CN109770363A - A kind of nanometer selenium Freeze-dry Powder of Probioctics and preparation method thereof - Google Patents
A kind of nanometer selenium Freeze-dry Powder of Probioctics and preparation method thereof Download PDFInfo
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- BUGBHKTXTAQXES-UHFFFAOYSA-N Selenium Chemical compound [Se] BUGBHKTXTAQXES-UHFFFAOYSA-N 0.000 title claims abstract description 109
- 239000011669 selenium Substances 0.000 title claims abstract description 106
- 229910052711 selenium Inorganic materials 0.000 title claims abstract description 105
- 239000000843 powder Substances 0.000 title claims abstract description 75
- 238000002360 preparation method Methods 0.000 title claims abstract description 37
- 229940091258 selenium supplement Drugs 0.000 claims abstract description 104
- 230000004913 activation Effects 0.000 claims abstract description 70
- 238000000855 fermentation Methods 0.000 claims abstract description 60
- 230000004151 fermentation Effects 0.000 claims abstract description 60
- 239000006041 probiotic Substances 0.000 claims abstract description 52
- 235000018291 probiotics Nutrition 0.000 claims abstract description 52
- 241000194020 Streptococcus thermophilus Species 0.000 claims abstract description 40
- 241000186000 Bifidobacterium Species 0.000 claims abstract description 37
- 240000001046 Lactobacillus acidophilus Species 0.000 claims abstract description 37
- 235000013956 Lactobacillus acidophilus Nutrition 0.000 claims abstract description 37
- 229940039695 lactobacillus acidophilus Drugs 0.000 claims abstract description 37
- 239000012530 fluid Substances 0.000 claims abstract description 36
- 235000015097 nutrients Nutrition 0.000 claims abstract description 36
- 239000007788 liquid Substances 0.000 claims abstract description 29
- BVTBRVFYZUCAKH-UHFFFAOYSA-L disodium selenite Chemical compound [Na+].[Na+].[O-][Se]([O-])=O BVTBRVFYZUCAKH-UHFFFAOYSA-L 0.000 claims abstract description 24
- 229960001471 sodium selenite Drugs 0.000 claims abstract description 24
- 235000015921 sodium selenite Nutrition 0.000 claims abstract description 24
- 239000011781 sodium selenite Substances 0.000 claims abstract description 24
- 230000001376 precipitating effect Effects 0.000 claims abstract description 10
- 238000005119 centrifugation Methods 0.000 claims abstract description 8
- 238000005406 washing Methods 0.000 claims abstract description 6
- 241000894006 Bacteria Species 0.000 claims description 34
- 239000002054 inoculum Substances 0.000 claims description 15
- 238000006243 chemical reaction Methods 0.000 claims description 13
- 238000013329 compounding Methods 0.000 claims description 6
- 238000007710 freezing Methods 0.000 claims description 6
- 230000008014 freezing Effects 0.000 claims description 6
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims description 5
- 239000011780 sodium chloride Substances 0.000 claims description 5
- HEBKCHPVOIAQTA-NGQZWQHPSA-N d-xylitol Chemical compound OC[C@H](O)C(O)[C@H](O)CO HEBKCHPVOIAQTA-NGQZWQHPSA-N 0.000 claims description 4
- MCAHWIHFGHIESP-UHFFFAOYSA-N selenous acid Chemical compound O[Se](O)=O MCAHWIHFGHIESP-UHFFFAOYSA-N 0.000 claims description 4
- 238000002156 mixing Methods 0.000 claims description 3
- 238000011081 inoculation Methods 0.000 claims description 2
- 239000002244 precipitate Substances 0.000 claims description 2
- 239000002609 medium Substances 0.000 claims 5
- 239000001963 growth medium Substances 0.000 claims 3
- 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 claims 2
- 229910052708 sodium Inorganic materials 0.000 claims 2
- 239000011734 sodium Substances 0.000 claims 2
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims 1
- 240000007594 Oryza sativa Species 0.000 claims 1
- 235000007164 Oryza sativa Nutrition 0.000 claims 1
- 241000194017 Streptococcus Species 0.000 claims 1
- 229910052799 carbon Inorganic materials 0.000 claims 1
- 229910052739 hydrogen Inorganic materials 0.000 claims 1
- 239000001257 hydrogen Substances 0.000 claims 1
- 125000004435 hydrogen atom Chemical class [H]* 0.000 claims 1
- 235000015110 jellies Nutrition 0.000 claims 1
- 239000008274 jelly Substances 0.000 claims 1
- 235000009566 rice Nutrition 0.000 claims 1
- 230000000694 effects Effects 0.000 abstract description 5
- 235000016709 nutrition Nutrition 0.000 abstract description 4
- 230000035764 nutrition Effects 0.000 abstract description 4
- 230000000529 probiotic effect Effects 0.000 abstract description 4
- 230000009286 beneficial effect Effects 0.000 abstract description 3
- 235000013325 dietary fiber Nutrition 0.000 abstract description 3
- 229910052500 inorganic mineral Inorganic materials 0.000 abstract description 3
- 230000031891 intestinal absorption Effects 0.000 abstract description 3
- 230000000968 intestinal effect Effects 0.000 abstract description 3
- 230000000813 microbial effect Effects 0.000 abstract description 3
- 235000010755 mineral Nutrition 0.000 abstract description 3
- 239000011707 mineral Substances 0.000 abstract description 3
- 235000013343 vitamin Nutrition 0.000 abstract description 3
- 239000011782 vitamin Substances 0.000 abstract description 3
- 229940088594 vitamin Drugs 0.000 abstract description 3
- 229930003231 vitamin Natural products 0.000 abstract description 3
- 150000003722 vitamin derivatives Chemical class 0.000 abstract description 3
- 229940065287 selenium compound Drugs 0.000 abstract description 2
- 150000003343 selenium compounds Chemical class 0.000 abstract description 2
- 235000011649 selenium Nutrition 0.000 description 85
- UIIMBOGNXHQVGW-UHFFFAOYSA-M Sodium bicarbonate Chemical compound [Na+].OC([O-])=O UIIMBOGNXHQVGW-UHFFFAOYSA-M 0.000 description 8
- 238000004108 freeze drying Methods 0.000 description 7
- 235000013406 prebiotics Nutrition 0.000 description 6
- 230000008859 change Effects 0.000 description 5
- 241000193830 Bacillus <bacterium> Species 0.000 description 4
- 241001147746 Lactobacillus delbrueckii subsp. lactis Species 0.000 description 4
- 230000015572 biosynthetic process Effects 0.000 description 4
- 238000000034 method Methods 0.000 description 4
- 235000017557 sodium bicarbonate Nutrition 0.000 description 4
- 229910000030 sodium bicarbonate Inorganic materials 0.000 description 4
- 238000003786 synthesis reaction Methods 0.000 description 4
- PMYDPQQPEAYXKD-UHFFFAOYSA-N 3-hydroxy-n-naphthalen-2-ylnaphthalene-2-carboxamide Chemical compound C1=CC=CC2=CC(NC(=O)C3=CC4=CC=CC=C4C=C3O)=CC=C21 PMYDPQQPEAYXKD-UHFFFAOYSA-N 0.000 description 3
- 241000588769 Proteus <enterobacteria> Species 0.000 description 3
- 230000008901 benefit Effects 0.000 description 3
- 230000037396 body weight Effects 0.000 description 3
- 150000001875 compounds Chemical class 0.000 description 3
- 244000005700 microbiome Species 0.000 description 3
- 239000002105 nanoparticle Substances 0.000 description 3
- 239000000047 product Substances 0.000 description 3
- 238000000926 separation method Methods 0.000 description 3
- 229960001881 sodium selenate Drugs 0.000 description 3
- 235000018716 sodium selenate Nutrition 0.000 description 3
- 239000011655 sodium selenate Substances 0.000 description 3
- 231100000419 toxicity Toxicity 0.000 description 3
- 230000001988 toxicity Effects 0.000 description 3
- 238000009777 vacuum freeze-drying Methods 0.000 description 3
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- SRBFZHDQGSBBOR-IOVATXLUSA-N D-xylopyranose Chemical compound O[C@@H]1COC(O)[C@H](O)[C@H]1O SRBFZHDQGSBBOR-IOVATXLUSA-N 0.000 description 2
- 241001052560 Thallis Species 0.000 description 2
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- 238000010586 diagram Methods 0.000 description 2
- 230000009977 dual effect Effects 0.000 description 2
- 230000036541 health Effects 0.000 description 2
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 description 2
- 238000009630 liquid culture Methods 0.000 description 2
- 239000002245 particle Substances 0.000 description 2
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- 230000009467 reduction Effects 0.000 description 2
- 150000003839 salts Chemical class 0.000 description 2
- 239000013049 sediment Substances 0.000 description 2
- VXVKGIZYLHTHMR-WCCKRBBISA-N (2s)-2-amino-4-methylselanylbutanoic acid;selenium Chemical compound [Se].C[Se]CC[C@H](N)C(O)=O VXVKGIZYLHTHMR-WCCKRBBISA-N 0.000 description 1
- 208000024172 Cardiovascular disease Diseases 0.000 description 1
- 241000588724 Escherichia coli Species 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- KIECCKFSGCJGTM-UHFFFAOYSA-N O[Se](O)=O.[SeH2] Chemical compound O[Se](O)=O.[SeH2] KIECCKFSGCJGTM-UHFFFAOYSA-N 0.000 description 1
- 241000589516 Pseudomonas Species 0.000 description 1
- 241000589180 Rhizobium Species 0.000 description 1
- 206010039921 Selenium deficiency Diseases 0.000 description 1
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- 239000002253 acid Substances 0.000 description 1
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- 230000003078 antioxidant effect Effects 0.000 description 1
- 235000006708 antioxidants Nutrition 0.000 description 1
- PYMYPHUHKUWMLA-UHFFFAOYSA-N arabinose Natural products OCC(O)C(O)C(O)C=O PYMYPHUHKUWMLA-UHFFFAOYSA-N 0.000 description 1
- SRBFZHDQGSBBOR-UHFFFAOYSA-N beta-D-Pyranose-Lyxose Natural products OC1COC(O)C(O)C1O SRBFZHDQGSBBOR-UHFFFAOYSA-N 0.000 description 1
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Landscapes
- Coloring Foods And Improving Nutritive Qualities (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
Abstract
The invention discloses a kind of nanometer selenium Freeze-dry Powder of Probioctics and preparation method thereof, first respectively by lactobacillus acidophilus, streptococcus thermophilus and Bifidobacterium Anaerobic culturel, activation；Then above-mentioned three kinds of strains after activation are inoculated with Anaerobic culturel in the MRS fluid nutrient medium containing sodium selenite respectively, obtain the fermentation liquid of three kinds of strains；Then it is centrifugated after the fermentation liquid of three kinds of strains being mixed, obtains probiotics precipitating, obtain nanometer selenium Freeze-dry Powder of Probioctics after washing, centrifugation, frozen dried；The present invention is by combining lactobacillus acidophilus, the health-care efficacy of three kinds of probiotics of streptococcus thermophilus and Bifidobacterium and the best selenium-supply form of nanometer selenium, it is prepared into high-activity nano selenium compound probiotic freeze-dried powder, the probiotics for being not only rich in the microbial flora balance beneficial to human body intestinal canal, minerals, vitamin, the dietary fiber for being easy to intestinal absorption, nanometer selenium double nutrition healthcare function is more added, and the partial size of nanometer selenium powder is 60-100nm.
Description
Technical field
The invention belongs to compound probiotic powder preparation technical field, in particular to a kind of nanometer selenium Freeze-dry Powder of Probioctics and its
Preparation method.
Background technique
Selenium (Se) is a kind of important microelement for having on biosystem and influencing extensively, including anti-oxidant, anticancer, antiviral
With promote the biological effects such as growth and breeding.Selenium deficiency can lead to cardiovascular disease, cancer, Kaschin-Beck disease, disease of viral infection
Generation.The form of human body selenium-supply is mainly traditional inorganic selenium, organic selenium at present.Inorganic selenium general security is low, takes slightly
One excessively will cause to be poisoned, and bioavailability is low.Organic selenium is current main selenium-supply form, organic selenium than inorganic selenium have compared with
High bioactivity, but price is high, scarcity of resources.Research finds LD50Inorganic selenium (selenite) is 22.0mg/kg/body
Weight, organic selenium (selenomethionine) is 25.6mg/kg/body weight, and nanometer selenium is then 92.1mg/kg/body
Weight illustrates that nanometer selenium has the efficient activity advantage of low toxicity, and the selenium-supply product as a kind of high activity, high safety will become
Substitute the optimal selection of other forms selemium nutrition preparation or medicament.
Currently, the synthesis of nanometer selenium has chemical method and biotransformation method, the nanometer selenium of bioanalysis conversion has structure single
Uniformly, the spherical of rule is presented, and stability is good, can be resistant to high temperature, is not easy to be converted to brown or black, performance is better than chemistry
The nanometer selenium of synthesis.Studied in terms of the reduction for carrying out selenium using microorganism both at home and abroad more, that reports at present has using solid
The red bacterium of nitrogen, pseudomonad, bacillus, rodlike Pseudomonas, existing germ oligotrophy unit cell, Escherichia coli, rhizobium, lactic acid bacteria
Selenous acid or selenate can be reduced into red nano elemental selenium by equal microorganisms.Most microorganisms are resistant to the sub- selenium of low concentration
Acid and its salt are simultaneously reduced into nanometer selenium, but with the raising of concentration of sodium selenite, are enhanced the toxicity of thallus, final to inhibit
Thalli growth influences the conversion ratio of nanometer selenium.
The Microbe synthesis of organic selenium has been reported, wherein Chinese patent application is using a kind of compound benefit rich in selenium
It is disclosed in raw bacterium powder preparation and its preparation (Publication No. CN107772477A) and organic selenium is synthesized using single bacterium Bifidobacterium, but
There is no the nanometer seleniums that open report is minimum using probiotics synthesis toxicity；Chinese patent application one kind contains nanometer selenium, rare earth
It is disclosed in probiotic feed additive of element and preparation method thereof (Publication No. CN108094716A) a kind of for animal
The additive of feed, but be subject to certain restrictions as selenium-enriched food exploitation, and step is more, preparation process is complicated.
Summary of the invention
For the technical problems in the prior art, the present invention provides a kind of nanometer selenium Freeze-dry Powder of Probioctics and its systems
Preparation Method, the present invention are mixed by using a variety of probiotics, prepare the selenium-enriched probiotics powder containing nanometer selenium, collect nanometer selenium
In one, the preparation for selenium-rich product provides new way for high physiological activity and probiotics health care dual function.
In order to achieve the above objectives, the technical solution adopted by the present invention are as follows:
A kind of preparation method of nanometer selenium Freeze-dry Powder of Probioctics, comprising the following steps:
Lactobacillus acidophilus is seeded in MRS fluid nutrient medium by step 1, actication of culture, and Anaerobic culturel, activation are lived
Lactobacillus acidophilus after change；Streptococcus thermophilus is seeded in MRS fluid nutrient medium, Anaerobic culturel, activation, after being activated
Streptococcus thermophilus；Bifidobacterium is seeded in MRS fluid nutrient medium, Anaerobic culturel, activation, the bifid bar after being activated
Bacterium；
Lactobacillus acidophilus after activation, is seeded in the MRS Liquid Culture containing sodium selenite by step 2, nanometer selenium conversion
Anaerobic culturel in base obtains lactobacillus acidophilus fermentation liquid；Streptococcus thermophilus after activation is seeded in containing sodium selenite
Anaerobic culturel in MRS fluid nutrient medium obtains streptococcus thermophilus fermentation mash；Bifidobacterium after activation is seeded in containing Asia
Anaerobic culturel in the MRS fluid nutrient medium of sodium selenate, obtains bifidobacterium fermentation mash；
Step 3, rich nanometer selenium probiotics compounding, by lactobacillus acidophilus fermentation liquid, streptococcus thermophilus obtained in step 2
Fermentation liquid and the mixing of bifidobacterium fermentation mash, obtain mixed fermentation mash；
Mixed fermentation mash in step 3 it is heavy to be centrifugally separating to obtain probiotics by the collection of step 4, nanometer selenium probiotics
It forms sediment, after probiotics washing of precipitate, centrifugation, obtains wet bacterium powder；
Step 5, the preparation of nanometer selenium Freeze-dry Powder of Probioctics, by the wet bacterium powder frozen dried in step 4, it is prebiotic to obtain nanometer selenium
Bacterium freeze-dried powder.
Further, in step 1 when actication of culture, the temperature of Anaerobic culturel is 32-37 DEG C；Actication of culture is using continuous living
Change 2-3 times, the time of first time activation culture is 24-48h, remaining activation culture time is 12-16h.
Further, the inoculum concentration of the lactobacillus acidophilus after activating in step 2 is 1-3%, the streptococcus thermophilus after activation
The inoculum concentration of inoculation is 1-3%, and the inoculum concentration of the Bifidobacterium after activation is 1-3%.
Further, in step 2 by after activation lactobacillus acidophilus or streptococcus thermophilus be seeded in containing concentration be 30-
Anaerobic culturel in the MRS fluid nutrient medium of 60mmol/L sodium selenite；Bifidobacterium after activation is seeded in and is containing concentration
Anaerobic culturel in the MRS fluid nutrient medium of 15-30mmol/L sodium selenite.
Further, the temperature of Anaerobic culturel is 32-37 DEG C in step 2, and the Anaerobic culturel time is 48-72h.
Further, lactobacillus acidophilus fermentation liquid in mixed fermentation mash in step 3, streptococcus thermophilus fermentation mash and
The volume ratio of bifidobacterium fermentation mash is (1-2): (1-3): (1-4).
Further, mixed fermentation liquid is centrifuged using high speed freezing centrifuge in step 4, centrifugation time 10-15min,
Revolving speed is 8000-10000r/min.
Further, probiotics precipitating is washed using sterile saline suspension, is centrifuged in step 4；Centrifugation time is 10-
15min, revolving speed 8000-10000r/min.
Further, in step 5 when wet bacterium powder frozen dried, pre-freeze is used first, and then freezing is dry under vacuum conditions
Dry 24-36h；Cryoprotector is added in wet bacterium powder before frozen dried；By mass percentage, cryoprotector is using oligomeric
Xylose 10-14%, synanthrin 5-7%, sodium bicarbonate 0.8-1%.
Compared with prior art, the invention has the benefit that
A kind of nanometer of Freeze-dry Powder of Probioctics of the invention and preparation method thereof, by by lactobacillus acidophilus, streptococcus thermophilus and
The health-care efficacy and the best selenium-supply form of nanometer selenium of three kinds of probiotics of Bifidobacterium combine, and it is compound to be prepared into high-activity nano selenium
Freeze-dry Powder of Probioctics is not only rich in the probiotics of the microbial flora balance beneficial to human body intestinal canal, is easy to the mineral of intestinal absorption
Matter, vitamin, dietary fiber more add nanometer selenium double nutrition healthcare function, and the partial size of nanometer selenium powder is 60-
100nm。
Detailed description of the invention
Fig. 1 is the nanometer selenium Freeze-dry Powder of Probioctics scanning electron microscope (SEM) photograph prepared in the embodiment of the present invention 1；
Fig. 2 is the SEM test chart of the probiotics conversion nanometer selenium probiotics in the embodiment of the present invention 1；
Fig. 3 is the EDX test chart of the probiotics conversion nanometer selenium probiotics in the embodiment of the present invention 1；
Fig. 4 is the XRD diagram of the nanometer selenium probiotics in the embodiment of the present invention 1.
Specific embodiment
Invention is further described in detail with reference to the accompanying drawings and detailed description.
A kind of preparation method of nanometer selenium Freeze-dry Powder of Probioctics of the present invention, comprising the following steps:
Lactobacillus acidophilus is seeded in MRS fluid nutrient medium by step 1, actication of culture, and Anaerobic culturel, activation are lived
Lactobacillus acidophilus after change；Streptococcus thermophilus is seeded in MRS fluid nutrient medium, Anaerobic culturel, activation, after being activated
Streptococcus thermophilus；Bifidobacterium is seeded in MRS fluid nutrient medium, Anaerobic culturel, activation, the bifid bar after being activated
Bacterium；
Lactobacillus acidophilus after activation, is seeded in the MRS Liquid Culture containing sodium selenite by step 2, nanometer selenium conversion
Anaerobic culturel in base obtains lactobacillus acidophilus fermentation liquid；Streptococcus thermophilus after activation is seeded in containing sodium selenite
Anaerobic culturel in MRS fluid nutrient medium obtains streptococcus thermophilus fermentation mash；Bifidobacterium after activation is seeded in containing Asia
Anaerobic culturel in the MRS fluid nutrient medium of sodium selenate, obtains bifidobacterium fermentation mash；
Step 3, rich nanometer selenium probiotics compounding, by lactobacillus acidophilus fermentation liquid, streptococcus thermophilus obtained in step 2
Fermentation liquid and the mixing of bifidobacterium fermentation mash, obtain mixed fermentation mash；
Mixed fermentation mash in step 3 it is heavy to be centrifugally separating to obtain probiotics by the collection of step 4, nanometer selenium probiotics
It forms sediment, washing, centrifugation obtain wet bacterium powder；
Step 5, the preparation of nanometer selenium Freeze-dry Powder of Probioctics, by the wet bacterium powder frozen dried in step 4, it is prebiotic to obtain nanometer selenium
Bacterium freeze-dried powder.
Embodiment 1
A kind of preparation method of nanometer selenium Freeze-dry Powder of Probioctics, comprising the following steps:
Lactobacillus acidophilus is seeded in MRS fluid nutrient medium by step 1, actication of culture, and anaerobism is trained under the conditions of 37 DEG C
It supports, continuous activation 3 times, activation culture time first time is that for 24 hours, second of activation culture time is 16h, thermophilic after being activated
Lactobacillus lactis；Streptococcus thermophilus is seeded in MRS fluid nutrient medium, Anaerobic culturel, continuous activation 3 times under the conditions of 37 DEG C,
Activation culture time first time is that for 24 hours, second of activation culture time is 16h, the streptococcus thermophilus after being activated, by bifid
Bacillus is seeded in MRS fluid nutrient medium, Anaerobic culturel, continuous activation 3 times, activation culture time first time under the conditions of 37 DEG C
For for 24 hours, second of activation culture time is 16h, the Bifidobacterium after being activated；
Lactobacillus acidophilus after activation, is pressed 2% inoculum concentration, is seeded in and is containing concentration by step 2, nanometer selenium conversion
In the MRS fluid nutrient medium of the sodium selenite of 60mmol/L, Anaerobic culturel 48h under conditions of 37 DEG C obtains lactobacillus acidophilus
Fermentation liquid；The inoculum concentration that streptococcus thermophilus after activation is pressed to 2% is seeded in the sodium selenite for being 60mmol/L containing concentration
MRS fluid nutrient medium in, Anaerobic culturel 48h under conditions of 37 DEG C obtains streptococcus thermophilus fermentation mash；After activation
Bifidobacterium presses 2% inoculum concentration, is seeded in the MRS fluid nutrient medium containing the sodium selenite that concentration is 20mmol/L,
Anaerobic culturel 48h under conditions of 37 DEG C, obtains bifidobacterium fermentation mash；
Step 3, rich nanometer selenium probiotics compounding, by lactobacillus acidophilus fermentation liquid, streptococcus thermophilus obtained in step 2
Fermentation liquid and bifidobacterium fermentation mash are mixed according to the ratio that volume ratio is 1:1:1, obtain mixed fermentation mash；
Mixed fermentation mash in step 3 is placed in high speed freezing centrifuge by the collection of step 4, nanometer selenium probiotics,
At 4 DEG C of temperature, under conditions of revolving speed is 10000r/min, it is centrifugated 10min, discards the isolated probiotics precipitating of clear liquid；
It is suspended washing probiotics precipitating using sterile saline again, using at 4 DEG C of temperature, under conditions of revolving speed is 10000r/min,
Centrifuge separation 10min obtains wet bacterium powder；
Step 5, the preparation of nanometer selenium Freeze-dry Powder of Probioctics, by the wet bacterium powder frozen dried in step 4, it is prebiotic to obtain nanometer selenium
Bacterium freeze-dried powder；When wet bacterium powder frozen dried, first using freeze drying protectant is added in wet bacterium powder, using in -20 DEG C of temperature
After lower pre-freeze 2h, vacuum freeze drying 36h；Wherein, as mass fraction, freeze drying protectant uses xylo-oligosaccharide 14%, synanthrin
7%, sodium bicarbonate 1%.
Embodiment 2
A kind of preparation method of nanometer selenium Freeze-dry Powder of Probioctics, comprising the following steps:
Lactobacillus acidophilus is seeded in MRS fluid nutrient medium by step 1, actication of culture, and anaerobism is trained under the conditions of 35 DEG C
It supports, continuous activation 3 times, activation culture time first time is 36h, and second of activation culture time is 14h, thermophilic after being activated
Lactobacillus lactis；Streptococcus thermophilus is seeded in MRS fluid nutrient medium, Anaerobic culturel, continuous activation 3 times under the conditions of 35 DEG C,
Activation culture time first time is 36h, and second of activation culture time is 14h, the streptococcus thermophilus after being activated, by bifid
Bacillus is seeded in MRS fluid nutrient medium, Anaerobic culturel, continuous activation 3 times, activation culture time first time under the conditions of 35 DEG C
For 36h, second of activation culture time is 14h, the Bifidobacterium after being activated；
Lactobacillus acidophilus after activation, is pressed 3% inoculum concentration, is seeded in and is containing concentration by step 2, nanometer selenium conversion
In the MRS fluid nutrient medium of the sodium selenite of 45mmol/L, Anaerobic culturel 60h under conditions of 35 DEG C obtains lactobacillus acidophilus
Fermentation liquid；The inoculum concentration that streptococcus thermophilus after activation is pressed to 3% is seeded in the sodium selenite for being 45mmol/L containing concentration
MRS fluid nutrient medium in, Anaerobic culturel 60h under conditions of 35 DEG C obtains streptococcus thermophilus fermentation mash；After activation
Bifidobacterium presses 3% inoculum concentration, is seeded in the MRS fluid nutrient medium containing the sodium selenite that concentration is 30mmol/L,
Anaerobic culturel 60h under conditions of 35 DEG C, obtains bifidobacterium fermentation mash；
Step 3, rich nanometer selenium probiotics compounding, by lactobacillus acidophilus fermentation liquid, streptococcus thermophilus obtained in step 2
Fermentation liquid and bifidobacterium fermentation mash are mixed according to the ratio that volume ratio is 1:1:2, obtain mixed fermentation mash；
Mixed fermentation mash in step 3 is placed in high speed freezing centrifuge by the collection of step 4, nanometer selenium probiotics,
At 4 DEG C of temperature, under conditions of revolving speed is 8000r/min, it is centrifugated 15min, discards the isolated probiotics precipitating of clear liquid；Again
It is suspended using sterile saline and washs probiotics precipitating, using at 4 DEG C of temperature, under conditions of revolving speed is 8000r/min, be centrifuged
Separation 15min obtains wet bacterium powder；
Step 5, the preparation of nanometer selenium Freeze-dry Powder of Probioctics, by the wet bacterium powder frozen dried in step 4, it is prebiotic to obtain nanometer selenium
Bacterium freeze-dried powder；When wet bacterium powder frozen dried, first using freeze drying protectant is added in wet bacterium powder, using in -20 DEG C of temperature
After lower pre-freeze 2h, vacuum freeze drying 30h；Wherein, as mass fraction, freeze drying protectant uses xylo-oligosaccharide 12%, synanthrin
6%, sodium bicarbonate 0.9%.
Embodiment 3
A kind of preparation method of nanometer selenium Freeze-dry Powder of Probioctics, comprising the following steps:
Lactobacillus acidophilus is seeded in MRS fluid nutrient medium by step 1, actication of culture, and anaerobism is trained under the conditions of 32 DEG C
It supports, continuous activation 2 times, activation culture time first time is 48h, and second of activation culture time is 12h, thermophilic after being activated
Lactobacillus lactis；Streptococcus thermophilus is seeded in MRS fluid nutrient medium, Anaerobic culturel, continuous activation 3 times under the conditions of 32 DEG C,
Activation culture time first time is 48h, and second of activation culture time is 12h, the streptococcus thermophilus after being activated, by bifid
Bacillus is seeded in MRS fluid nutrient medium, Anaerobic culturel, continuous activation 3 times, activation culture time first time under the conditions of 32 DEG C
For 48h, second of activation culture time is 12h, the Bifidobacterium after being activated；
Lactobacillus acidophilus after activation, is pressed 1% inoculum concentration, is seeded in and is containing concentration by step 2, nanometer selenium conversion
In the MRS fluid nutrient medium of the sodium selenite of 30mmol/L, Anaerobic culturel 72h under conditions of 32 DEG C obtains lactobacillus acidophilus
Fermentation liquid；The inoculum concentration that streptococcus thermophilus after activation is pressed to 1% is seeded in the sodium selenite for being 50mmol/L containing concentration
MRS fluid nutrient medium in, Anaerobic culturel 72h under conditions of 32 DEG C obtains streptococcus thermophilus fermentation mash；After activation
Bifidobacterium presses 1% inoculum concentration, is seeded in the MRS fluid nutrient medium containing the sodium selenite that concentration is 15mmol/L,
Anaerobic culturel 72h under conditions of 32 DEG C, obtains bifidobacterium fermentation mash；
Step 3, rich nanometer selenium probiotics compounding, by lactobacillus acidophilus fermentation liquid, streptococcus thermophilus obtained in step 2
Fermentation liquid and bifidobacterium fermentation mash are mixed according to the ratio that volume ratio is 2:3:4, obtain mixed fermentation mash；
Mixed fermentation mash in step 3 is placed in high speed freezing centrifuge by the collection of step 4, nanometer selenium probiotics,
At 4 DEG C of temperature, under conditions of revolving speed is 10000r/min, it is centrifugated 10min, discards the isolated probiotics precipitating of clear liquid；
It is suspended washing probiotics precipitating using sterile saline again, using at 4 DEG C of temperature, under conditions of revolving speed is 10000r/min,
Centrifuge separation 10min obtains wet bacterium powder；
Step 5, the preparation of nanometer selenium Freeze-dry Powder of Probioctics, by the wet bacterium powder frozen dried in step 4, it is prebiotic to obtain nanometer selenium
Bacterium freeze-dried powder；When wet bacterium powder frozen dried, first using freeze drying protectant is added in wet bacterium powder, using in -20 DEG C of temperature
After lower pre-freeze 2h, vacuum freeze drying is for 24 hours；Wherein, as mass fraction, freeze drying protectant uses xylo-oligosaccharide 10%, synanthrin
5%, sodium bicarbonate 0.8%.
The nanometer selenium Freeze-dry Powder of Probioctics that the present invention is prepared, powder are in cerise, are the characteristic colors of nanometer selenium,
Uniformly, color does not change grain after room temperature 6 months, illustrates freeze-dried powder nanoparticles stable；With reference to shown in attached drawing 1, give
The nanometer selenium Freeze-dry Powder of Probioctics scanning electron microscope (SEM) photograph for having gone out the preparation of the embodiment of the present invention 1, it can be seen that probiotics is thin from attached drawing 1
There are a large amount of compacted grains around born of the same parents and in cytoplasm, granular size is about 60~100nm, and particle is uniform；
With reference to shown in attached drawing 2-3, SEM and the EDX test of probiotics conversion nanometer selenium in the embodiment of the present invention 1 are given
Figure, it can be seen that the particle on probiotics surface is mainly nanometer selenium from attached drawing 2-3；
With reference to shown in attached drawing 4, the XRD diagram of the nanometer selenium Freeze-dry Powder of Probioctics prepared in the embodiment of the present invention 1 is given, from
It can be seen that the nano particle of nanometer selenium Freeze-dry Powder of Probioctics is the amorphous volume morphing without stationary state in attached drawing 4.
A kind of nanometer selenium Freeze-dry Powder of Probioctics of the present invention, by by lactobacillus acidophilus, streptococcus thermophilus and Bifidobacterium three
The health-care efficacy and the best selenium-supply form of nanometer selenium of kind probiotics combine, and are prepared into the freeze-drying of high-activity nano selenium compound probiotic
Powder, not only rich in beneficial to human body intestinal canal microbial flora balance probiotics, be easy to intestinal absorption minerals, vitamin,
Dietary fiber more adds nanometer selenium double nutrition healthcare function, and the partial size of nanometer selenium powder is 60-100nm, and the present invention obtains
The nanometer selenium Freeze-dry Powder of Probioctics powder arrived is in cerise, is the characteristic color of nanometer selenium, and powder granule is uniform；Nanometer selenium is prebiotic
The nanoparticles stable of bacterium freeze-dried powder places six months or more colors under room temperature and does not change.
Many probiotics are resistant to the selenous acid and its salt of low concentration, and are reduced into nanometer selenium, but with selenous acid
The raising of na concn enhances the toxicity of thallus, finally inhibits thalli growth, influence the conversion ratio of nanometer selenium；The present invention is with Asia
The color of sodium selenate reduction rate and red nano selenium is index, is obtained using the methods of condition domestication, segmentation addition screening thermophilic
Lactobacillus lactis, streptococcus thermophilus reach as high as 60mmol/L to the tolerable concentration of sodium selenite, and Bifidobacterium is to sodium selenite
Tolerable concentration reaches as high as 30mmol/L, and sodium selenite high conversion rate is up to 84.5% or more；Multi-joint richness is prepared into using three kinds of bacterium
Nanometer selenium probiotics composite preparation combines the high physiological activity of nanometer selenium and probiotics health care dual function, produces for selenium-rich
The preparation of product provides new way.
Claims (10)
1. a kind of preparation method of nanometer selenium Freeze-dry Powder of Probioctics, which comprises the following steps:
Lactobacillus acidophilus is seeded in MRS fluid nutrient medium by step 1, actication of culture, Anaerobic culturel, activation, after obtaining activation
Lactobacillus acidophilus；Streptococcus thermophilus is seeded in MRS fluid nutrient medium, Anaerobic culturel, activation are thermophilic after being activated
Streptococcus；Bifidobacterium is seeded in MRS fluid nutrient medium, Anaerobic culturel, activation, the Bifidobacterium after being activated；
Step 2, nanometer selenium conversion, the lactobacillus acidophilus after activation is seeded in the MRS fluid nutrient medium containing sodium selenite
Anaerobic culturel obtains lactobacillus acidophilus fermentation liquid；Streptococcus thermophilus after activation is seeded in the MRS liquid containing sodium selenite
Anaerobic culturel in body culture medium obtains streptococcus thermophilus fermentation mash；Bifidobacterium after activation is seeded in containing selenous acid
Anaerobic culturel in the MRS fluid nutrient medium of sodium, obtains bifidobacterium fermentation mash；
Step 3, rich nanometer selenium probiotics compounding, by lactobacillus acidophilus fermentation liquid, streptococcus thermophilus fermentation obtained in step 2
Mash and the mixing of bifidobacterium fermentation mash, obtain mixed fermentation mash；
Mixed fermentation mash in step 3 is centrifugally separating to obtain probiotics precipitating by the collection of step 4, nanometer selenium probiotics,
After probiotics washing of precipitate, centrifugation, wet bacterium powder is obtained；
Wet bacterium powder frozen dried in step 4 is obtained nanometer selenium probiotics jelly by step 5, the preparation of nanometer selenium Freeze-dry Powder of Probioctics
Dry powder.
2. a kind of preparation method of nanometer selenium Freeze-dry Powder of Probioctics according to claim 1, which is characterized in that in step 1
When actication of culture, the temperature of Anaerobic culturel is 32-37 DEG C；Actication of culture is activated 2-3 times using continuous, first time activation culture
Time is 24-48h, remaining activation culture time is 12-16h.
3. a kind of preparation method of nanometer selenium Freeze-dry Powder of Probioctics according to claim 1, which is characterized in that in step 2
The inoculum concentration of lactobacillus acidophilus after activation is 1-3%, and the inoculum concentration of the streptococcus thermophilus inoculation after activation is 1-3%, activation
The inoculum concentration of Bifidobacterium afterwards is 1-3%.
4. a kind of preparation method of nanometer selenium Freeze-dry Powder of Probioctics according to claim 1, which is characterized in that in step 2
By after activation lactobacillus acidophilus or streptococcus thermophilus be seeded in containing concentration be 30-60mmol/L sodium selenite MRS liquid
Anaerobic culturel in culture medium；Bifidobacterium after activation is seeded in the MRS liquid containing concentration for 15-30mmol/L sodium selenite
Anaerobic culturel in body culture medium.
5. a kind of preparation method of nanometer selenium Freeze-dry Powder of Probioctics according to claim 1, which is characterized in that in step 2
The temperature of Anaerobic culturel is 32-37 DEG C, and the Anaerobic culturel time is 48-72h.
6. a kind of preparation method of nanometer selenium Freeze-dry Powder of Probioctics according to claim 1, which is characterized in that in step 3
The volume ratio of lactobacillus acidophilus fermentation liquid, streptococcus thermophilus fermentation mash and bifidobacterium fermentation mash in mixed fermentation mash
For (1-2): (1-3): (1-4).
7. a kind of preparation method of nanometer selenium Freeze-dry Powder of Probioctics according to claim 1, which is characterized in that in step 4
Mixed fermentation liquid is centrifuged using high speed freezing centrifuge, centrifugation time 10-15min, revolving speed 8000-10000r/min.
8. a kind of preparation method of nanometer selenium Freeze-dry Powder of Probioctics according to claim 1, which is characterized in that in step 4
Probiotics precipitating is suspended using sterile saline washs, is centrifuged；Centrifugation time is 10-15min, revolving speed 8000-10000r/
min。
9. a kind of preparation method of nanometer selenium Freeze-dry Powder of Probioctics according to claim 1, which is characterized in that in step 5
When wet bacterium powder frozen dried, pre-freeze is used first, is then freeze-dried 24-36h under vacuum conditions；In wet bacterium before frozen dried
Cryoprotector is added in powder；By mass percentage, cryoprotector uses xylo-oligosaccharide 10-14%, synanthrin 5-7%, carbon
Sour hydrogen sodium 0.8-1%.
10. what the preparation method of nanometer selenium Freeze-dry Powder of Probioctics described in -9 any one was prepared according to claim 1 receives
Rice selenium Freeze-dry Powder of Probioctics.
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| CN117866857A (en) * | 2024-03-13 | 2024-04-12 | 中国农业大学 | Selenium-enriched corynebacterium H8 capable of relieving intestinal inflammation and preparation method and application thereof |
| CN117866857B (en) * | 2024-03-13 | 2024-05-28 | 中国农业大学 | Selenium-enriched corynebacterium H8 capable of relieving intestinal inflammation and preparation method and application thereof |
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