CN111206003B - 弱后酸化瑞士乳杆菌sh2-5-66及其应用 - Google Patents
弱后酸化瑞士乳杆菌sh2-5-66及其应用 Download PDFInfo
- Publication number
- CN111206003B CN111206003B CN202010104236.5A CN202010104236A CN111206003B CN 111206003 B CN111206003 B CN 111206003B CN 202010104236 A CN202010104236 A CN 202010104236A CN 111206003 B CN111206003 B CN 111206003B
- Authority
- CN
- China
- Prior art keywords
- lactobacillus helveticus
- fermented milk
- acidification
- post
- fermented
- 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.)
- Active
Links
- 240000002605 Lactobacillus helveticus Species 0.000 title claims abstract description 35
- 235000013967 Lactobacillus helveticus Nutrition 0.000 title claims abstract description 28
- 229940054346 lactobacillus helveticus Drugs 0.000 title claims abstract description 28
- 235000015140 cultured milk Nutrition 0.000 claims abstract description 84
- 241000194020 Streptococcus thermophilus Species 0.000 claims abstract description 18
- 238000013329 compounding Methods 0.000 claims abstract description 18
- 238000004321 preservation Methods 0.000 claims abstract description 6
- 241000894006 Bacteria Species 0.000 claims description 23
- 238000000034 method Methods 0.000 claims description 12
- 235000013373 food additive Nutrition 0.000 claims description 6
- 239000002778 food additive Substances 0.000 claims description 6
- 239000000203 mixture Substances 0.000 claims description 6
- 238000002360 preparation method Methods 0.000 claims description 6
- 235000021107 fermented food Nutrition 0.000 claims description 5
- 239000007858 starting material Substances 0.000 claims description 3
- 235000013351 cheese Nutrition 0.000 claims description 2
- 230000000630 rising effect Effects 0.000 claims description 2
- 235000013580 sausages Nutrition 0.000 claims description 2
- 238000003860 storage Methods 0.000 abstract description 33
- 230000000694 effects Effects 0.000 abstract description 24
- 238000000855 fermentation Methods 0.000 abstract description 21
- 230000004151 fermentation Effects 0.000 abstract description 21
- 238000012216 screening Methods 0.000 abstract description 12
- 231100000350 mutagenesis Toxicity 0.000 abstract description 11
- 238000002703 mutagenesis Methods 0.000 abstract description 10
- 108091006112 ATPases Proteins 0.000 abstract description 9
- 102000057290 Adenosine Triphosphatases Human genes 0.000 abstract description 9
- 239000002253 acid Substances 0.000 abstract description 7
- 230000002068 genetic effect Effects 0.000 abstract description 6
- 238000004519 manufacturing process Methods 0.000 abstract description 2
- 239000006041 probiotic Substances 0.000 abstract description 2
- 235000018291 probiotics Nutrition 0.000 abstract description 2
- 230000000529 probiotic effect Effects 0.000 abstract 1
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 description 22
- 239000000523 sample Substances 0.000 description 20
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 17
- 102000004190 Enzymes Human genes 0.000 description 14
- 108090000790 Enzymes Proteins 0.000 description 14
- 229940088598 enzyme Drugs 0.000 description 14
- 239000004310 lactic acid Substances 0.000 description 11
- 235000014655 lactic acid Nutrition 0.000 description 11
- IKHGUXGNUITLKF-UHFFFAOYSA-N Acetaldehyde Chemical compound CC=O IKHGUXGNUITLKF-UHFFFAOYSA-N 0.000 description 10
- 231100000225 lethality Toxicity 0.000 description 10
- 239000000243 solution Substances 0.000 description 10
- 102000005936 beta-Galactosidase Human genes 0.000 description 9
- 108010005774 beta-Galactosidase Proteins 0.000 description 9
- 239000001963 growth medium Substances 0.000 description 9
- 238000005259 measurement Methods 0.000 description 9
- 230000035772 mutation Effects 0.000 description 9
- 238000009395 breeding Methods 0.000 description 8
- 230000001488 breeding effect Effects 0.000 description 8
- 238000002474 experimental method Methods 0.000 description 7
- 239000000796 flavoring agent Substances 0.000 description 7
- 235000019634 flavors Nutrition 0.000 description 7
- 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 6
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 6
- 150000001875 compounds Chemical class 0.000 description 6
- 239000011521 glass Substances 0.000 description 6
- 239000008101 lactose Substances 0.000 description 6
- 235000020183 skimmed milk Nutrition 0.000 description 6
- 239000000126 substance Substances 0.000 description 6
- QSJXEFYPDANLFS-UHFFFAOYSA-N Diacetyl Chemical group CC(=O)C(C)=O QSJXEFYPDANLFS-UHFFFAOYSA-N 0.000 description 5
- 238000009630 liquid culture Methods 0.000 description 5
- 239000006228 supernatant Substances 0.000 description 5
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 4
- 241000186660 Lactobacillus Species 0.000 description 4
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 description 4
- 230000001580 bacterial effect Effects 0.000 description 4
- 238000012258 culturing Methods 0.000 description 4
- 229940039696 lactobacillus Drugs 0.000 description 4
- 238000002156 mixing Methods 0.000 description 4
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 3
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 3
- 241000186673 Lactobacillus delbrueckii Species 0.000 description 3
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- 239000006285 cell suspension Substances 0.000 description 3
- 230000008859 change Effects 0.000 description 3
- 239000012153 distilled water Substances 0.000 description 3
- 235000021001 fermented dairy product Nutrition 0.000 description 3
- 235000010855 food raising agent Nutrition 0.000 description 3
- 238000011081 inoculation Methods 0.000 description 3
- 239000012528 membrane Substances 0.000 description 3
- 235000013336 milk Nutrition 0.000 description 3
- 239000008267 milk Substances 0.000 description 3
- 210000004080 milk Anatomy 0.000 description 3
- 231100000219 mutagenic Toxicity 0.000 description 3
- 230000003505 mutagenic effect Effects 0.000 description 3
- 230000008569 process Effects 0.000 description 3
- 239000000047 product Substances 0.000 description 3
- IQUPABOKLQSFBK-UHFFFAOYSA-N 2-nitrophenol Chemical compound OC1=CC=CC=C1[N+]([O-])=O IQUPABOKLQSFBK-UHFFFAOYSA-N 0.000 description 2
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- 238000012408 PCR amplification Methods 0.000 description 2
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 description 2
- 241001052560 Thallis Species 0.000 description 2
- 238000003556 assay Methods 0.000 description 2
- 235000015278 beef Nutrition 0.000 description 2
- 229940041514 candida albicans extract Drugs 0.000 description 2
- HVYWMOMLDIMFJA-DPAQBDIFSA-N cholesterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)CCCC(C)C)[C@@]1(C)CC2 HVYWMOMLDIMFJA-DPAQBDIFSA-N 0.000 description 2
- 239000002131 composite material Substances 0.000 description 2
- 230000001276 controlling effect Effects 0.000 description 2
- 238000001816 cooling Methods 0.000 description 2
- 230000003247 decreasing effect Effects 0.000 description 2
- QELUYTUMUWHWMC-UHFFFAOYSA-N edaravone Chemical compound O=C1CC(C)=NN1C1=CC=CC=C1 QELUYTUMUWHWMC-UHFFFAOYSA-N 0.000 description 2
- 239000000284 extract Substances 0.000 description 2
- 238000001914 filtration Methods 0.000 description 2
- 235000013305 food Nutrition 0.000 description 2
- 239000007788 liquid Substances 0.000 description 2
- 229910052943 magnesium sulfate Inorganic materials 0.000 description 2
- 230000035800 maturation Effects 0.000 description 2
- 239000002609 medium Substances 0.000 description 2
- 230000020477 pH reduction Effects 0.000 description 2
- 239000000843 powder Substances 0.000 description 2
- 239000002244 precipitate Substances 0.000 description 2
- 238000005057 refrigeration Methods 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 238000005070 sampling Methods 0.000 description 2
- 238000012163 sequencing technique Methods 0.000 description 2
- 239000000725 suspension Substances 0.000 description 2
- 238000010200 validation analysis Methods 0.000 description 2
- 238000005406 washing Methods 0.000 description 2
- 238000005303 weighing Methods 0.000 description 2
- 239000012138 yeast extract Substances 0.000 description 2
- KUWPCJHYPSUOFW-YBXAARCKSA-N 2-nitrophenyl beta-D-galactoside Chemical compound O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@H]1OC1=CC=CC=C1[N+]([O-])=O KUWPCJHYPSUOFW-YBXAARCKSA-N 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
- USFZMSVCRYTOJT-UHFFFAOYSA-N Ammonium acetate Chemical compound N.CC(O)=O USFZMSVCRYTOJT-UHFFFAOYSA-N 0.000 description 1
- 239000005695 Ammonium acetate Substances 0.000 description 1
- 229920001661 Chitosan Polymers 0.000 description 1
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 description 1
- 238000007400 DNA extraction Methods 0.000 description 1
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 1
- 241000196324 Embryophyta Species 0.000 description 1
- 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 1
- 201000010538 Lactose Intolerance Diseases 0.000 description 1
- 102000016943 Muramidase Human genes 0.000 description 1
- 108010014251 Muramidase Proteins 0.000 description 1
- 108010062010 N-Acetylmuramoyl-L-alanine Amidase Proteins 0.000 description 1
- 239000001888 Peptone Substances 0.000 description 1
- 108010080698 Peptones Proteins 0.000 description 1
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 1
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 1
- 229930006000 Sucrose Natural products 0.000 description 1
- 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 1
- 239000007984 Tris EDTA buffer Substances 0.000 description 1
- 241001464778 Zymobacter Species 0.000 description 1
- 238000002835 absorbance Methods 0.000 description 1
- ZOIORXHNWRGPMV-UHFFFAOYSA-N acetic acid;zinc Chemical compound [Zn].CC(O)=O.CC(O)=O ZOIORXHNWRGPMV-UHFFFAOYSA-N 0.000 description 1
- 230000004913 activation Effects 0.000 description 1
- 229910000147 aluminium phosphate Inorganic materials 0.000 description 1
- 235000019257 ammonium acetate Nutrition 0.000 description 1
- 229940043376 ammonium acetate Drugs 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 230000001174 ascending effect Effects 0.000 description 1
- 238000003287 bathing Methods 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- 239000007853 buffer solution Substances 0.000 description 1
- 239000002962 chemical mutagen Substances 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 229960001701 chloroform Drugs 0.000 description 1
- 235000012000 cholesterol Nutrition 0.000 description 1
- 238000004587 chromatography analysis Methods 0.000 description 1
- 238000005520 cutting process Methods 0.000 description 1
- 235000013365 dairy product Nutrition 0.000 description 1
- 230000029087 digestion Effects 0.000 description 1
- 238000007865 diluting Methods 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 description 1
- BNIILDVGGAEEIG-UHFFFAOYSA-L disodium hydrogen phosphate Chemical compound [Na+].[Na+].OP([O-])([O-])=O BNIILDVGGAEEIG-UHFFFAOYSA-L 0.000 description 1
- 239000012154 double-distilled water Substances 0.000 description 1
- 238000004043 dyeing Methods 0.000 description 1
- 238000001952 enzyme assay Methods 0.000 description 1
- 239000000706 filtrate Substances 0.000 description 1
- 238000009472 formulation Methods 0.000 description 1
- 238000010353 genetic engineering Methods 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 238000003988 headspace gas chromatography Methods 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 238000000265 homogenisation Methods 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 239000007924 injection Substances 0.000 description 1
- 238000002347 injection Methods 0.000 description 1
- 230000000968 intestinal effect Effects 0.000 description 1
- 230000001678 irradiating effect Effects 0.000 description 1
- 235000010335 lysozyme Nutrition 0.000 description 1
- 229960000274 lysozyme Drugs 0.000 description 1
- 239000004325 lysozyme Substances 0.000 description 1
- 229940099596 manganese sulfate Drugs 0.000 description 1
- 239000011702 manganese sulphate Substances 0.000 description 1
- 235000007079 manganese sulphate Nutrition 0.000 description 1
- SQQMAOCOWKFBNP-UHFFFAOYSA-L manganese(II) sulfate Chemical compound [Mn+2].[O-]S([O-])(=O)=O SQQMAOCOWKFBNP-UHFFFAOYSA-L 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- 230000037323 metabolic rate Effects 0.000 description 1
- 238000009629 microbiological culture Methods 0.000 description 1
- 238000001000 micrograph Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 150000002772 monosaccharides Chemical class 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 239000002773 nucleotide Substances 0.000 description 1
- 125000003729 nucleotide group Chemical group 0.000 description 1
- 235000016709 nutrition Nutrition 0.000 description 1
- 230000035764 nutrition Effects 0.000 description 1
- 230000010355 oscillation Effects 0.000 description 1
- 238000001139 pH measurement Methods 0.000 description 1
- 239000008188 pellet Substances 0.000 description 1
- 235000019319 peptone Nutrition 0.000 description 1
- 239000002504 physiological saline solution Substances 0.000 description 1
- 229920000136 polysorbate Polymers 0.000 description 1
- 229910052700 potassium Inorganic materials 0.000 description 1
- 239000011591 potassium Substances 0.000 description 1
- 239000000276 potassium ferrocyanide Substances 0.000 description 1
- 239000003755 preservative agent Substances 0.000 description 1
- 238000003825 pressing Methods 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- PYWVYCXTNDRMGF-UHFFFAOYSA-N rhodamine B Chemical compound [Cl-].C=12C=CC(=[N+](CC)CC)C=C2OC2=CC(N(CC)CC)=CC=C2C=1C1=CC=CC=C1C(O)=O PYWVYCXTNDRMGF-UHFFFAOYSA-N 0.000 description 1
- 229940043267 rhodamine b Drugs 0.000 description 1
- 239000012488 sample solution Substances 0.000 description 1
- 238000010008 shearing Methods 0.000 description 1
- 235000010378 sodium ascorbate Nutrition 0.000 description 1
- PPASLZSBLFJQEF-RKJRWTFHSA-M sodium ascorbate Substances [Na+].OC[C@@H](O)[C@H]1OC(=O)C(O)=C1[O-] PPASLZSBLFJQEF-RKJRWTFHSA-M 0.000 description 1
- 229960005055 sodium ascorbate Drugs 0.000 description 1
- CDBYLPFSWZWCQE-UHFFFAOYSA-L sodium carbonate Substances [Na+].[Na+].[O-]C([O-])=O CDBYLPFSWZWCQE-UHFFFAOYSA-L 0.000 description 1
- 229910000029 sodium carbonate Inorganic materials 0.000 description 1
- PPASLZSBLFJQEF-RXSVEWSESA-M sodium-L-ascorbate Chemical compound [Na+].OC[C@H](O)[C@H]1OC(=O)C(O)=C1[O-] PPASLZSBLFJQEF-RXSVEWSESA-M 0.000 description 1
- 239000012086 standard solution Substances 0.000 description 1
- 230000001954 sterilising effect Effects 0.000 description 1
- 238000003756 stirring Methods 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 238000010998 test method Methods 0.000 description 1
- XOGGUFAVLNCTRS-UHFFFAOYSA-N tetrapotassium;iron(2+);hexacyanide Chemical compound [K+].[K+].[K+].[K+].[Fe+2].N#[C-].N#[C-].N#[C-].N#[C-].N#[C-].N#[C-] XOGGUFAVLNCTRS-UHFFFAOYSA-N 0.000 description 1
- 238000004448 titration Methods 0.000 description 1
- YWYZEGXAUVWDED-UHFFFAOYSA-N triammonium citrate Chemical compound [NH4+].[NH4+].[NH4+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O YWYZEGXAUVWDED-UHFFFAOYSA-N 0.000 description 1
- 239000001393 triammonium citrate Substances 0.000 description 1
- 235000011046 triammonium citrate Nutrition 0.000 description 1
- 239000012137 tryptone Substances 0.000 description 1
- 238000012795 verification Methods 0.000 description 1
- 239000004246 zinc acetate Substances 0.000 description 1
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
- C12N1/205—Bacterial isolates
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
- A23C19/00—Cheese; Cheese preparations; Making thereof
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
- A23C9/00—Milk preparations; Milk powder or milk powder preparations
- A23C9/12—Fermented milk preparations; Treatment using microorganisms or enzymes
- A23C9/123—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L13/00—Meat products; Meat meal; Preparation or treatment thereof
- A23L13/40—Meat products; Meat meal; Preparation or treatment thereof containing additives
- A23L13/45—Addition of, or treatment with, microorganisms
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L13/00—Meat products; Meat meal; Preparation or treatment thereof
- A23L13/60—Comminuted or emulsified meat products, e.g. sausages; Reformed meat from comminuted meat product
- A23L13/65—Sausages
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2400/00—Lactic or propionic acid bacteria
- A23V2400/11—Lactobacillus
- A23V2400/147—Helveticus
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/225—Lactobacillus
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Health & Medical Sciences (AREA)
- Polymers & Plastics (AREA)
- Biotechnology (AREA)
- Zoology (AREA)
- Food Science & Technology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Organic Chemistry (AREA)
- Genetics & Genomics (AREA)
- Microbiology (AREA)
- Wood Science & Technology (AREA)
- Medicinal Chemistry (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Biomedical Technology (AREA)
- Virology (AREA)
- Tropical Medicine & Parasitology (AREA)
- Nutrition Science (AREA)
- Dairy Products (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
本发明公开了一株弱后酸化瑞士乳杆菌sh2‑5‑66及其应用。本发明以益生特性优良的人源瑞士乳杆菌SH2‑1为出发菌株,通过紫外诱变,并根据H+‑ATPase酶与瑞士乳杆菌SH2‑1贮藏期产酸的关系,以H+‑ATPase酶活为筛选指标,获得弱后酸化瑞士乳杆菌sh2‑5‑66,保藏编号为CGMCC No.19123。利用本发明的瑞士乳杆菌sh2‑5‑66进行发酵乳生产,与瑞士乳杆菌SH2‑1相比,酸度降低57°T,单独利用瑞士乳杆菌sh2‑5‑66或者与商业用嗜热链球菌st447复配后生产的发酵乳各项发酵指标优良,同时瑞士乳杆菌sh2‑5‑66具有优良的遗传稳定性。
Description
技术领域
本发明属于乳酸菌技术领域,涉及一株弱后酸化瑞士乳杆菌sh2-5-66及其应用。
背景技术
发酵乳是以奶或奶粉为原料,经过杀菌后加入乳酸菌发酵而成的一种具有特殊风味、营养丰富的乳制品。因其具有促消化、减缓乳糖不耐症、调节肠道菌群平衡、降低胆固醇等作用而受到消费者的青睐。发酵乳发酵结束后需在2~4℃的温度下冷藏12~24h,这称之为发酵乳的后熟期或后酸化。消费者能接受的发酵乳的最低pH一般在4.2。但在运输、销售前的低温环境中乳酸菌依然具有一定活性,会继续生长繁殖,使发酵乳的pH值降至3.5甚至更低,导致发酵乳的外观、口感、质地变差,这不仅降低了发酵乳的品质,还严重影响了其货架期。如何通过简单有效的方法控制发酵乳的后酸化过度、延长货架期已经成为目前发酵乳制品研究的热点。
目前国内外研究者提出了很多种方法来控制发酵乳后酸化,大体上可以分为以下三类:1)物理、化学方法,如发酵后使发酵乳快速冷却、添加壳聚糖等防腐剂、改变发酵剂的比例和组成等方法,主要是通过抑制菌体活性,降低乳糖代谢速率来控制发酵后乳酸的代谢;2)基因工程,近年来国外一些公司运用基因手段筛选出在高温的条件下产酸快、在低温条件下基本不产酸或产酸甚微的菌株,例如丹麦汉森公司已研制出在冷藏条件下发酵乳糖产酸能力很弱的发酵剂,丹尼斯克公司也已经筛选出后酸化低的菌株;3)诱变育种,利用物理或化学诱变剂进行诱变育种,获得弱后酸化菌株。目前国内对乳酸菌的改性还停留在人工诱变水平上,操作复杂,并且所得性状均不能稳定遗传。
发明内容
本发明的目的在于提供一株适用于弱后酸化发酵乳制品制备或者直接食用的瑞士乳杆菌L.helveticus sh2-5-66及其应用。
发明人以L.helveticus SH2-1为出发菌株,通过紫外诱变育种获得一株弱后酸化乳酸菌菌株,该菌株与突变前菌株相比,发酵乳贮藏期20天的酸度降低57°T,经分子生物学鉴定为瑞士乳杆菌(Lactobacillus helveticus),命名为Lactobacillus helveticussh2-5-66。该菌株已于2019年12月11日在中国微生物菌种保藏管理委员会普通微生物中心(CGMCC)保藏,保藏地址为北京市朝阳区北辰西路1号院3号,保藏编号为CGMCC No.19123。
本发明还提供上述瑞士乳杆菌L.helveticus sh2-5-66在制备弱后酸化发酵食品或食品添加剂中的应用。
本发明所述的发酵食品或食品添加剂为本领域中瑞士乳杆菌作为益生菌常规使用的食品或食品添加剂,例如发酵乳、奶酪、香肠及发酵剂等。
进一步地,上述瑞士乳杆菌L.helveticus sh2-5-66在制备弱后酸化发酵食品或食品添加剂中的应用,具体方法为:采用L.helveticus sh2-5-66单菌,或者L.helveticussh2-5-66与商业用嗜热链球菌S.thermophilus st447复配后制备。
具体地,本发明提供一种弱后酸化发酵剂,由L.helveticus sh2-5-66单菌组成,或由L.helveticus sh2-5-66与商业用S.thermophilus st447复配组成。
作为本发明的优选技术方案,由L.helveticus sh2-5-66与商业用S.thermophilus st447复配组成的弱后酸化发酵剂中,L.helveticus sh2-5-66与商业用S.thermophilus st447的复配比例为1:2。
具体地,本发明提供瑞士乳杆菌L.helveticus sh2-5-66在制备弱后酸化发酵乳中的应用,具体方法为:将L.helveticus sh2-5-66单菌,或者L.helveticus sh2-5-66与商业用S.thermophilus st447复配后制备发酵乳。
作为本发明的优选技术方案,L.helveticus sh2-5-66与S.thermophilus st447复配制备发酵乳时,L.helveticus sh2-5-66与S.thermophilus st447的复配比为1:2。
与现有技术相比,本发明具有以下优点:
利用本发明的瑞士乳杆菌L.helveticus sh2-5-66生产发酵乳,比突变前菌株酸度降低57°T,发酵乳贮藏20天后酸度低于110°T。与商用发酵剂Streptococcusthermophilus st447复配后生产发酵乳的各项发酵指标优良。并且本发明的瑞士乳杆菌L.helveticus sh2-5-66具有良好的遗传稳定性,能够稳定传代100代以上。
附图表说明
图1为不同菌株贮藏期发酵乳活菌数(a)、乳酸度和pH(b)、β-半乳糖酶酶活(c)和H+-ATPase酶酶活(d)。
图2为L.helveticus sh2-5-66不同传代菌株的发酵pH。
图3为SH2-1、sh2-5-66与st447不同复配比例得到的发酵乳的持水力(a)、触变性(b)、表观粘度(c)和粘度(d)。
图4为贮藏期发酵乳活菌数(a)及酸度、pH(b)。
图5为发酵乳贮藏期表观粘度(a-e分别为st447、SH2-1、sh2-5-66、st447+SH2-1及st447+sh2-5-66)、持水力(f)、触变性(g)和粘度(h)。
图6为发酵乳贮藏期风味物质乙醛(a)和双乙酰(b)含量变化。
表1为诱变时间对L.helveticus SH2-1致死率的影响。
表2为诱变菌株H+-ATPase酶活(U)。
表3为不同诱变菌株发酵乳贮藏期酸度及pH。
表4为发酵乳贮藏期质构特性。
具体实施方式
下面结合实施例、附图和表对本发明作进一步详述。
本发明所述的L.helveticus SH2-1为发明人早期筛选得到的菌株,已在【许丽君.混合型瑞士乳杆菌发酵剂弱后酸化机制初步研究[D].2017.】中公开。S.thermophilusst447,L.delbrueckii frs4-1购自上海昊岳食品科技有限公司。Streptococcusthermophilus grx02已在中国专利200810023012.0公开,来自扬州大学江苏省乳品生物技术与安全控制重点实验室。
下述实施例中的相关培养基、试剂配方以及测试方法如下。
一、菌株活化及培养基
菌株经MRS或M17液体培养基活化两次后,按3%接种量接入到脱脂乳中培养至凝乳。
M17培养基:植物蛋白胨5.0g、聚蛋白胨5.0g、牛肉浸膏2.5g、酵母提取物5.0g、抗坏血酸钠0.5g、MgSO4·7H2O 0.25g、乳糖10g、甘油10g、磷酸氢二钠5g、蒸馏水1L,高压灭菌(101Kpa,121℃)15min;
MRS培养基:葡萄糖20.0g,胰蛋白胨10.0g,牛肉浸膏10.0g,酵母提取物5.0g,磷酸氢二钾2.0g,柠檬酸三铵2.0g,乙酸钠5.0g,吐温80 1mL,MgSO4·7H2O 0.2g,硫酸锰0.05g,蒸馏水1L,高压灭菌(101Kpa,121℃)15min;
脱脂乳培养基(RSM):脱脂乳粉120g,蒸馏水1L,高压灭菌(101Kpa,121℃)15min。
二、发酵乳制备
12%复原脱脂乳→添加6%蔗糖→均质→热处理(95℃,5min)→冷却(42℃)→接种乳酸菌(3%)→37/42℃发酵至凝乳→4℃冷藏后熟12h。
三、贮藏期发酵乳指标测定
将4℃冷藏后熟12h时定为贮藏第1d,分别测定贮藏第1、5、10、15、20d的发酵乳相关指标。
1.贮藏期发酵乳中活菌数、酸度、pH、质构、粘度、持水力、流变特性和微观结构的测定
(1)活菌数测定:参照GB 4789.35-2016。
(2)滴定酸度:参照GB 5413.34-2016。
(3)pH测定:采用pH计直接测定。
(4)质构分析测定:TMS-PRO质构仪(美国,TMS公司),采用12.7mm的圆柱型探头,测定速度60mm/min,测定前速度60mm/min,返回速度60mm/min,穿刺距离为15mm,力量元0.05N。记录硬度、弹性、内聚性、胶黏性等数据。
(5)粘度测定:经4℃后熟的发酵乳用玻璃棒顺时针搅拌5圈,再逆时针搅拌5圈,用RVD-Ⅶ+黏度计测定。
(6)持水力测定:取发酵乳样品10g装入离心管,室温下离心,3500r/min离心15min,倒掉上清液称重,并按以下公式计算:持水力(%)=(离心沉淀物重量/样品重量)×100%。
(7)流变特性测定:利用旋转流变仪,选用CP2/50SR0162 SS锥板,对样品的触变性和表观粘度进行测定,表观粘度测定时在恒温25℃、恒定剪切速率1.0s-1条件下,剪切时间为2min,每10s采集一个数据点,检测样品的表观粘度随时间的变化情况。
在恒温25℃条件下,分升速和降速两个步骤进行测定,首先升速过程剪切速率由0.1r·s-1线性升髙到100r·s-1,每2s采集一个数据,测定时间为2min;降速过程剪切速率到达100r·s-1后再线性降速到0.1r·s-1,每2s采集一个数据,测定时间为2min,检测样品剪切应力随剪切速率的变化关系。
(8)风味物质测定:采用顶空气相色谱法测定风味物质,取4mL发酵乳样品6000r/min离心15min,取上清用0.22μm的微孔滤膜过滤,取0.2μL滤液进行色谱分析,检测样品中风味物质乙醛、双乙酰的含量。
(9)微观结构测定:将后熟后的发酵乳样品搅拌20次,取一滴滴于载玻片中心,滴加罗丹明B水溶液(0.2%,w/w,BBI)染色10min,盖上盖玻片轻压,使样品均匀分布于载玻片中央,用双蒸水冲洗掉多余染料,用滤纸在盖玻片另一侧将水吸干。待玻片干燥后,使用双光子激光共聚焦显微镜(ZEISS,德国),He/Ne激光束激发出的488nm波长下利用63倍油镜观察发酵乳的微观结构。观察多个视野,存储具有典型特征的显微照片。
以上实验均设置三组平行。
2.贮藏期乳糖、乳酸含量的测定
(1)乳糖含量的测定
准确称取贮藏1、5、10、15、20d的发酵乳样品2.00g,缓慢加入1mL的106g/L亚铁***和220g/L的乙酸锌,再加水定容至50.00g,混合均匀,过滤后放置于4℃备用。
取单糖、混标溶液以及样品溶液各1mL,加入200μL 0.3mol/L氢氧化钠和150μL0.5mol/L 1-苯基-3-甲基-5-吡唑啉酮(PMP)的甲醇溶液,混合均匀后,静置70℃水浴锅中30min,取出后加入200μL 0.3mol/L的盐酸溶液,再加入3mL三氯甲烷,涡旋震荡1min后静置取上层水相,经0.45μm滤膜过滤,置于4℃保存。
测定条件:ZORBAX Eclipse XDB-C18色谱柱(4.6×150mm,5μm),流动相为纯乙腈和浓度为100mmol/L,pH 5.5的乙酸铵溶液,体积比为22:78;流速为1mL/min;柱温为30℃;进样量为10μL;测定波长选择244nm。
(2)乳酸含量的测定
在1.00g发酵乳样品中加入3mL 1mol/L的盐酸溶液,6000r/min离心15min,上清液过0.22μm滤膜后置于4℃备用。
测定条件:Agilent XDB-C18色谱柱(4.6×150mm,5μm);以甲醇和浓度10mmol/L的磷酸缓冲溶液(pH 2.5)为流动相,体积比为3:97,流速1mL/min;柱温为35℃,20μL的进样量,测定波长选择210nm。
3.贮藏期相关酶活力的测定
(1)发酵乳粗酶液的制备
分别取10.0g贮藏第1、5、10、15、20d发酵乳,用1%EDTA溶液(pH=12,4℃)去蛋白后用TE缓冲液重悬,加入溶菌酶,37℃水浴1h,离心去掉上清液,用生理盐水重悬,高压破碎细胞仪破碎细胞悬浮液提取粗酶液用于测定β-半乳糖苷酶和H+-ATPase酶的活性。
(2)β-半乳糖苷酶活性测定
(a)绘制ONP(邻硝基酚)标准曲线
(b)β-半乳糖苷酶活性测定
取粗酶液1mL,37℃的水浴锅中放置10min后加入1mL ONPG溶液(pH 7.0,20mmol/L),37℃反应10min后立即加入3mL的0.5mol/L Na2CO3溶液终止反应,420nm下测定吸光度。。
β-半乳糖苷酶活力(U)=A/(B×C×D)
式中:A-ONP浓度,μmol/L;B-反应时间,min;C-取样体积,mL;D-样品蛋白浓度,mg/mL。
(3)H+-ATPase酶活性测定
按照试剂盒测定,采用氢-钾ATP酶试剂盒,购自南京建成生物工程研究所。
四、诱变育种
1.细胞悬浮液的制备
将菌株用MRS液体培养基培养至对数期(12h),10000g离心10min,收集沉淀得菌体,用0.85%灭菌生理盐水洗涤菌体两次,将菌悬液稀释制备108cfu/mL细胞悬浮液,4℃放置备用。
2.紫外诱变处理及致死率的测定
紫外灯(20W)开启预热30min,取5mL菌悬液置于直径9cm的培养皿中,放置于磁力搅拌器上,紫外灯照射距离为30cm,开启磁力搅拌器分别照射0s、30s、60s、90s、120s之后立即盖上培养皿盖。4℃、10000g离心10min,收集沉淀得菌体。将菌体悬浮在5mL的液体MRS液体培养基中,42℃恒温培养箱预培养12h,稀释不同倍数倒平板,42℃培养72h,进行活菌计数。以未经紫外处理的各稀释度菌液为对照,计算不同紫外诱变时间的致死率,整个诱变过程在避光条件下进行。
致死率公式:S=1-M/M0×100%
式中:S为致死率;M0为对照平板上的菌落数;M为经紫外线照射后平板上的菌落数。
五、弱后酸化菌株筛选
选取致死率为80%~90%的诱变菌株培养皿,经中间培养后长出单菌落,挑选菌落42℃恒温培养48h,以H+-ATPase酶活力为指标进行弱后酸化菌株的筛选。
将筛选菌株接种于脱脂乳培养基,测定12h、贮藏1d、3d、5d的pH和酸度进行进一步的验证。
六、遗传稳定性
以6%的接菌量每隔8h传代一次,共计传代100次,传代过程中取样后在相同条件下培养后测定pH值、活菌数。
实施例1
弱后酸化菌株的诱变育种、筛选及鉴定
1.诱变育种
采用20W紫外灯,照射距离30cm,分别照射0s、30s、60s、90s、120s,测定诱变时间下的致死率(表1)。照射50s后菌株致死率在80%以上,因此诱变照射的时间定为50s~60s。
表1诱变时间对L.helveticus SH2-1致死率的影响
2.筛选
(1)后酸化筛选指标的确立
为确定与Lactobacillus helveticus SH2-1后酸化偶联的酶用于后期诱变菌株的筛选指标,本部分实验对L.helveticus SH2-1发酵后贮藏期指标进行测定,同时选取两株后酸化较弱的菌株L.delbrueckii frs4-1和Streptococcus thermophilus grx02作为对照。
图1为不同菌株贮藏期发酵乳活菌数(a)、乳酸度和pH(b)、β-半乳糖酶酶活(c)和H+-ATPase酶酶活(d)。贮藏10天前,三株菌均保持生长(图1a),与此同时发酵乳pH降低、酸度升高(图1b),10天后活菌数开始下降,但pH仍保持下降、酸度上升,与L.delbrueckii frs4-1和S.thermophilus grx02相比,L.helveticus SH2-1发酵乳在贮藏15天后pH下降、酸度升高的幅度更大。根据文献报道,β-半乳糖苷酶和H+-ATPase酶是乳酸菌代谢乳糖产生乳酸的关键酶,但存在菌株差异性,因此,本实验同时测定了贮藏期间,β-半乳糖苷酶(图1c)和H+-ATPase酶(图1d)酶活的变化,由结果可知,L.helveticus SH2-1中的β-半乳糖苷酶在整个贮藏期先上升后下降,至后期除S.thermophilus grx02外,其他两株菌酶活接近零。H+-ATPase酶在整个贮藏期间均保持较高酶活,与发酵乳的pH和酸度变化趋势一致。基于以上结果,将H+-ATPase酶用作弱后酸化菌株诱变育种的筛选指标。
(2)筛菌
选取紫外诱导突变致死率为80%~90%的突变菌株培养皿,培养后选取80个菌落接种至MRS液体培养基,42℃恒温培养48h,对55株正常生长的样品测定H+-ATPase酶活,未突变的L.helveticus SH2-1作为对照(表2)。测得的H+-ATPase酶活最低为0.03U,最高为5.25U。
(3)后酸化验证
选取酶活低于对照L.helveticus SH2-1(2.32U)的20株菌进行后酸化验证,除样品2,3,11,20,21,28,44,45,49,51为凝乳外,其他十株后酸化结果如表3。结合贮藏期发酵乳的酸度和pH,选取的菌株的发酵乳后酸化均弱于L.helveticus SH2-1,说明该方法可以用于L.helveticus SH2-1诱变育种弱后酸化菌株的筛选。最终,通过本实验筛选获得的样品52的贮藏期酸度最低,较L.helveticus SH2-1贮藏期酸度下降约57°T。
(4)菌株鉴定及遗传稳定性
采用Ezup柱式细菌基因组DNA抽提试剂盒提取菌株52的DNA,进行PCR扩增,PCR扩增后将产物切胶、纯化、回收后,送至生工生物工程(上海)有限公司进行测序,核苷酸序列见序列表SEQ ID NO.1所示。测序结果经EZBioCloud中16S-based ID在线序列比对:菌株52的16SrDNA与瑞士乳杆菌L.helveticus strain KLDS 1.0603同源性为99%,相差7个碱基,具有较高的同源性,但尚有部分序列存在一定差异,属新菌株,命名为L.helveticus sh2-5-66。
将L.helveticus sh2-5-66经MRS液体培养基进行传代100代培养,传代期间取样培养后测定pH,结果如图2所示,pH基本保持一致,该菌株发酵产酸方面具有良好的遗传稳定性。
实施例2
(1)L.helveticus sh2-5-66的发酵效果实验
将SH2-1(L.helveticus SH2-1)与sh2-5-66(L.helveticus sh2-5-66)进行单菌发酵,或者与商用发酵剂st447(S.thermophilus st447)不同比例混合后进行混菌发酵,测定发酵乳的品质指标,具体实验方法如下:
单菌发酵:将活化好的菌株按照3%的量接种于配置好的脱脂乳培养基,放置于42℃恒温培养箱中培养,观察凝乳情况,记录凝乳时间,放置于4℃冰箱中贮存。
混菌发酵:将st447分别与诱变前后的瑞士乳杆菌SH2-1与sh2-5-66按照1:1、1.5:1、2:1的比例进行复配,且加入总活菌数为6×106cfu/mL进行接种发酵,放置于42℃恒温培养箱中培养,观察凝乳情况,记录凝乳时间。
(2)SH2-1、sh2-5-66与st447复配比例对发酵乳品质的影响
将st447分别与SH2-1与sh2-5-66按照1:1、1.5:1、2:1的比例进行复配发酵,测定发酵乳的持水力、触变性、表观粘度及粘度。图3为SH2-1、sh2-5-66与st447不同复配比例得到的发酵乳的持水力(a)、触变性(b)、表观粘度(c)和粘度(d)结果图。从图中可以看出,随着复配比例的增加,这两组复配发酵乳的持水力、表观粘度和粘度均增加,st447与SH2-1与sh2-5-66分别按1:1.5复配时的触变性高于其他两个复配比例,但综合四个指标,选择st447与SH2-1与sh2-5-66分别按1:2复配进行下一步实验。
(3)SH2-1、sh2-5-66与st447复配发酵乳的发酵特性
将SH2-1、sh2-5-66、st447分别进行单菌发酵乳的制备,以及SH2-1与st447(st447+SH2-1)、sh2-5-66与st447(st447+sh2-5-66)按2:1复配后制备发酵乳,测定发酵乳及其贮藏期的各项指标。
(3.1)发酵乳特性
测定发酵乳贮藏期间的活菌数及酸度和pH,结果如图4所示,发酵菌株在贮藏期5天内均继续生长,5天后活菌数开始呈下降趋势,st447+sh2-5-66比st447+SH2-1活菌数低,但比单菌发酵时活菌数高,st447+sh2-5-66发酵乳贮藏第一天活菌数高达8.98Logcfu/mL,贮藏20天后仍有7.86Logcfu/mL。
贮藏期间发酵乳的pH和酸度分别呈现下降和上升趋势,sh2-5-66单菌发酵及st447+sh2-5-66复合发酵的酸度均明显低于其他几组,贮藏20天时酸度低于120°T,比SH2-1及st447+SH2-1低20到30°T。pH趋势与酸度保持一致。
(3.2)发酵乳质构特性
测定贮藏期发酵乳质构特性,结果如表4和图5所示。贮藏期间发酵乳硬度、内聚性、弹性、胶粘及咀嚼性呈现先上升后下降的趋势,其中,硬度和内聚性几组之间差别不大,弹性、胶粘及咀嚼性方面,复合菌发酵高于单菌发酵,同时st447+sh2-5-66略高于st447+SH2-1。
五组发酵乳的表观粘度在贮藏期内均呈现先上升后下降的趋势,SH2-1发酵乳的表观粘度明显低于其他四组,sh2-5-66与st447复配后能够提高发酵乳的表观粘度,st447+sh2-5-66复配发酵乳在贮藏期20天时表观粘度仍高于其他几组,同时,随着剪切力的升高,表观粘度变化平稳,没有出现较大的波动。st447+sh2-5-66复配发酵乳在贮藏1天时持水力在70%以上,贮藏20天时持水力仍保持55%以上,结合触变性和粘度数据,st447+sh2-5-66复配发酵乳在贮藏期间复弹性好,凝胶强度高,能够较好的保持发酵乳的理化状态。
(3.3)发酵乳风味特性
测定发酵乳中风味物质乙醛和双乙酰含量,结果如图6所示,st447+sh2-5-66发酵乳中乙醛含量低于st447+SH2-1,但高于其他单菌发酵,双乙酰低于st447发酵乳,但高于其他三组发酵乳。在贮藏20天时,st447+sh2-5-66发酵乳中乙醛和双乙酰含量仍分别含有15.6mg/L和1.25mg/mL。
序列表
<110> 扬州大学
<120> 弱后酸化瑞士乳杆菌sh2-5-66及其应用
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1477
<212> DNA
<213> 瑞士乳杆菌(Lactobacillus helveticus)
<400> 1
cggccgggcg gcgtgctata catgcaagtc gagcgagcag aaccagcaga tttacttcgg 60
taatgacgct ggggacgcga gcggcggatg ggtgagtaac acgtggggaa cctgccccat 120
agtctgggat accacttgga aacaggtgct aataccggat aagaaagcag atcgcatgat 180
cagcttataa aaggcggcgt aagctgtcgc tatgggatgg ccccgcggtg cattagctag 240
ttggtaaggt aacggcttac caaggcaatg atgcatagcc gagttgagag actgatcggc 300
cacattggga ctgagacacg gcccaaactc ctacgggagg cagcagtagg gaatcttcca 360
caatggacgc aagtctgatg gagcaacgcc gcgtgaggtg aagaaggttt tcggatcgta 420
aagctctgtt gttggtgaag aaggatagag gtagtaactg gcctttattt gacggtaatc 480
aaccagaaag tcacggctaa ctacgtgcca gcagccgcgg taatacgtag gtggcaagcg 540
ttgtccggat ttattgggcg taaagcgagc gcaggcggaa gaataagtct gatgtgaaag 600
ccctcggctt aaccgaggaa ctgcatcgga aactgttttt cttgagtgca gaagaggaga 660
gtggaattcc atgtgtagcg gtggaatgcg tagatatatg gaagaacacc agtggcgaag 720
gcgactctct ggtctgcaac tgacgctgag gctcgaaagc atgggtagcg aacaggatta 780
gataccctgg tagtccatgc cgtaaacgat gagtgctaag tgttgggagg tttccgcctc 840
tcagtgctgc agctaacgca ttaagcactc cgcctgggga gtacgaccgc aaaggttgaa 900
actcaaagga attgacgggg ggcccgcaca agcggtggga gcatgtggtt taattcgaag 960
caacgcgaag aaccttacca ggtcttgaca tctagtgcca tcctaagaga ttaggagttc 1020
ccttcgggga cgctaagaca ggtggtgcat ggcctgtcgt cagctcgtgt cgtgagaatg 1080
ttgggttaag tcccgcaaac gagcgcaacc cttgttatta gttgccagca ttaagttggg 1140
cactctaatg agactgccgg tgataaaccg gaggaaggtg gggatgacgt caagtcatca 1200
tgccccttat gacctgggct acacacgtgc tacaatggac agtacaacga gaagcgagcc 1260
tgcgaaggca agcgaatctc tgaaagctgt tctcagttcg gactgcagtc tgcaactcga 1320
ctgcacgaag ctggaatcgc tagtaatcgc ggatcagaac gccgcggtga atacgttccc 1380
gggccttgta cacaccgccc gtcacaccat ggaagtctgc aatgcccaaa gccggtggcc 1440
taaccttcgg gaaggagccg tctaagcagg tagattg 1477
Claims (8)
1.弱后酸化瑞士乳杆菌(Lactobacillus helveticus)sh2-5-66,保藏编号为CGMCCNo. 19123,保藏时间为2019年12月11日,保藏地址为北京市朝阳区北辰西路1号院3号。
2.根据权利要求1所述的弱后酸化瑞士乳杆菌sh2-5-66在制备弱后酸化发酵食品或食品添加剂中的应用。
3.根据权利要求2所述的应用,其特征在于,所述的发酵食品或食品添加剂为发酵乳、奶酪、香肠或发酵剂。
4.根据权利要求2或3所述的应用,其特征在于,具体方法为:采用sh2-5-66单菌,或者sh2-5-66与嗜热链球菌(Streptococcus thermophilus)st447复配后制备。
5.弱后酸化发酵剂,其特征在于,由权利要求1所述的弱后酸化瑞士乳杆菌sh2-5-66单菌组成,或由权利要求1所述的弱后酸化瑞士乳杆菌sh2-5-66与嗜热链球菌st447复配组成。
6.根据权利要求5所述的弱后酸化发酵剂,其特征在于,由sh2-5-66与st447复配组成的弱后酸化发酵剂中, sh2-5-66与st447的复配比例为1:2。
7.根据权利要求4所述的应用,其特征在于,sh2-5-66在制备弱后酸化发酵乳中的应用,具体方法为:将sh2-5-66单菌,或者sh2-5-66与st447复配后制备发酵乳。
8.根据权利要求7所述的应用,其特征在于, sh2-5-66与st447复配制备发酵乳时,sh2-5-66与st447的复配比为1:2。
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202010104236.5A CN111206003B (zh) | 2020-02-20 | 2020-02-20 | 弱后酸化瑞士乳杆菌sh2-5-66及其应用 |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202010104236.5A CN111206003B (zh) | 2020-02-20 | 2020-02-20 | 弱后酸化瑞士乳杆菌sh2-5-66及其应用 |
Publications (2)
Publication Number | Publication Date |
---|---|
CN111206003A CN111206003A (zh) | 2020-05-29 |
CN111206003B true CN111206003B (zh) | 2021-09-10 |
Family
ID=70781195
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN202010104236.5A Active CN111206003B (zh) | 2020-02-20 | 2020-02-20 | 弱后酸化瑞士乳杆菌sh2-5-66及其应用 |
Country Status (1)
Country | Link |
---|---|
CN (1) | CN111206003B (zh) |
Families Citing this family (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN114395579B (zh) * | 2021-10-29 | 2024-04-19 | 扬州大学 | 一种基因i型与基因iii型日本脑炎病毒感染性克隆及其构建方法与应用 |
Citations (9)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
EP0575010A1 (en) * | 1992-06-19 | 1993-12-22 | Verenigde Coöperatieve Melkindustrie Coberco B.A. | Method of preparing a fermented milk product |
EP0652285A2 (en) * | 1993-11-04 | 1995-05-10 | The Calpis Food Industry Co., Ltd. | Lactic acid bacteria and fermented milk product |
CN104059866A (zh) * | 2014-06-17 | 2014-09-24 | 克拉玛依绿成农业开发有限责任公司 | 用于发酵豆乳的乳酸菌组合发酵剂 |
CN104480187A (zh) * | 2014-12-05 | 2015-04-01 | 临沂格瑞食品有限公司 | 一株高产γ-氨基丁酸乳酸菌及其筛选方法 |
CN105920049A (zh) * | 2016-06-02 | 2016-09-07 | 扬州大学 | 鼠李糖乳杆菌grx19在调节肠道菌群中的应用 |
CN106234891A (zh) * | 2016-02-05 | 2016-12-21 | 上海交大昂立股份有限公司 | 一种常温保存的高活性益生菌发酵植物饮料及其制备方法 |
CN106754472A (zh) * | 2016-11-23 | 2017-05-31 | 扬州大学 | 一株果蔬发酵用植物乳杆菌grx15及其应用 |
CN107189959A (zh) * | 2017-05-19 | 2017-09-22 | 扬州大学 | 瑞士乳杆菌grx09及其应用 |
CN108684816A (zh) * | 2017-04-12 | 2018-10-23 | 内蒙古乳业技术研究院有限责任公司 | 一种可控制后酸的发酵乳制品及其制备方法 |
Family Cites Families (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
ES2384096T3 (es) * | 2006-02-20 | 2012-06-29 | Compagnie Gervais Danone | Cepas de Lactobacillus helveticus que no fermentan la lactosa |
CN103215199B (zh) * | 2013-03-13 | 2014-08-20 | 内蒙古农业大学 | 一株具有延缓后酸化作用保加利亚乳杆菌菌株及其应用 |
CN109504636B (zh) * | 2018-12-27 | 2022-04-12 | 内蒙古农业大学 | 一种植物乳杆菌p12及其用途 |
CN109517763B (zh) * | 2018-12-27 | 2022-03-22 | 内蒙古农业大学 | 瑞士乳杆菌h11及其应用 |
-
2020
- 2020-02-20 CN CN202010104236.5A patent/CN111206003B/zh active Active
Patent Citations (9)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
EP0575010A1 (en) * | 1992-06-19 | 1993-12-22 | Verenigde Coöperatieve Melkindustrie Coberco B.A. | Method of preparing a fermented milk product |
EP0652285A2 (en) * | 1993-11-04 | 1995-05-10 | The Calpis Food Industry Co., Ltd. | Lactic acid bacteria and fermented milk product |
CN104059866A (zh) * | 2014-06-17 | 2014-09-24 | 克拉玛依绿成农业开发有限责任公司 | 用于发酵豆乳的乳酸菌组合发酵剂 |
CN104480187A (zh) * | 2014-12-05 | 2015-04-01 | 临沂格瑞食品有限公司 | 一株高产γ-氨基丁酸乳酸菌及其筛选方法 |
CN106234891A (zh) * | 2016-02-05 | 2016-12-21 | 上海交大昂立股份有限公司 | 一种常温保存的高活性益生菌发酵植物饮料及其制备方法 |
CN105920049A (zh) * | 2016-06-02 | 2016-09-07 | 扬州大学 | 鼠李糖乳杆菌grx19在调节肠道菌群中的应用 |
CN106754472A (zh) * | 2016-11-23 | 2017-05-31 | 扬州大学 | 一株果蔬发酵用植物乳杆菌grx15及其应用 |
CN108684816A (zh) * | 2017-04-12 | 2018-10-23 | 内蒙古乳业技术研究院有限责任公司 | 一种可控制后酸的发酵乳制品及其制备方法 |
CN107189959A (zh) * | 2017-05-19 | 2017-09-22 | 扬州大学 | 瑞士乳杆菌grx09及其应用 |
Non-Patent Citations (3)
Title |
---|
A weak post-acidification lactobacillus helveticus mutant with improved textural properties;Chengran Guan等;《Food Science & Nutrition》;20201115(第9期);第469-479页 * |
嗜热链球菌对瑞士乳杆菌发酵乳后酸化的影响;张臣臣等;《乳液科学与技术》;20180901;第41卷(第5期);第1-5页 * |
混合型瑞士乳杆菌发酵剂弱后酸化机制初步研究;许丽君;《中国优秀硕士学位论文全文数据库》;20180115;第16页倒数第1段,表2-1、表2-2 * |
Also Published As
Publication number | Publication date |
---|---|
CN111206003A (zh) | 2020-05-29 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
JP4772131B2 (ja) | 新規乳酸菌を用いた発酵乳の製造方法 | |
CN108728382B (zh) | 一株具降胆固醇及促进肠道短链脂肪酸产生能力的植物乳杆菌及其应用 | |
CN101472481B (zh) | 低后酸化乳酸菌 | |
CN110066750B (zh) | 嗜热链球菌jmcc0024及分离纯化方法、应用 | |
JP4862053B2 (ja) | 新規乳酸菌 | |
CN107586741B (zh) | 一种植物乳杆菌及其在水果酵素产品中的应用 | |
CN109536406B (zh) | 弱后酸化的嗜热链球菌jmcc16、分离纯化方法及应用 | |
CN107674852B (zh) | 一种产丁二酮的植物乳杆菌及其应用 | |
CN110157650B (zh) | 一株分离自母乳的乳双歧杆菌m8及其应用 | |
CN111206003B (zh) | 弱后酸化瑞士乳杆菌sh2-5-66及其应用 | |
CN116240128A (zh) | 植物乳杆菌、发酵剂及其在制备鲜湿米粉中的应用 | |
Han et al. | Milk fermentation by monocultures or co-cultures of Streptococcus thermophilus strains | |
CN117099848A (zh) | 一种清酒乳杆菌常温凝固型胡萝卜酸奶及其制备方法 | |
CN110484477B (zh) | 一株德氏乳杆菌保加利亚亚种菌株及其应用 | |
CN115895979A (zh) | 一种低温益生菌酸奶发酵剂及酸奶制备方法 | |
Özdemir et al. | Effect of Gluconacetobacter spp. on Kefir grains and Kefir Quality | |
CN115820512A (zh) | 一株嗜热链球菌菌株EuAS-D-S6及其应用 | |
CN1793325A (zh) | 芳香型直投式酸奶发酵剂 | |
CN109749962A (zh) | 一株耐受性强、产酸高的山西老陈醋优势土著风味植物乳杆菌及应用 | |
CN108330082A (zh) | 一株副干酪乳杆菌及其应用 | |
CN114532400A (zh) | 嗜热链球菌及应用其制备的酸奶 | |
CN109022307B (zh) | 一种具有高脲酶活力的嗜热链球菌及其应用 | |
JP3953777B2 (ja) | 食品醗酵のための炭素含有添加剤及びそれを含有する食品組成物 | |
JP4794593B2 (ja) | 新規乳酸菌を用いた発酵乳の製造方法 | |
CN108060099B (zh) | 一种辅助发酵剂、其制备方法及其在发酵制品中的应用 |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
PB01 | Publication | ||
PB01 | Publication | ||
SE01 | Entry into force of request for substantive examination | ||
SE01 | Entry into force of request for substantive examination | ||
GR01 | Patent grant | ||
GR01 | Patent grant |