, ease back: 3.0,40 ℃ of liquor ratios, permeate agent 0.3% changes 120min, stops drum and spends the night, and changes liquid, 2.5,40 ℃ of liquor ratios, permeate agent 0.3%, formic acid 0.5% changes 30min;
11, retanning: liquor ratio 2.0, normal temperature, AA3.0% changes 30min, and MB 4.0%, and L-3 1.0%, changes 60min, and chromium powder 4.0% changes 90min, pH4.0, washing 10min;
12, neutralization: liquor ratio 2.0, sodium bicarbonate 1.0%, sodium-acetate 0.6% changes 60min, pH6.5 (tetrabromo-mcresolsulfonphthalein whether check full impregnated) washing 20min;
13, dyeing and stuffing: 2.0,55 ℃ of liquor ratios, erie black ATT 1%, direct fast black 2%, bright red 0.01%, change 30min, 201 6%, 801 3%, L-36%, ST 2%, the acid fish oil 1% of sulfurous, modified wool fat 2%, change 60min, formic acid 0.5% * 2/20min, pH3.8, washing goes out drum.
Annotate: immersion, depilation, the softening middle pH that regulates all uses yellow soda ash.
The present invention has following features:
The present invention finds depilation reactions vary surely to want multiple protein enzyme participation, by add entry, at mechanism and albumen of the present invention Just can finish whole depilation reaction under the effect of enzyme fully; And protease of the present invention not only can lose hair or feathers, and can make Collagenous fibres disperse and reach the purpose that makes bating; Also can be used for the technology of soaking. Protease of the present invention is to pigskin, continuous Skin and goat skin and ox-hide all have good hair removal effect, and be especially good to the hair removal effect of sheepskin and ox-hide. Adopt egg of the present invention When the depilation experimental size of white enzyme was amplified to 50/batches, effect was still very good. The leather that utilizes protease of the present invention to carry out Softening test shows: this protease all has good bating effect to pigskin, sheepskin and ox-hide. Softening finished leather rerum natura all reaches To international standard. Therefore protease of the present invention has not only solved the pollution problem of present chemical method depilation, has also overcome routine The unstable properties of enzyme unhairing, the easy damaged collagen is the shortcoming of bad operation again, and protease of the present invention also can be used for system Immersion technology and softening process in the leather industry.
Proteinase activity height of the present invention, the action condition gentleness, the activity keeping time is long, is subjected to such environmental effects few, can be routinely The method separation and purification. Produce the microbial strains of protease of the present invention, its Media Components is common, and cultural method is easy.
Claims (18)
1, a kind of Sumizyme MP is characterized in that satisfying at least one condition of following (a)-(d) defined:
(a), under the room temperature, under the EDTA of 1mM effect, the residue vigor reaches 14.7%;
(b), under the room temperature, under the EGTA of 1mM effect, the residue vigor reaches 12.7%;
(c), under the room temperature, under the dipotassium sodium arseniate effect of 1mM, the residue vigor reaches 90.3%;
(d), under the room temperature, under the methyl-phenoxide effect of 1mM, the residue vigor reaches 93.4%.
2,, it is characterized in that said Sumizyme MP has following properties according to the Sumizyme MP of claim 1:
(1), effect
This proteolytic enzyme energy protein hydrolysate and peptide;
(2), optimal pH
Under 50 ℃ when pH9.6 enzyme live the highest;
(3), stable pH range
It is stable to live at pH6-pH11 scope endoenzyme under 50 ℃;
(4), optimum temperuture
The optimal reactive temperature of this enzyme is 50 ℃;
(5), molecular weight
The molecular weight of this enzyme that records by the SDS-polyacrylamide gel electrophoresis is 40,000 dalton.
3,, it is characterized in that said Sumizyme MP is by the microorganisms that belongs to bacillus (Bacillus) according to the Sumizyme MP of claim 1 or claim 2.
4, according to the Sumizyme MP of claim 3, the microorganism that it is characterized in that said generation Sumizyme MP is bacillus pumilus UN-31-C-42 (preserving number CGMCC NO.0518).
5, a kind of method of producing Sumizyme MP is characterized in that producing the said proteolytic enzyme of acquisition bacterium bacillus pumilus UN-31-C-42 (preserving number CGMCC NO.0518) or its mutation culture from producing the said Sumizyme MP of claim 1-4.
6,, it is characterized in that said microorganism is bacillus pumilus UN-31-C-42 (preserving number CGMCC NO.0518) according to the method for claim 5.
7, a kind of microorganism that produces Sumizyme MP is characterized in that it is bacillus pumilus UN-31-C-42 (preserving number CGMCC NO.0518) and its mutant.
8, the immersion leather softener and depilatory is characterized in that the Sumizyme MP comprising claim 1-4 defined.
9, a kind of immersion leather softener and depilatory is characterized in that wherein comprising the Sumizyme MP of claim 1-4 defined.
10, a kind of preparation that can be used for the immersion technology of tanning industry is characterized in that wherein comprising the Sumizyme MP of claim 1-4 defined.
11, a kind of preparation that can be used for the dehairing process of tanning industry is characterized in that wherein comprising the Sumizyme MP of claim 1-4 defined.
12, a kind of preparation that can be used for the softening process of tanning industry is characterized in that wherein comprising the Sumizyme MP of claim 1-4 defined.
13, a kind of method of soaking of hide or skin is characterized in that soaking rawhide with the proteolytic enzyme and the water of any one defined among the claim 1-4 under mechanical effect.
14, a kind of method of dehairing of hides is characterized in that making proteolytic enzyme and the rawhide hair follicle protein or the reactive polypeptide of any one defined among the claim 1-4.
15, a kind of method that can be used for the softening process in the process hides is characterized in that using the proteolytic enzyme and the skin reaction of any one defined of claim 1-4, causes skin remollescent method.
16, according to the method for claim 13, it is characterized in that said method concrete steps are: use wet salted hide, soak and ease back; Enzyme dosage 30U/g or 50U/g during soaking of hide or skin, temperature is 18 ℃ in the drum, and liquor ratio 3 adds yellow soda ash and regulates pH10, and time 15h eases back.
17, according to the method for claim 14, it is characterized in that said method concrete steps are: wet salted hide eases back by the condition immersion of the immersion of claim 16; Be used to the enzyme unhairing of banking up behind the drip-dry water; The condition that is adopted in the epilation process is: 18 ℃ of temperature, and with the sodium carbonate solution dilution enzyme liquid of pH10, enzyme dosage is 100U/g, 16h banks up.
18., it is characterized in that said method concrete steps are according to the method for claim 15: through the skin behind immersion, depilation, liming, multiple ash, the deliming in drum, 38 ℃ of temperature, pH10, softening 2h, liquor ratio 2, enzyme dosage is 100U/g or 150U/g.
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CNB001206419A CN1303210C (en) | 2000-12-29 | 2000-12-29 | Detersile proteinase and its production process and usage and microbe producing the said proteinase |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CNB001206419A CN1303210C (en) | 2000-12-29 | 2000-12-29 | Detersile proteinase and its production process and usage and microbe producing the said proteinase |
Publications (2)
Publication Number | Publication Date |
---|---|
CN1361278A true CN1361278A (en) | 2002-07-31 |
CN1303210C CN1303210C (en) | 2007-03-07 |
Family
ID=4588317
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CNB001206419A Expired - Fee Related CN1303210C (en) | 2000-12-29 | 2000-12-29 | Detersile proteinase and its production process and usage and microbe producing the said proteinase |
Country Status (1)
Country | Link |
---|---|
CN (1) | CN1303210C (en) |
Cited By (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN100374558C (en) * | 2003-06-24 | 2008-03-12 | 四川大学 | Unhairing protease and polynucleotide encoding same |
CN103627825A (en) * | 2013-11-20 | 2014-03-12 | 邱占伟 | Method for carrying out softening treatment on leather |
CN105199983A (en) * | 2015-09-23 | 2015-12-30 | 刘彦 | Bacillus amyloliquefaciens and application of protease produced by same in tanning depilation |
CN113061062A (en) * | 2021-03-22 | 2021-07-02 | 四川大学 | Method for preparing short peptide-growth-promoting bacterium compound fertilizer by dechromization fermentation of wet blue dander and application |
CN115323081A (en) * | 2022-09-23 | 2022-11-11 | 四川大学 | Softening method for preventing damaged surface and loose surface of leather |
Family Cites Families (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
DE4411223A1 (en) * | 1994-03-31 | 1995-10-05 | Solvay Enzymes Gmbh & Co Kg | Use of alkaline proteases in commercial textile washing processes |
WO1996019590A1 (en) * | 1994-12-21 | 1996-06-27 | Novo Nordisk A/S | Method for dehairing of hides or skins by means of enzymes |
JP4030603B2 (en) * | 1995-11-02 | 2008-01-09 | ノボザイムス アクティーゼルスカブ | Alkaline protease, method for producing the same, use and microorganism producing the protease |
-
2000
- 2000-12-29 CN CNB001206419A patent/CN1303210C/en not_active Expired - Fee Related
Cited By (7)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN100374558C (en) * | 2003-06-24 | 2008-03-12 | 四川大学 | Unhairing protease and polynucleotide encoding same |
CN103627825A (en) * | 2013-11-20 | 2014-03-12 | 邱占伟 | Method for carrying out softening treatment on leather |
CN105199983A (en) * | 2015-09-23 | 2015-12-30 | 刘彦 | Bacillus amyloliquefaciens and application of protease produced by same in tanning depilation |
CN105199983B (en) * | 2015-09-23 | 2018-10-09 | 四川大学 | The application of one bacillus amyloliquefaciens and its protease of generation in terms of process hides depilation |
CN113061062A (en) * | 2021-03-22 | 2021-07-02 | 四川大学 | Method for preparing short peptide-growth-promoting bacterium compound fertilizer by dechromization fermentation of wet blue dander and application |
CN115323081A (en) * | 2022-09-23 | 2022-11-11 | 四川大学 | Softening method for preventing damaged surface and loose surface of leather |
CN115323081B (en) * | 2022-09-23 | 2023-08-18 | 四川大学 | Softening method for preventing leather from damaging and loosening |
Also Published As
Publication number | Publication date |
---|---|
CN1303210C (en) | 2007-03-07 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
Kalaikumari et al. | Bioutilization of poultry feather for keratinase production and its application in leather industry | |
Lo et al. | Production of keratinolytic enzyme by an indigenous feather–degrading strain Bacillus cereus Wu2 | |
Senthilvelan et al. | Application of enzymes for dehairing of skins: cleaner leather processing | |
Hameed et al. | Production of alkaline protease by a new Bacillus subtilis isolate for use as a bating enzyme in leather treatment | |
CN105821023B (en) | A kind of isolation and purification method of keratinase and application | |
Arunachalam et al. | Protease enzyme: an eco-friendly alternative for leather industry | |
CN103069014B (en) | The enzyme unhairing of skin and animal skin | |
CN1708590A (en) | A novel ecofriendly bio-process for leather processing | |
Łaba et al. | Enzymatic degradation of pretreated pig bristles with crude keratinase of Bacillus cereus PCM 2849 | |
Shaikh et al. | Streptomyces sp. MM-3 from rhizosphere of Psidium guajava: a potential candidate for protease with dehairing properties | |
CN1171135A (en) | Method for dehairing of hides by enzymes | |
CN1303210C (en) | Detersile proteinase and its production process and usage and microbe producing the said proteinase | |
CN1678759A (en) | Process for lime and sulfide free unhairing of skins or hides using animal and/or plant enzymes | |
CN112094759B (en) | Method for preparing white rot fungus liquid culture medium by utilizing tanning wastes and application | |
RU2052506C1 (en) | Method of treatment of hide and depilated hide | |
CN110656209B (en) | Method for making leather from sturgeon skin | |
Isaac et al. | Dehairing capability of alkaline keratinase produced by new isolated Cochliobolus hawaiiensis AUMC 8606 grown on chicken feather | |
CN109609484B (en) | Preparation method of composite depilatory enzyme preparation and application of composite depilatory enzyme preparation in depilation of imported oxhide containing feather | |
JP2004043660A (en) | Enzymic depilatory and enzymic depilation method | |
US6777219B2 (en) | Process for the preparation of alkaline protease | |
CN105199983A (en) | Bacillus amyloliquefaciens and application of protease produced by same in tanning depilation | |
CN1245523C (en) | Method for preparing leather using protease and method for treating wastes dervied from leather processing using same | |
Nagal et al. | Production of alkaline protease from Elizabethkingia meningoseptica KB042 using chicken feathers | |
WO2001042513A1 (en) | Enzymatic unhairing agent for use in tanning for producing leather and method for enzymatic unhairing treatment | |
FITRIYANTO et al. | Enzymatic activity of alkaline protease from bacillus cereus TD5B and its application as sheep skin dehairing agent |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
C06 | Publication | ||
PB01 | Publication | ||
C10 | Entry into substantive examination | ||
C57 | Notification of unclear or unknown address | ||
DD01 | Delivery of document by public notice |
Addressee: Sichuan University Document name: Notification before expiration of term |
|
SE01 | Entry into force of request for substantive examination | ||
C57 | Notification of unclear or unknown address | ||
DD01 | Delivery of document by public notice |
Addressee: Sichuan University Document name: Notification of the application for patent for invention to go through the substantive examination procedure |
|
C14 | Grant of patent or utility model | ||
GR01 | Patent grant | ||
CF01 | Termination of patent right due to non-payment of annual fee | ||
CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20070307 Termination date: 20161229 |