CN115478028B - Lactobacillus acidophilus F02, preparation prepared from lactobacillus acidophilus F02 and application of lactobacillus acidophilus F02 - Google Patents
Lactobacillus acidophilus F02, preparation prepared from lactobacillus acidophilus F02 and application of lactobacillus acidophilus F02 Download PDFInfo
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- 238000002360 preparation method Methods 0.000 title claims abstract description 8
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- 229940116269 uric acid Drugs 0.000 claims abstract description 22
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- PUNSOBLTEWFAMJ-UHFFFAOYSA-N 4-(diethylamino)benzohydrazide Chemical compound CCN(CC)C1=CC=C(C(=O)NN)C=C1 PUNSOBLTEWFAMJ-UHFFFAOYSA-N 0.000 description 1
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- 208000027418 Wounds and injury Diseases 0.000 description 1
- 108010093894 Xanthine oxidase Proteins 0.000 description 1
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- RQFCJASXJCIDSX-UUOKFMHZSA-N guanosine 5'-monophosphate Chemical compound C1=2NC(N)=NC(=O)C=2N=CN1[C@@H]1O[C@H](COP(O)(O)=O)[C@@H](O)[C@H]1O RQFCJASXJCIDSX-UUOKFMHZSA-N 0.000 description 1
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- 229940039696 lactobacillus Drugs 0.000 description 1
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- 102000004169 proteins and genes Human genes 0.000 description 1
- 239000002213 purine nucleotide Substances 0.000 description 1
- 150000003212 purines Chemical class 0.000 description 1
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- SUKJFIGYRHOWBL-UHFFFAOYSA-N sodium hypochlorite Chemical compound [Na+].Cl[O-] SUKJFIGYRHOWBL-UHFFFAOYSA-N 0.000 description 1
- 208000024891 symptom Diseases 0.000 description 1
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- A61K35/66—Microorganisms or materials therefrom
- A61K35/74—Bacteria
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- A61K35/744—Lactic acid bacteria, e.g. enterococci, pediococci, lactococci, streptococci or leuconostocs
- A61K35/747—Lactobacilli, e.g. L. acidophilus or L. brevis
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- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P19/00—Drugs for skeletal disorders
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Abstract
The invention belongs to the field of microorganisms and application thereof, and discloses lactobacillus acidophilus F02, a preparation prepared from the lactobacillus acidophilus F02 and application of the lactobacillus acidophilus F. The lactobacillus acidophilus F02 is preserved in China Center for Type Culture Collection (CCTCC) M2022088 in 2022, 01 and 18 days. The lactobacillus acidophilus F02 can efficiently degrade purine metabolic intermediates inosine and guanosine in vitro, effectively control uric acid, creatinine and urea nitrogen levels in serum, can be used for preparing an auxiliary pharmaceutical composition for reducing uric acid, has the advantages of low cost, strong safety, good degradation efficiency and the like, and has very wide potential development prospect.
Description
Technical Field
The invention belongs to the field of microorganisms and application thereof, and particularly relates to lactobacillus acidophilus F02, a preparation prepared from the lactobacillus acidophilus F02 and application of the lactobacillus acidophilus.
Background
Hyperuricemia is a metabolic disorder caused by purine metabolic disorder and unbalanced uric acid formation and secretion, and is typically characterized by abnormally elevated uric acid levels in the blood, i.e., greater than 0.36mmol/L (6.0 mg/dL) in blood for females and greater than 0.42mmol/L (7.0 mg/dL) in males. With the continuous improvement of the living standard of people, the life style and the diet mode are changed, the food intake of high protein, high fructose and high purine is continuously increased, the incidence of hyperuricemia in the people is continuously increased, and the sick age tends to be younger. It is a matter of statistics that,
the prevalence rate of hyperuricemia in China is over 13 percent. Hyperuricemia is not only the main risk factor for gout, but also induces kidney function injury, accelerates the development of type II diabetes mellitus, cardiovascular diseases and the like, and has become a second largest metabolic disease threatening human health.
Purine is an important substance mainly existing in human body in the form of purine nucleotide, and plays an important role in energy supply, metabolism regulation, composition coenzyme and the like. Because of the lack of uricase caused by human evolution, various derivatives containing the purine skeleton eventually form uric acid through a series of metabolic changes, rendering uric acid unable to metabolize into soluble allantoin. Thus, the intake of purine substances directly affects uric acid levels in the blood.
At present, two medicines for inhibiting uric acid generation and regulating uric acid transporter through competing xanthine oxidase to promote uric acid excretion are mainly used for clinically treating hyperuricemia, but the medicines can damage kidney and gastrointestinal tract functions and the like.
Disclosure of Invention
The invention aims to solve the defects in the prior art, and based on the defects, the inventor discovers a lactobacillus acidophilus F02 with the name of Lactobacillus acidophilus F, which is preserved in China center for type culture collection (CCTCC M2022088) in the year 2022, 01 and 18. The thallus of lactobacillus acidophilus F02 is in a rod shape, and the colony shape in an MRS agar plate culture medium is round, milky white, opaque and irregular in edge.
Lactobacillus acidophilus is one of the most important genera in the lactobacillus family, which is widely present in the human gastrointestinal tract, and it is currently reported in great numbers that lactobacillus acidophilus exists mainly as a probiotic agent for the intestinal tract. The lactobacillus acidophilus F02 discovered by the invention can have stronger degradation effect on purine metabolic intermediate products inosine and guanosine, and the degradation rate can reach 94% and 60% respectively; compared with lactobacillus gasseri OLL2922 (application number is CN 102016004A) with the accession number of NITE BP-462, the lactobacillus gasseri F02 can only reduce serum uric acid level to 15.2%, the lactobacillus acidophilus F02 can reduce uric acid level in serum to be higher, can reach 38% degradation rate, can also degrade creatinine and urea nitrogen level in serum effectively, has obvious relieving effect on serum indexes of mice with hyperuricemia induced by high fructose and adenine, can be used for preparing related preparations/medicines, can also be used in the food field, for example, can be used for producing fermented dairy products, fermented fruit and vegetable products and fermented bean products by using a starter containing lactobacillus acidophilus F02, and the starter can endow the products with specific fragrance of probiotics, and simultaneously prolong the storage time of the products and improve the nutritional value and the safety of the products.
The lactobacillus acidophilus F02 can achieve the effect of preventing hyperuricemia and gout, has the advantages of low cost, strong safety, good degradation efficiency and the like, and has very broad potential development prospect.
In the preparation of the preparation/medicament for reducing uric acid, creatinine and urea nitrogen levels in serum by using the lactobacillus acidophilus F02, the lactobacillus acidophilus F02 is a live bacterium or an inactivated bacterium. The concentration is 10 8 CFU/mL-10 9 CFU/mL。
The beneficial effects of the invention are as follows: the lactobacillus acidophilus F02 can efficiently degrade purine metabolic intermediates inosine and guanosine in vitro, effectively control uric acid, creatinine and urea nitrogen levels in serum, can be used for preparing an auxiliary pharmaceutical composition for reducing uric acid, has the advantages of low cost, strong safety, good degradation efficiency and the like, and has very wide potential development prospect.
Additional aspects and advantages of the invention will be set forth in part in the description which follows, and in part will be obvious from the description, or may be learned by practice of the invention.
Drawings
FIG. 1 is a graph showing colony characteristics of Lactobacillus acidophilus F02;
FIG. 2 is a graph showing the results of a phylogenetic tree of Lactobacillus acidophilus F02;
FIG. 3 is a graph showing the results of measuring the degradation rates of inosine and guanosine by Lactobacillus acidophilus F02 by HPLC in example 1;
FIG. 4 is a graph showing the results of measurement of serum uric acid from mice suffering from hyperuricemia by Lactobacillus acidophilus F02 in example 2;
FIG. 5 is a graph showing the measurement result of serum creatinine of mice with hyperuricemia by Lactobacillus acidophilus F02 in example 3;
FIG. 6 is a graph showing the results of measurement of serum urea nitrogen of hyperuricemia mice by Lactobacillus acidophilus F02 in example 4.
Detailed Description
The conception and the technical effects produced by the present invention will be clearly and completely described below with reference to the embodiments and the drawings to fully understand the objects, aspects and effects of the present invention.
The colony characteristics of Lactobacillus acidophilus F02 are shown in FIG. 1, and the results of phylogenetic tree are shown in FIG. 2.
Example 1:
high Performance Liquid Chromatograph (HPLC) determines the ability of lactobacillus acidophilus F02 to degrade inosine and guanosine:
the frozen strain (Lactobacillus acidophilus F02, which is named Lactobacillus acidophilus F and was deposited in China center for type culture collection, no. CCTCC M2022088, for example, for year 2022, month 18) was removed from the refrigerator at-80 ℃, streaked into MRS plates with an inoculating loop, anaerobic cultured at 37℃for 24 hours, single colonies were picked up and inoculated into 5mL MRS liquid medium, and anaerobic cultured at 37℃for 12 hours. Subcultured twice at 1% inoculum size. 2mL of the activated bacterial liquid is taken and centrifuged at 6000r/min for 10min at 4 ℃, bacterial precipitate is collected, and the bacterial precipitate is washed three times with 1mL of physiological saline. To the bacterial cell pellet, 700. Mu.L of 3.78mmol/L inosine and guanosine phosphate buffer solution were added, and after mixing, anaerobic culture was performed at 37℃for 45 minutes. The culture broth was then centrifuged at 6000r/min at 4℃for 10min and filtered through a 0.22 μm filter to obtain a supernatant. 270. Mu.L of the supernatant was mixed with 30. Mu.L of a perchloric acid solution as a reaction terminator and analyzed by high performance liquid chromatography.
And (3) determining the retention time of the standard inosine and guanosine by an external standard method, preparing a standard curve according to different concentrations and retention time, and calculating the degradation rate of the lactobacillus acidophilus F02 on the inosine and guanosine according to the liquid chromatogram and the standard curve.
High performance liquid chromatography conditions: the high performance liquid chromatograph is LC-20AD, the reversed phase chromatographic column is ZORBAX SB-C18.6X1250 mm, and the mobile phase is 0.1 mu mol/L NaClO 4 -0.187mol/L H 3 PO 4 -ddH 2 0, the flow rate is 1mL/min, the column temperature is 25 ℃, the retention time is 10min, the measurement wavelength is 254nm, and the measurement result is shown in FIG. 1.
As shown in FIG. 3, the Lactobacillus acidophilus F02 provided by the invention can be used for strongly degrading purine metabolic intermediate products inosine and guanosine, and the degradation rate can reach 94% and 60% respectively.
Example 2:
testing the ability of lactobacillus acidophilus F02 to reduce uric acid in hyperuricemia mice:
30 7 week old Kunming male mice were selected and randomly assigned to the normal group, the hyperuricemia group and the Lactobacillus acidophilus group. Normal group drinking normal saline, hyperuricemia group and lactobacillus acidophilus group drinking 10% high fructose water and lavaging 50 mg/kg adenine according to weight standard to jointly induce for 24 days to form hyperuricemia model; after 25 days, normal group and hyperuricemia group were perfused with the same dose of physiological saline, and lactobacillus acidophilus group was perfused with lactobacillus acidophilus F02 (1.0X10) 9 CFU/mL). After 8d treatment, mice were sacrificed by cervical dislocation and blood was collected from the eyeballs. Blood collected from eyeballs is placed at room temperature for overnight coagulation, the temperature is 4 ℃, the blood is obtained by centrifugation at 4000 r/min for 10min, uric acid level in a serum sample is measured, and the measurement result is shown in figure 2.
As can be seen from FIG. 4, lactobacillus acidophilus F02 according to the present invention was effective in lowering uric acid levels in serum.
Example 3:
testing the ability of lactobacillus acidophilus F02 to reduce creatinine in hyperuricemia mice:
30 7 week old Kunming male mice were selected and randomly assigned to the normal group, the hyperuricemia group and the Lactobacillus acidophilus group. Normal group drinking normal saline, hyperuricemia group and lactobacillus acidophilus group drinking 10% high fructose water, and forming hyperuricemia model by combined induction of 50 mg/kg adenine according to weight standard; after 25 days, normal group and hyperuricemia group were perfused with the same dose of physiological saline, and lactobacillus acidophilus group was perfused with lactobacillus acidophilus F02 (1.0X10) 9 CFU/mL). After 8d treatment, mice were sacrificed by cervical dislocation and blood was collected from the eyeballs. Blood collected from eyeballs was allowed to coagulate overnight at room temperature, centrifuged at 4000 r/min for 10min at 4℃to obtain serum, and creatinine levels in the serum samples were measured as shown in FIG. 3.
As can be seen from FIG. 5, lactobacillus acidophilus F02 according to the present invention was effective in reducing serum creatinine levels.
Example 4:
testing the ability of lactobacillus acidophilus F02 to reduce urea nitrogen in hyperuricemia mice:
30 7 week old Kunming male mice were selected and randomly assigned to the normal group, the hyperuricemia group and the Lactobacillus acidophilus group. Normal group drinking normal saline, hyperuricemia group and lactobacillus acidophilus group drinking 10% high fructose water, and forming hyperuricemia model by combined induction of 50 mg/kg adenine according to weight standard; after 25 days, normal group and hyperuricemia group were perfused with the same dose of physiological saline, and lactobacillus acidophilus group was perfused with lactobacillus acidophilus F02 (1.0X10) 9 CFU/mL). After 8d treatment, mice were sacrificed by cervical dislocation and blood was collected from the eyeballs. Blood collected from eyeballs is placed at room temperature for overnight coagulation, serum is obtained by centrifugation at 4000 r/min for 10min at 4 ℃, urea nitrogen level in a serum sample is measured, and the measurement result is shown in fig. 4.
As can be seen from fig. 6, lactobacillus acidophilus F02 of the present invention was effective in reducing urea nitrogen levels in serum.
Intake of hyperuricemia mice 10 9 After CFU/mL of lactobacillus acidophilus F02, uric acid, creatinine and urea nitrogen levels in serum are effectively reduced, and hyperuricemia symptoms can be relieved.
The present invention is not limited to the above embodiments, but is merely preferred embodiments of the present invention, and the present invention should be construed as being limited to the above embodiments as long as the technical effects of the present invention are achieved by the same means. Various modifications and variations are possible in the technical solution and/or in the embodiments within the scope of the invention.
Claims (7)
1. Lactobacillus acidophilus F02, wherein the lactobacillus acidophilus F02 is named lactobacillus acidophilus (Lactobacillus acidophilus) F02 and has been deposited at the collection of chinese typical cultures at year 2022, month 01 and day 18, and has the number cctccc NO: m2022088.
2. Use of lactobacillus acidophilus F02 as claimed in claim 1 in the manufacture of a formulation for reducing uric acid levels in serum.
3. The use according to claim 2, wherein the lactobacillus acidophilus F02 is a live or inactivated bacterium.
4. Use of lactobacillus acidophilus F02 as claimed in claim 1 in the manufacture of a formulation for reducing creatinine levels in serum.
5. Use of lactobacillus acidophilus F02 as claimed in claim 1 in the manufacture of a formulation for reducing urea nitrogen levels in serum.
6. A preparation for reducing uric acid level in serum, comprising lactobacillus acidophilus F02 as defined in claim 1.
7. The formulation of claim 6, wherein the lactobacillus acidophilus F02 is at a concentration of 10 8 CFU/mL-10 9 CFU/mL。
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| CN114574382A (en) * | 2022-01-17 | 2022-06-03 | 南京农业大学 | Lactobacillus rhamnosus fmb14 with function of preventing or relieving hyperuricemia and application |
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| CN111388509A (en) * | 2019-01-03 | 2020-07-10 | 丰华生物科技股份有限公司 | Food composition and pharmaceutical composition of lactic acid bacteria strain for reducing uric acid concentration in blood |
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| JP2008005834A (en) * | 2006-05-31 | 2008-01-17 | Meiji Milk Prod Co Ltd | Lactobacillus having blood uric acid level-reducing activity |
| CA2718157A1 (en) * | 2007-11-29 | 2009-06-04 | Meiji Dairies Corporation | Lactobacillus gasseri capable of lowering blood uric acid level |
| WO2016002757A1 (en) * | 2014-06-30 | 2016-01-07 | 株式会社明治 | Lactic acid bacterium for suppressing purine absorption and use for same |
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| CN114574382A (en) * | 2022-01-17 | 2022-06-03 | 南京农业大学 | Lactobacillus rhamnosus fmb14 with function of preventing or relieving hyperuricemia and application |
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