CN116694528B - Application of bifidobacterium longum LF04 in preparation of antioxidant and anti-aging products - Google Patents
Application of bifidobacterium longum LF04 in preparation of antioxidant and anti-aging products Download PDFInfo
- Publication number
- CN116694528B CN116694528B CN202310731195.6A CN202310731195A CN116694528B CN 116694528 B CN116694528 B CN 116694528B CN 202310731195 A CN202310731195 A CN 202310731195A CN 116694528 B CN116694528 B CN 116694528B
- Authority
- CN
- China
- Prior art keywords
- bifidobacterium longum
- zebra fish
- group
- antioxidant
- preparation
- 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
- 241001608472 Bifidobacterium longum Species 0.000 title claims abstract description 65
- 229940009291 bifidobacterium longum Drugs 0.000 title claims abstract description 63
- 230000003078 antioxidant effect Effects 0.000 title claims abstract description 13
- 239000003963 antioxidant agent Substances 0.000 title claims abstract description 12
- 230000003712 anti-aging effect Effects 0.000 title claims abstract description 11
- 238000002360 preparation method Methods 0.000 title abstract description 11
- 230000000694 effects Effects 0.000 claims abstract description 24
- 238000004321 preservation Methods 0.000 claims description 7
- 238000001727 in vivo Methods 0.000 claims description 5
- 241000252212 Danio rerio Species 0.000 abstract description 49
- 230000036542 oxidative stress Effects 0.000 abstract description 16
- 239000006041 probiotic Substances 0.000 abstract description 3
- 235000018291 probiotics Nutrition 0.000 abstract description 3
- 244000005700 microbiome Species 0.000 abstract description 2
- 230000000529 probiotic effect Effects 0.000 abstract description 2
- RWSXRVCMGQZWBV-WDSKDSINSA-N glutathione Chemical compound OC(=O)[C@@H](N)CCC(=O)N[C@@H](CS)C(=O)NCC(O)=O RWSXRVCMGQZWBV-WDSKDSINSA-N 0.000 description 19
- 102000019197 Superoxide Dismutase Human genes 0.000 description 18
- 108010012715 Superoxide dismutase Proteins 0.000 description 18
- HYPYXGZDOYTYDR-HAJWAVTHSA-N 2-methyl-3-[(2e,6e,10e,14e)-3,7,11,15,19-pentamethylicosa-2,6,10,14,18-pentaenyl]naphthalene-1,4-dione Chemical compound C1=CC=C2C(=O)C(C/C=C(C)/CC/C=C(C)/CC/C=C(C)/CC/C=C(C)/CCC=C(C)C)=C(C)C(=O)C2=C1 HYPYXGZDOYTYDR-HAJWAVTHSA-N 0.000 description 12
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 12
- 235000019143 vitamin K2 Nutrition 0.000 description 12
- 239000011728 vitamin K2 Substances 0.000 description 12
- 229940041603 vitamin k 3 Drugs 0.000 description 12
- 239000013641 positive control Substances 0.000 description 10
- 210000004027 cell Anatomy 0.000 description 9
- 229960003180 glutathione Drugs 0.000 description 9
- 239000001963 growth medium Substances 0.000 description 8
- 239000000243 solution Substances 0.000 description 8
- 239000000047 product Substances 0.000 description 7
- 230000001580 bacterial effect Effects 0.000 description 6
- 238000011534 incubation Methods 0.000 description 6
- 230000003647 oxidation Effects 0.000 description 5
- 238000007254 oxidation reaction Methods 0.000 description 5
- 150000003254 radicals Chemical class 0.000 description 5
- 239000007787 solid Substances 0.000 description 5
- 108020004465 16S ribosomal RNA Proteins 0.000 description 4
- 230000032683 aging Effects 0.000 description 4
- 238000000576 coating method Methods 0.000 description 4
- 239000012895 dilution Substances 0.000 description 4
- 238000010790 dilution Methods 0.000 description 4
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 3
- 102000004316 Oxidoreductases Human genes 0.000 description 3
- 108090000854 Oxidoreductases Proteins 0.000 description 3
- 239000011248 coating agent Substances 0.000 description 3
- 238000012258 culturing Methods 0.000 description 3
- 238000001514 detection method Methods 0.000 description 3
- 239000007788 liquid Substances 0.000 description 3
- 238000009630 liquid culture Methods 0.000 description 3
- 239000000725 suspension Substances 0.000 description 3
- 241000251468 Actinopterygii Species 0.000 description 2
- 241000186000 Bifidobacterium Species 0.000 description 2
- 230000003064 anti-oxidating effect Effects 0.000 description 2
- 239000007853 buffer solution Substances 0.000 description 2
- 238000004113 cell culture Methods 0.000 description 2
- 239000006285 cell suspension Substances 0.000 description 2
- 239000003085 diluting agent Substances 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 210000003608 fece Anatomy 0.000 description 2
- 239000012634 fragment Substances 0.000 description 2
- 230000004060 metabolic process Effects 0.000 description 2
- 238000009629 microbiological culture Methods 0.000 description 2
- 239000008188 pellet Substances 0.000 description 2
- 239000002504 physiological saline solution Substances 0.000 description 2
- 230000000270 postfertilization Effects 0.000 description 2
- 239000002244 precipitate Substances 0.000 description 2
- 238000012545 processing Methods 0.000 description 2
- 102000004169 proteins and genes Human genes 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- 239000012898 sample dilution Substances 0.000 description 2
- 239000011550 stock solution Substances 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 210000001519 tissue Anatomy 0.000 description 2
- PXEZTIWVRVSYOK-UHFFFAOYSA-N 2-(3,6-diacetyloxy-2,7-dichloro-9h-xanthen-9-yl)benzoic acid Chemical compound C1=2C=C(Cl)C(OC(=O)C)=CC=2OC2=CC(OC(C)=O)=C(Cl)C=C2C1C1=CC=CC=C1C(O)=O PXEZTIWVRVSYOK-UHFFFAOYSA-N 0.000 description 1
- 229920001817 Agar Polymers 0.000 description 1
- 108020004414 DNA Proteins 0.000 description 1
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 1
- 108010024636 Glutathione Proteins 0.000 description 1
- 241000237903 Hirudo Species 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- 101150053185 P450 gene Proteins 0.000 description 1
- 230000003213 activating effect Effects 0.000 description 1
- 230000004913 activation Effects 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 239000003242 anti bacterial agent Substances 0.000 description 1
- 229940088710 antibiotic agent Drugs 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- 230000030833 cell death Effects 0.000 description 1
- 210000000170 cell membrane Anatomy 0.000 description 1
- 210000003855 cell nucleus Anatomy 0.000 description 1
- 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 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 230000002708 enhancing effect Effects 0.000 description 1
- 239000007850 fluorescent dye Substances 0.000 description 1
- 230000012010 growth Effects 0.000 description 1
- 230000036039 immunity Effects 0.000 description 1
- 230000000968 intestinal effect Effects 0.000 description 1
- 230000003834 intracellular effect Effects 0.000 description 1
- 230000006799 invasive growth in response to glucose limitation Effects 0.000 description 1
- 238000002955 isolation Methods 0.000 description 1
- 230000003859 lipid peroxidation Effects 0.000 description 1
- 229920002521 macromolecule Polymers 0.000 description 1
- 239000002609 medium Substances 0.000 description 1
- 238000000034 method Methods 0.000 description 1
- 230000003228 microsomal effect Effects 0.000 description 1
- 210000003470 mitochondria Anatomy 0.000 description 1
- 230000008811 mitochondrial respiratory chain Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 230000000877 morphologic effect Effects 0.000 description 1
- 108020004707 nucleic acids Proteins 0.000 description 1
- 102000039446 nucleic acids Human genes 0.000 description 1
- 150000007523 nucleic acids Chemical class 0.000 description 1
- 239000007800 oxidant agent Substances 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 108010008359 protein kinase C lambda Proteins 0.000 description 1
- 239000003642 reactive oxygen metabolite Substances 0.000 description 1
- 230000011506 response to oxidative stress Effects 0.000 description 1
- 230000002000 scavenging effect Effects 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 238000012163 sequencing technique Methods 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 238000007619 statistical method Methods 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 230000000007 visual effect Effects 0.000 description 1
- 229940088594 vitamin Drugs 0.000 description 1
- 235000013343 vitamin Nutrition 0.000 description 1
- 239000011782 vitamin Substances 0.000 description 1
- 229930003231 vitamin Natural products 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
- 108700024526 zebrafish sox32 Proteins 0.000 description 1
Classifications
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
Landscapes
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
Abstract
The invention discloses application of bifidobacterium longum LF04 in preparation of antioxidant and anti-aging products, and belongs to the technical field of microorganisms. The bifidobacterium longum LF04 disclosed by the invention has the potential of obviously reducing the ROS level in the zebra fish body and obviously improving the SOD activity in the zebra fish body in a zebra fish oxidative stress model, and provides theoretical reference and guiding basis for developing an antioxidant probiotic preparation by using the bifidobacterium longum LF 04.
Description
Technical Field
The invention relates to the technical field of microorganisms, in particular to application of bifidobacterium longum LF04 in preparation of antioxidant and anti-aging products.
Background
Oxidation is an essential metabolic process of the body, but excessive oxidation causes damage to biological macromolecules. Oxidative stress is the root cause of aging and aging-related diseases in the body. The free radical has strong oxidation effect, and can trigger lipid peroxidation to damage cell membrane, thereby leading to cell death. When the human body has excessive free radical generation, the free radical can combine with biomacromolecules such as protein, nucleic acid and the like in cells of the human body and destroy the structure and the function of the biomacromolecules. The probiotics can regulate intestinal flora balance, improve metabolism of protein and vitamins, generate antibiotics to inhibit growth of harmful flora, resist tumor, enhance immunity, reduce serum cholesterol, resist oxidation, delay aging, etc.
Therefore, providing the use of bifidobacterium longum LF04 in the preparation of antioxidant and anti-aging products is a problem to be solved by the person skilled in the art.
Disclosure of Invention
In view of this, the present invention provides the use of bifidobacterium longum LF04 in the preparation of antioxidant and anti-ageing products.
Menaquinone is an oxidizing agent that produces unstable semiquinones through the intracellular reductase system (microsomal P450 reductase and mitochondrial respiratory chain reductase), which enter the redox cycle, producing reactive oxygen species. Menaquinone can induce zebra fish to establish an oxidative stress model.
Through specific fluorescent staining (green, mainly located in cell nuclei and mitochondria), the whole body of the zebra fish subjected to oxidative stress reaction is obviously much more green than that of normal zebra fish, and the active oxygen content in the zebra fish can be observed under a fluorescent microscope.
In order to achieve the above purpose, the present invention adopts the following technical scheme:
The application of bifidobacterium longum (Bifidobacterium longum) LF04 in preparing antioxidant and anti-aging products is characterized in that the preservation number of the bifidobacterium longum LF04 is CGMCC No.23316, the bifidobacterium longum LF04 is preserved in China general microbiological culture Collection center (CGMCC), the institute of microbiology, national academy of sciences 3, the Xiya 1, north Star, the Beijing city, and the preservation date is 2021, 08, 30, and the bifidobacterium longum Bifidobacterium longum.
Further, the bifidobacterium longum LF04 is applied to the preparation of products for reducing the ROS level in vivo and improving the SOD activity in vivo.
Further, the bifidobacterium longum LF04 is a bacterial suspension.
The bifidobacterium longum LF04 can obviously reduce the ROS level in the zebra fish body and obviously improve the SOD activity in the zebra fish body in an in-vivo oxidative stress model, enhances the capability of the organism for scavenging free radicals, and has good antioxidant and anti-aging effects.
Compared with the prior art, the application of the bifidobacterium longum LF04 in preparing antioxidant and anti-aging products is disclosed, the bifidobacterium longum LF04 has the potential of obviously reducing the ROS level in the zebra fish body and obviously improving the SOD activity in the zebra fish body in a zebra fish oxidative stress model, and theoretical reference and guiding basis are provided for developing antioxidant probiotic preparations by using the bifidobacterium longum LF 04.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings that are required to be used in the embodiments or the description of the prior art will be briefly described below, and it is obvious that the drawings in the following description are only embodiments of the present invention, and that other drawings can be obtained according to the provided drawings without inventive effort for a person skilled in the art.
FIG. 1 is a drawing showing the colony morphology of bifidobacterium longum LF04 on BS agar plates;
FIG. 2 is a visual graph showing the effect of bifidobacterium longum LF04 on ROS levels in a menaquinone-induced zebra fish oxidative stress model;
Wherein A: normal group; b: a model group; c: a positive control group; d: 1X 10 6 CFU/mL Bifidobacterium longum 15708; e: 1X 10 6 CFU/mL Bifidobacterium longum LF04;
FIG. 3 is a graph showing the statistical effect of bifidobacterium longum LF04 on ROS levels in a menaquinone-induced zebra fish oxidative stress model;
FIG. 4 is a graph showing the effect of bifidobacterium longum LF04 on SOD activity in menaquinone-induced zebra fish oxidative stress models.
Detailed Description
The following description of the embodiments of the present invention will be made clearly and completely with reference to the accompanying drawings, in which it is apparent that the embodiments described are only some embodiments of the present invention, but not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
Reduced Glutathione (GSH), menaquinone, dimethyl sulfoxide (DMSO) were all purchased from shanghai source leaf biotechnology limited; 2',7' -dichloro-dihydro-fluorescein diacetate (DCFH-DA) and superoxide dismutase (SOD) detection kits were purchased from Sigma-Aldrich company; bifidobacterium longum 15708 (ATCC 15708) was purchased from Beijing Bai Oubo Biotechnology Co.
EXAMPLE 1 isolation, identification and preservation of bifidobacterium longum LF04
(1) Separating:
1) The feces (about 0.1 g) of the elder with long life are dissolved in a 1.5mL centrifuge tube filled with 1mL sterile physiological saline, and are fully blown and evenly mixed by a 1mL sterile gun head for standby.
2) Into each of 6 sterile 1.5mL centrifuge tubes, 900. Mu.L of sterile physiological saline was added.
3) From the 1 st centrifuge tube containing 10 -1 sample dilutions, 100 μl of the liquid was pipetted into the 2 nd centrifuge tube (10 -2) and diluted to 10 -2;
4) From the 2 nd centrifuge tube containing 10 -2 sample dilutions, 100 μl of liquid was pipetted into the 3 rd centrifuge tube (10 -3) and diluted to 10 -3;
5) The above steps were repeated until the dilution was 10 -4、10-5、10-6、10-7.
6) 100 Mu L of sample diluent is sucked from a centrifuge tube filled with 10 -4 sample diluent and respectively inoculated to MRS solid culture medium, BS solid culture medium and BHI solid culture medium, and 100 mu L of bacterial liquid is spread and dried, and the coating method is mild, fast in action and needs to be operated near the flame of an alcohol lamp. After coating, the side of the dish was marked, including information on name, sample number, medium name, incubation time, dilution gradient, incubation conditions (anaerobic/aerobic), etc.
7) The above steps were repeated to complete the dilution coating at a dilution gradient of 10 -5、10-6、10-7.
8) After the coating, the dishes were cultured at 37℃under anaerobic conditions for 48 hours, and then were subjected to observation and recording.
9) Single colony on the plate is picked up by an inoculating loop and streaked into a BS solid culture medium, anaerobic culture is carried out for 48 hours at 37 ℃, and pure colony is obtained by separation.
10 Inoculating pure bacterial colony on the flat plate into BS liquid culture medium, anaerobic culturing at 37 ℃ for 12-16 h, adding 20% glycerol, and storing in a refrigerator at-80 ℃.
(2) Molecular biological identification of strains: genomic DNA was extracted from the obtained strain, and a full-length fragment of 16S rDNA was amplified by PCR technique using the universal primers 27F and 1492R of 16S rDNA, followed by sequencing to identify the species of the strain.
The primer sequences of the universal primers 27F and 1492R are as follows:
27F：5’-AGAGTTTGATCCTGGCTCAG-3’；SEQ ID NO.1；
1492R：5’-GGTTACCTTGTTACGACTT-3’；SEQ ID NO.2。
Experimental results: the strain screened from feces of the elderly in Jiangling county of plum state, guangdong province is identified by morphological observation and 16S rDNA, wherein the strain LF04 is identified as bifidobacterium longum, and the 16S rDNA sequence is shown as SEQ ID NO. 3.
TGCAAGTCGAACGGGATCCATCAGGCTTTGCTTGGTGGTGAGAGTGGCGAACGGGTGAGTAATGCGTGACCGACCTGCCCCATACACCGGAATAGCTCCTGGAAACGGGTGGTAATGCCGGATGCTCCAGTTGATCGCATGGTCTTCTGGGAAAGCTTTCGCGGTATGGGATGGGGTCGCGTCCTATCAGCTTGACGGCGGGGTAACGGCCCACCGTGGCTTCGACGGGTAGCCGGCCTGAGAGGGCGACCGGCCACATTGGGACTGAGATACGGCCCAGACTCCTACGGGAGGCAGCAGTGGGGAATATTGCACAATGGGCGCAAGCCTGATGCAGCGACGCCGCGTGAGGGATGGAGGCCTTCGGGTTGTAAACCTCTTTTATCGGGGAGCAAGCGAGAGTGAGTTTACCCGTTGAATAAGCACCGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGGTGCAAGCGTTATCCGGAATTATTGGGCGTAAAGGGCTCGTAGGCGGTTCGTCGCGTCCGGTGTGAAAGTCCATCGCTTAACGGTGGATCCGCGCCGGGTACGGGCGGGCTTGAGTGCGGTAGGGGAGACTGGAATTCCCGGTGTAACGGTGGAATGTGTAGATATCGGGAAGAACACCAATGGCGAAGGCAGGTCTCTGGGCCGTTACTGACGCTGAGGAGCGAAAGCGTGGGGAGCGAACAGGATTAGATACCCTGGTAGTCCACGCCGTAAACGGTGGATGCTGGATGTGGGGCCCGTTCCACGGGTTCCGTGTCGGAGCTAACGCGTTAAGCATCCCGCCTGGGGAGTACGGCCGCAAGGCTAAAACTCAAAGAAATTGACGGGGGCCCGCACAAGCGGCGGAGCATGCGGATTAATTCGATGCAACGCGAAGAACCTTACCTGGGCTTGACATGTTCCCGACGGTCGTAGAGATACGGCTTCCCTTCGGGGCGGGTTCACAGGTGGTGCATGGTCGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTCGCCCCGTGTTGCCAGCGGATTATGCCGGGAACTCACGGGGGACCGCCGGGGTTAACTCGGAGGAAGGTGGGGATGACGTCAGATCATCATGCCCCTTACGTCCAGGGCTTCACGCATGCTACAATGGCCGGTACAACGGGATGCGACGCGGCGACGCGGAGCGGATCCCTGAAAACCGGTCTCAGTTCGGATCGCAGTCTGCAACTCGACTGCGTGAAGGCGGAGTCGCTAGTAATCGCGAATCAGCAACGTCGCGGTGAATGCGTTCCCGGGCCTTGTACACACCGCCCGTCAAGTCATGAAAGTGGGCAGCACCCGAAGCCGGTGGCCTAACCCCTTGTGGGATGAGCCGTCTAA;SEQ ID NO.3.
The strain LF04 single colony is inoculated on a BS solid culture medium, grows well under anaerobic conditions at 37 ℃, and has milky white, spherical, neat-edged and smooth surface (figure 1). The strain BS is preserved in China general microbiological culture Collection center (CGMCC), and has the preservation date of 2021, 08 and 30 days, the classification name of bifidobacterium longum Bifidobacterium longum and the preservation number of CGMCC No.23316, wherein the CGMCC is called No. 1 and No.3 of West Hirudo in the Korean region North Star of Beijing.
EXAMPLE 2 preparation of Bifidobacterium longum LF04 bacterial suspension (thallus)
Inoculating bifidobacterium longum LF04 into a BS liquid culture medium after activating and culturing for 24 hours at 37 ℃, and centrifuging for 10 minutes at 4 ℃ and 6000r/min to obtain bacterial precipitate; after the cell pellet was washed twice with PBS, the cell was resuspended in PBS, and the cell concentration was adjusted to 1X 10 6 CFU/mL to obtain a cell suspension (cell).
EXAMPLE 3 preparation of Bifidobacterium longum 15708 suspension (thallus)
Inoculating bifidobacterium longum 15708 after activation culture in a BS liquid culture medium, culturing at 37 ℃ for 24 hours, and centrifuging at 4 ℃ for 10 minutes at 6000r/min to obtain bacterial precipitate; after the cell pellet was washed twice with PBS, the cell was resuspended in PBS, and the cell concentration was adjusted to 1X 10 6 CFU/mL to obtain a cell suspension (cell).
Example 4 Effect of Bifidobacterium longum LF04 on ROS levels in the zebra fish oxidative stress model
Healthy wild-type AB zebra fish that developed to 4dpf (days post fertilization) were selected and placed in 6-well cell culture plates with 20 fish per well. The experiments set up a normal group, a model group, a positive control Group (GSH), a bifidobacterium longum 15708 intervention group, and a bifidobacterium longum LF04 intervention group. PBS was added to both the normal and model groups, GSH solution (100. Mu.M) was added to the positive control group, and bifidobacterium longum 15708 was added to the intervention group (1X 10 6 CFU/mL) and 1X 10 6 CFU/M bifidobacterium longum 15708; bifidobacterium longum LF04 intervention group (1×10 6 CFU/mL) 1×10 6 CFU/mL Bifidobacterium longum LF04 was added, incubated at 28℃2.5mL per well, and new solution was changed after every 24 h; after 48h incubation, 2.5mL of PBS (1% DMSO) was added to the normal group, and 6 μm menaquinone was added to the model group, positive control group, bifidobacterium longum 15708 intervention group, bifidobacterium longum LF04 intervention group, respectively (menaquinone was first formulated with DMSO as 600 μm stock solution and then diluted with PBS to 6 μm), 2.5mL per well; after 24h incubation at 28 ℃, the solution is discarded, the zebra fish is washed 3 times by PBS, 20 mug/mL DCFH-DA solution is added, 3mL of each hole is incubated for 1h at 28 ℃ in a dark place, the zebra fish is washed 3 times by PBS, and the fluorescence intensity in the zebra fish is observed under a fluorescence microscope and recorded by photographing. Quantitative statistical analysis of fluorescence intensity (S) in zebra fish was performed using Image J software. ROS levels in zebra fish were calculated as follows:
SPSS 19.0 software is adopted for statistical data processing, and experimental data are all adopted Data represent, analyzed by T-test, compared to normal group: ### P <0.005, compared to model group: *** P <0.005.
The results are shown in fig. 2 and 3; as can be seen from fig. 2 and 3, the intensity of green fluorescence in the zebra fish body reflects the level of ROS; compared with the normal group, the green fluorescence intensity in the zebra fish body of the model group is enhanced, which indicates that the ROS level in the zebra fish body of the model group is increased; meanwhile, compared with a normal group (100.00+/-7.21%), the ROS level (211.80 +/-10.19%) in the zebra fish body of the model group is obviously increased (p < 0.005), which indicates that the current zebra fish oxidative stress model is successfully established.
Compared with the model group, the positive control Group (GSH) has reduced green fluorescence intensity in the zebra fish body, which indicates that GSH can reduce the ROS level in the zebra fish body in a menaquinone induced zebra fish oxidative stress model; meanwhile, the ROS level in the zebra fish of the positive control group is 114.22 +/-6.37%, and the difference is obvious (P < 0.005) compared with the model group (211.80 +/-10.19 percent); thus, GSH has a pronounced antioxidant effect, consistent with clinical results. Bifidobacterium longum 15708 intervention group (1X 10 6 CFU/mL) red fluorescence intensity in zebra fish was similar to model group; the ROS levels in zebra fish of the same duration bifidobacterium 15708 intervention group (1 x 10 6 CFU/mL) were 205.60 ±9.92%, with no significant differences (P > 0.05) compared to the model group (211.80 ±10.19%). Compared with the model group, the green fluorescence intensity of the bifidobacterium longum LF04 in the zebra fish body is weakened, which shows that the bifidobacterium longum LF04 can reduce the ROS level in the zebra fish body in a menaquinone-induced zebra fish oxidative stress model; the ROS level in zebra fish of the same duration bifidobacterium LF04 intervention group (1 x 10 6 CFU/mL) was 126.78 ±9.72%, with significant worse variability (P < 0.005) compared to the model group (211.80 ±10.19%). Therefore, the results show that at the same concentration, the bifidobacterium longum LF04 has stronger effect of reducing the ROS level in the zebra fish body than the bifidobacterium longum 15708 in the body oxidative stress model, and has good anti-oxidation and anti-aging effects.
Example 5 Effect of Bifidobacterium longum LF04 on SOD Activity in a zebra fish oxidative stress model
Healthy wild-type AB zebra fish that developed to 4dpf (days post fertilization) were selected and placed in 6-well cell culture plates with 20 fish per well. The experiments set up a normal group, a model group, a positive control Group (GSH), a bifidobacterium longum 15708-interfered group, and a bifidobacterium longum LF 04-interfered group, 3 duplicate wells per group. PBS was added to both the normal and model groups, GSH solution (100. Mu.M) was added to the positive control group, and bifidobacterium longum 15708 was added to the intervention group (1X 10 6 CFU/mL) and 1X 10 6 CFU/mL bifidobacterium longum 15708; bifidobacterium longum LF04 intervention group (1×10 6 CFU/mL) 1×10 6 CFU/mL Bifidobacterium longum LF04 was added, incubated at 28℃2.5mL per well, and new solution was changed after every 24 h; after 48h incubation, 2.5mL of PBS (1% DMSO) was added to the normal group, and 6 μm menaquinone was added to the model group, positive control group, bifidobacterium longum 15708 intervention group, bifidobacterium longum LF04 intervention group, respectively (menaquinone was first formulated with DMSO as 600 μm stock solution and then diluted with PBS to 6 μm), 2.5mL per well; after 24h incubation at 28 ℃, the solution was discarded, the zebra fish was washed 3 times with PBS, and the zebra fish was collected into 1.5mL centrifuge tubes, 50mg zebra fish per tube, 6 tubes per experimental group; after the water in the centrifuge tube was sucked dry, 250. Mu.L of a buffer solution (a buffer solution of a superoxide dismutase (SOD) detection kit) was added. The zebra fish homogenate was broken up by holding a micro-electric tissue homogenizer with S-18KS until no distinct tissue fragments were present, centrifuged at 15000 Xg at 4℃for 15min and the supernatant was collected. The SOD activity of each group was measured using a superoxide dismutase (SOD) detection kit (Sigma-Aldrich).
SPSS 19.0 software is adopted for statistical data processing, and experimental data are all adoptedData represent, analyzed by T-test, compared to normal group: ### P <0.005, compared to model group: *** P <0.005.
The results are shown in FIG. 4; as can be seen from FIG. 4, compared with the normal group (2.79+ -0.17U/mg), the SOD activity (0.82+ -0.05U/mg) in the zebra fish of the model group is significantly reduced (p < 0.005), which indicates that the current zebra fish oxidative stress model is successfully established.
The SOD activity in the zebra fish of the positive control group is 2.17+/-0.13U/mg, and the difference is obvious (P < 0.005) compared with the model group (0.82+/-0.05U/mg), which shows that GSH has obvious antioxidation effect and is consistent with clinical results. The SOD activity in zebra fish of the bifidobacterium longum 15708 intervention group (1X 10 6 CFU/mL) is 0.98+/-0.11U/mg, and compared with the model group (0.82+/-0.05U/mg), the SOD activity in zebra fish has no significant difference (P > 0.05). Whereas the activity of SOD in zebra fish in the interference group of bifidobacterium longum LF04 (1X 10 6 CFU/mL) is 1.98+/-0.13U/mg, the dissimilarity is obvious (P < 0.005) compared with the model group (0.82+/-0.05U/mg). Therefore, the result shows that at the same concentration, the bifidobacterium longum LF04 has stronger effect of improving the SOD activity in the zebra fish body than the bifidobacterium longum 15708 in the in-vivo oxidative stress model, namely the capability of enhancing the body to remove free radicals, and has good effects of resisting oxidation and aging.
The previous description of the disclosed embodiments is provided to enable any person skilled in the art to make or use the present invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the generic principles defined herein may be applied to other embodiments without departing from the spirit or scope of the invention. Thus, the present invention is not intended to be limited to the embodiments shown herein but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.
Claims (2)
1. The application of bifidobacterium longum (Bifidobacterium longum) LF04 in preparing antioxidant and anti-aging products is characterized in that the preservation number of the bifidobacterium longum LF04 is CGMCC No.23316.
2. The application of bifidobacterium longum LF04 in preparing a product for reducing the ROS level and/or improving the SOD activity in vivo is characterized in that the preservation number of the bifidobacterium longum LF04 is CGMCC No.23316.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202310731195.6A CN116694528B (en) | 2023-06-20 | 2023-06-20 | Application of bifidobacterium longum LF04 in preparation of antioxidant and anti-aging products |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202310731195.6A CN116694528B (en) | 2023-06-20 | 2023-06-20 | Application of bifidobacterium longum LF04 in preparation of antioxidant and anti-aging products |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN116694528A CN116694528A (en) | 2023-09-05 |
| CN116694528B true CN116694528B (en) | 2024-08-02 |
Family
ID=87833698
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202310731195.6A Active CN116694528B (en) | 2023-06-20 | 2023-06-20 | Application of bifidobacterium longum LF04 in preparation of antioxidant and anti-aging products |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN116694528B (en) |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN115279385A (en) * | 2020-01-15 | 2022-11-01 | 佰欧波利斯有限公司 | Probiotic compositions as antioxidants |
Family Cites Families (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20170232047A1 (en) * | 2012-01-16 | 2017-08-17 | Elizabeth McKenna | Control of Cellular Redox Levels |
| EP4091673A4 (en) * | 2020-01-17 | 2024-01-31 | Morinaga Milk Industry Co., Ltd. | Anti-aging composition |
-
2023
- 2023-06-20 CN CN202310731195.6A patent/CN116694528B/en active Active
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN115279385A (en) * | 2020-01-15 | 2022-11-01 | 佰欧波利斯有限公司 | Probiotic compositions as antioxidants |
Also Published As
| Publication number | Publication date |
|---|---|
| CN116694528A (en) | 2023-09-05 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN111849836B (en) | Lactobacillus rhamnosus with antioxidant function and application thereof | |
| CN114790430B (en) | Lactobacillus rhamnosus E2 for producing hyaluronic acid and application thereof | |
| CN114507622B (en) | Bacillus amyloliquefaciens WF2020 strain product and application thereof | |
| CN114574393B (en) | Lactobacillus delbrueckii SEUNEU-110 and application thereof in skin | |
| CN114591861B (en) | Lactobacillus fermentum WC2020 and application thereof | |
| CN116162569A (en) | Bifidobacterium animalis subspecies BL03 and application thereof | |
| CN117645941A (en) | Fermented lactobacillus mucilaginosus capable of relieving hyperuricemia through simultaneous degradation of uric acid and purine nucleoside and application thereof | |
| CN113151072A (en) | Bifidobacterium breve NX-5 and application thereof in antioxidation | |
| CN115093985A (en) | Bifidobacterium lactis and fermentation method and application thereof | |
| CN107653199B (en) | Healthy human intestinal escherichia coli and application thereof | |
| CN116622584A (en) | Application of lactobacillus gasseri LS03 in preparation of antioxidant and anti-aging products | |
| CN116769661A (en) | Application of lactobacillus reuteri E9 in preparation of antioxidant and anti-aging products | |
| CN115786190B (en) | Lactobacillus plantarum capable of producing urolithin A and resisting aging and application thereof | |
| CN116694528B (en) | Application of bifidobacterium longum LF04 in preparation of antioxidant and anti-aging products | |
| CN116790430A (en) | Application of lactobacillus salivarius LS08 in preparation of antioxidant and anti-aging products | |
| KR101201420B1 (en) | A feed additive containing novel Lactobacillus jonhsonnii | |
| CN112877231A (en) | Lactic acid bacteria with bacteriostatic and antioxidant activity and application thereof | |
| CN114854623B (en) | Lactobacillus delbrueckii subspecies bulgaricus, microbial inoculum containing lactobacillus delbrueckii subspecies bulgaricus and application of lactobacillus delbrueckii subspecies bulgaricus | |
| CN117089480B (en) | Bacillus marinus BH03 and application thereof in preparation of antioxidant and anti-aging products | |
| CN115992079A (en) | Lactobacillus fermentum E1 and application thereof in preparation of uric acid reducing drugs | |
| CN116286507A (en) | Lactobacillus fermentum E1 and application thereof in preparation of medicines for preventing or treating intestinal inflammation | |
| CN115812968A (en) | Application of lactobacillus harbin in antioxidation, anti-aging and fat reduction | |
| CN115466702A (en) | Intestinal probiotic and application thereof in preventing and treating atrial fibrillation | |
| Lavanya et al. | Isolation and characterization of probiotic bacteria from the soil samples of the coastal areas of (Gudur division, Nellore Dt.) for utilization in Shrimp farming | |
| CN117187102A (en) | Application of bacillus subtilis BS02 in preparation of antioxidant and anti-aging products |
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 |