CN115851508A - Streptococcus thermophilus JYST-26 capable of reducing oxalic acid and improving kidney stone, and product and application thereof - Google Patents
Streptococcus thermophilus JYST-26 capable of reducing oxalic acid and improving kidney stone, and product and application thereof Download PDFInfo
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- CN115851508A CN115851508A CN202211253422.0A CN202211253422A CN115851508A CN 115851508 A CN115851508 A CN 115851508A CN 202211253422 A CN202211253422 A CN 202211253422A CN 115851508 A CN115851508 A CN 115851508A
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- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
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Abstract
The invention relates to the technical field of probiotics, in particular to streptococcus thermophilus JYST-26 capable of reducing oxalic acid and improving kidney stone, and a product and application thereof. Streptococcus thermophilus (Streptococcus thermophilus) JYST-26 is preserved in China general microbiological culture Collection center on 27.06.2019, with the preservation address of No.3 of Xilu No.1 of Beijing Korean district, the preservation number of CGMCC NO.18045, and the 16S rRNA sequence of Streptococcus thermophilus JYST-26 is shown in SEQ ID NO. 3. The streptococcus thermophilus JYST-26 has strong intestinal tract colonization ability,can reduce the formation of calcium oxalate kidney stones and the dissolving capacity of the calcium oxalate kidney stones, and can avoid uncomfortable symptoms caused by medication and operation without any side effect.
Description
Technical Field
The invention relates to the technical field of probiotics, in particular to streptococcus thermophilus JYST-26 capable of reducing oxalic acid and improving kidney stone, and a product and application thereof.
Background
Urolithiasis is one of the common diseases of the urinary system, and can be divided into upper urinary tract stones (kidney stones and ureteral stones) and lower urinary tract stones (bladder stones and urethral stones). Calcium oxalate kidney stones are the common type of kidney stones, and have relatively high stone density and relatively hard texture. Often, the oxalic acid content in the body is too high, and the oxalic acid is combined with calcium in urine to form calcium oxalate calculi in a long time. At present, the treatment means of calcium oxalate kidney stones mainly comprise western medicine treatment, traditional Chinese medicine treatment, acupuncture treatment, surgical treatment and the like.
And (3) treatment with western medicines: patients with chronic urolithiasis often develop severe inflammation of the urethra or wing skin secondary to bacterial infection, even causing pyelonephritis, renal failure and sepsis. Therefore, the urinary calculus must be treated by matching with local and systemic antibiotics, such as ampicillin and compound sulfamethoxazole, or barking drugs, such as barktanadine. But the side effects of antibiotics are great.
Treatment with traditional Chinese medicines: the traditional Chinese medicine treatment mainly comprises clearing heat, promoting diuresis, treating stranguria and removing urinary calculus, and is assisted by regulating qi, activating blood circulation, softening hardness and dissipating stagnation. The application of the BAZHENG powder is more, and the other formulas include SHI SHUANG SHI TANG, HUASHI TANG, pig decoction, and HUASHI powder. Traditional Chinese medicine can only assist itself, and can not be used for rapid treatment of severe diseases.
Acupuncture and moxibustion treatment: acupuncture of corresponding acupuncture points can obviously expand renal pelvis, ureter and urethral smooth muscle, enhance renal function, increase renal blood flow, promote renal secretion function and increase urine volume. The bidirectional regulation of needle puncture makes ureter contract and relax, and can promote the peristalsis of kidney and ureter and expand ureter cavity, so as to decrease calculus and accelerate its discharge. But still requires the drug to be used together.
And (3) surgical treatment: the operation treatment effect is rapid, but the side effect is large, and part of patients cannot bear the operation process. And the recurrence rate of the operation is high, and the problem cannot be fundamentally solved.
Disclosure of Invention
Aiming at the technical problems of large side effect, high cost and the like of the existing treatment means of calcium oxalate kidney stone, the invention provides streptococcus thermophilus JYST-26 for reducing oxalic acid and improving kidney stone, and a product and application thereof. The streptococcus thermophilus JYST-26 strain provided by the invention has the functions of reducing oxalic acid and improving kidney stone.
In a first aspect, the invention provides a strain of Streptococcus thermophilus (S.) (Streptococcus thermophilus) JYST-26 has been deposited in China general microbiological culture Collection center (CGMCC) at 27.06.2019 at the No.3 of Xilu No.1 of the North Chen of the Chaoyang region in Beijing, with the deposition number of CGMCC No.18045, and the 16S rRNA sequence of Streptococcus thermophilus JYST-26 is shown in SEQ ID No. 3.
In a second aspect, the invention provides an application of streptococcus thermophilus JYST-26 in preparing a product for reducing oxalic acid and improving kidney stones.
In a third aspect, the invention provides an oxalic acid reducing and kidney stone improving product prepared from streptococcus thermophilus JYST-26, and a preparation method of the oxalic acid reducing and kidney stone improving product comprises the following steps:
(1) The streptococcus thermophilus JYST-26 is firstly activated on an MRS plate culture medium;
(2) Inoculating the activated streptococcus thermophilus JYST-26 to an MRS liquid culture medium for culture to obtain a bacterial liquid;
(3) Centrifugally collecting thalli by using bacterial liquid, washing by using sterile normal saline, and then suspending in 15wt% of recovered skim milk to obtain suspension;
(4) Adjusting the concentration of the suspension to 1.0-2.0 × 10 10 cfu/mL to obtain bacterial suspension, and freeze-drying the bacterial suspension to obtain the product for reducing oxalic acid and improving kidney stone.
Further, the raw materials of the MRS plate culture medium in the step (1) comprise: 10g of peptone, 5g of beef powder, 5g of sodium acetate trihydrate, 2g of dipotassium phosphate heptahydrate, 1mL of tween 80, 0.05g of manganese sulfate tetrahydrate, 2g of triammonium citrate, 20g of glucose, 0.2g of magnesium sulfate heptahydrate, 15g of agar and 1000mL of distilled water;
the preparation method comprises mixing the above raw materials, stirring at natural pH, sterilizing at 121 deg.C under 0.1MPa for 20min, pouring the sterilized culture medium into a plate, and cooling.
Further, the raw materials of the MRS liquid culture medium in the step (2) comprise: 10g of peptone, 5g of beef powder, 5g of sodium acetate trihydrate, 2g of dipotassium phosphate heptahydrate, 80 mL of tween, 0.05g of manganese sulfate tetrahydrate, 2g of triammonium citrate, 20g of glucose, 0.2g of magnesium sulfate heptahydrate and 1000mL of distilled water;
the preparation method comprises mixing the above materials, adjusting pH to 6.8, stirring, and sterilizing at 121 deg.C and 0.1MPa for 20 min.
Further, the culture conditions in the step (2) are 37 ℃ and 24h.
Furthermore, the product for reducing oxalic acid and improving kidney stone also comprises glucose.
Furthermore, the thallus quantity of streptococcus thermophilus JYST-26 in the product for reducing oxalic acid and improving kidney stone is 10 10 cfu/g。
The invention has the beneficial effects that:
the streptococcus thermophilus JYST-26 has strong intestinal tract colonization ability, can reduce the formation of calcium oxalate kidney stones and the dissolving ability of calcium oxalate kidney stones, and can avoid uncomfortable symptoms caused by medication and operation without any side effect.
Drawings
In order to more clearly illustrate the embodiments or technical solutions in the prior art of the present invention, the drawings used in the description of the embodiments or prior art will be briefly described below, and it is obvious for those skilled in the art that other drawings can be obtained based on these drawings without creative efforts.
FIG. 1 shows the results of the adhesion test in example 3.
FIG. 2 shows the results of the in vitro oxalic acid decomposition test in example 4.
FIG. 3 is a graph showing the effect of inhibiting the formation of calcium oxalate calculi in example 5.
FIG. 4 is a graph showing the effect of dissolving a fruit fly Marble's tube stone in example 6.
Detailed Description
In order to make those skilled in the art better understand the technical solution of the present invention, the technical solution in the embodiment of the present invention will be clearly and completely described below with reference to the drawings in the embodiment of the present invention, and it is obvious that the described embodiment is only a part of the embodiment of the present invention, and not all embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Glucose used in the embodiments of the present invention was purchased from Shangyiwang group, inc.
The raw materials of the MRS plate culture medium used in the specific embodiment of the invention comprise: 10g of peptone, 5g of beef powder, 5g of sodium acetate trihydrate, 2g of dipotassium phosphate heptahydrate, 1mL of tween 80, 0.05g of manganese sulfate tetrahydrate, 2g of triammonium citrate, 20g of glucose, 0.2g of magnesium sulfate heptahydrate, 15g of agar and 1000mL of distilled water;
the preparation method comprises mixing the above raw materials, stirring at natural pH, sterilizing at 121 deg.C and 0.1MPa for 20min, pouring the sterilized culture medium into a plate, and cooling.
The raw materials of the MRS liquid culture medium used in the specific embodiment of the invention comprise: 10g of peptone, 5g of beef powder, 5g of sodium acetate trihydrate, 2g of dipotassium phosphate heptahydrate, 80 mL of tween, 0.05g of manganese sulfate tetrahydrate, 2g of triammonium citrate, 20g of glucose, 0.2g of magnesium sulfate heptahydrate and 1000mL of distilled water;
the preparation method comprises mixing the above materials, adjusting pH to 6.8, stirring, and sterilizing at 121 deg.C and 0.1MPa for 20 min.
The raw materials of the MRS-sodium oxalate culture medium used in the specific embodiment of the invention comprise: 10g of peptone, 5g of beef powder, 5g of sodium acetate trihydrate, 2g of dipotassium phosphate heptahydrate, 80 mL of tween, 0.05g of manganese sulfate tetrahydrate, 2g of triammonium citrate, 20g of glucose, 0.2g of magnesium sulfate heptahydrate, 2.68g of sodium oxalate and 1000mL of distilled water;
the preparation method comprises mixing the above raw materials, stirring under natural pH, sterilizing at 121 deg.C and 0.1MPa for 20min, and making into MRS-oxalic acid culture medium with sodium oxalate content of 20 mmol/L.
The PBS buffer used in the embodiments of the present invention is prepared as follows: 0.27g of monopotassium phosphate, 1.42g of disodium hydrogen phosphate, 8g of sodium chloride and 0.2g of potassium chloride, adding about 800mL of deionized water, fully stirring and dissolving, adding concentrated hydrochloric acid to adjust the pH value to 7.4, adding deionized water to a constant volume of 1L, and sterilizing at 121 ℃ for 20min to obtain the compound.
The general standard food used in the embodiment of the method is prepared according to the following method: mixing 40g of sucrose, 25g of yeast, 67g of corn flour, 10g of soybean flour, 10g of agar powder, 1g of sodium benzoate, 40g of maltose and 1000mL of distilled water, carrying out water bath at 33 ℃ for 1h, boiling, adding 7mL of propionic acid when the temperature is reduced to 60-70 ℃, mixing uniformly and subpackaging.
The rat food used in the specific implementation mode of the method is prepared by mixing the following raw materials: 19g of flour, 23g of corn flour, 6g of sorghum flour, 10g of bran, 15g of soybean meal, 2g of vegetable oil, 1g of cod-liver oil, 10g of fish meal, 1g of bone meal, 1g of beer yeast, 1g of salt, 6.6g of starch, 3.4g of glycine, 0.5g of methionine and 0.5g of calcium carbonate.
Example 1
1. Screening of strains
1. A bacterium source:
naturally fermented cheese in 7 months of 2012 and in the families of herdsmen in inner Mongolia Dala.
2. Preparing a sample:
(1) Placing sterilized normal saline (0.85%) into a sterile triangular flask, adding 1g of bacteria source sample, and oscillating for later use;
(2) Diluting the solution obtained in the step (1) to prepare samples with different concentration gradients, wherein the concentration gradients are respectively 10 -1 、10 -2 、10 -3 、10 -4 、10 -5 、10 -6 、10 -7 The labels are 1#, 2#, 3#, 4#, 5#, 6#, and 7# respectively for standby;
3. culturing:
coating the solutions 1#, 2#, 3#, 4#, 5#, 6# and 7# in MRS plate culture medium with coater, and culturing at 37 deg.C under anaerobic condition for 48 hr;
4. and (3) selecting colonies:
the colony features 1-2mm diameter, round colony, regular edge, white middle bulge, and 5 single colonies.
5. Separation and purification:
inoculating the single colony to a screening culture medium by a scribing method, culturing at 37 ℃ under anaerobic condition for 48h, selecting the single colony, and preserving at-70 ℃ in a glycerol tube.
2. Identification of the species
And (3) sending the separated and purified single colony to be identified, wherein the identification unit is as follows: biometrics bioengineering (Shanghai) Ltd
1. In the identification process, the following primers were used:
27F：5'-AGAGTTTGATCCTGGCTCAG-3'；
1492R：5'-GGTTACCTTGTTACGACTT-3'。
2. the gene sequences identified were as follows:
CGCTCAGGACGAACGCTGGCGGCGTGCCTAATACATGCAAGTAGAACGCTGAAGAGAGGAGCTTGCTCTTCTTGGATGAGTTGCGAACGGGTGAGTAACGCGTAGGTAACCTGCCTTGTAGCGGGGGATAACTATTGGAAACGATAGCTAATACCGCATAACAATGGATGACACATGTCATTTATTTGAAAGGGGCAATTGCTCCACTACAAGATGGACCTGCGTTGTATTAGCTAGTAGGTGAGGTAATGGCTCACCTAGGCGACGATACATAGCCGACCTGAGAGGGTGATCGGCCACACTGGGACTGAGACACGGCCCAGACTCCTACGGGAGGCAGCAGTAGGGAATCTTCGGCAATGGGGGCAACCCTGACCGAGCAACGCCGCGTGAGTGAAGAAGGTTTTCGGATCGTAAAGCTCTGTTGTAAGTCAAGAACGGGTGTGAGAGTGGAAAGTTCACACTGTGACGGTAGCTTACCAGAAAGGGACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTCCCGAGCGTTGTCCGGATTTATTGGGCGTAAAGCGAGCGCAGGCGGTTTGATAAGTCTGAAGTTAAAGGCTGTGGCTCAACCATAGTTCGCTTTGGAAACTGTCAAACTTGAGTGCAGAAGGGGAGAGTGGAATTCCATGTGTAGCGGTGAAATGCGTAGATATATGGAGGAACACCGGTGGCGAAAGCGGCTCTCTGGTCTGTAACTGACGCTGAGGCTCGAAAGCGTGGGGAGCGAACAGGATTAGATACCCTGGTAGTCCACGCCGTAAACGATGAGTGCTAGGTGTTGGATCCTTTCCGGGATTCAGTGCCGCAGCTAACGCATTAAGCACTCCGCCTGGGGAGTACGACCGCAAGGTTGAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGAAGCAACGCGAAGAACCTTACCAGGTCTTGACATCCCGATGCTATTTCTAGAGATAGAAAGTTACTTCGGTACATCGGTGACAGGTGGTGCATGGTTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCCTATTGTTAGTTGCCATCATTCAGTTGGGCACTCTAGCGAGACTGCCGGTAATAAACCGGAGGAAGGTGGGGATGACGTCAAATCATCATGCCCCTTATGACCTGGGCTACACACGTGCTACAATGGTTGGTACAACGAGTTGCGAGTCGGTGACGGCGAGCTAATCTCTTAAAGCCAATCTCAGTTCGGATTGTAGGCTGCAACTCGCCTACATGAAGTCGGAATCGCTAGTAATCGCGGATCAGCACGCCGCGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCACGAGAGTTTGTAACACCCGAAGTCGGTGAGGTAACCTTTTGGAGCCAGCCGCCTAAGGTGGGACAGATGATTGGGGTGAAGTCGC。
3. the identified bacteria are streptococcus thermophilus(s) (B)Streptococcus thermophilus) The classification unit is as follows: (ii) Bacteria; firmicutes; bacillus; lactobacillus; streptococcus; streptococcus.
4. The strain is named as streptococcus thermophilus JYST-26 and is sent to the China general microbiological culture Collection center of the Committee for culture preservation, and the preservation information is as follows;
the streptococcus thermophilus JYST-26 is preserved in China general microbiological culture Collection center in 2019, 06 months and 27 days, and the address is as follows: west way No.1 hospital No.3, north beijing, chaoyang district, zip code: 100101, accession number: CGMCC No.18045.
Example 2
According to the bacterial number of the streptococcus thermophilus JYST-26 in the product as 10 10 cfu/g, and compounding the JYST-26 streptococcus thermophilus powder and the glucose.
The streptococcus thermophilus JYST-26 strain powder is prepared by the following preparation method:
(1) The streptococcus thermophilus JYST-26 is firstly activated on an MRS plate culture medium;
(2) Inoculating the activated streptococcus thermophilus JYST-26 to an MRS liquid culture medium according to the inoculation amount of 1% (v/v) for culturing for 24h at 37 ℃ to obtain a bacterial liquid;
(3) Centrifugally collecting thalli by using bacterial liquid, washing by using sterile normal saline, and then suspending in 15wt% of recovered skim milk to obtain suspension;
(4) Adjusting the concentration of the suspension to 1.0-2.0 × 10 10 cfu/mL to obtain bacterial suspension, and freeze-drying the bacterial suspension to obtain bacterial powder.
Example 3
The adhesion colonization effect of streptococcus thermophilus JYST-26 on human colorectal adenocarcinoma cell line HT-29 is detected, the human colorectal adenocarcinoma cell line HT-29 is purchased from the institute of basic medicine of Chinese academy of medical sciences, and meanwhile, an LGG strain (ATCC 53103) is used as a control. The detection method comprises the following steps:
1. culturing:
(1) The streptococcus thermophilus JYST-26 and LGG strains are activated in MRS liquid culture medium and cultured for 16h at 37 ℃.
(2) HT-29 cells were treated with DMEM medium (containing 10% fetal bovine serum) at 37 ℃ with 5% CO 2 The cells were cultured in an incubator at a concentration such that when the cells were completely differentiated, they were digested with 0.25% trypsin and then passaged. Cells grew adherent to the wall and the culture medium was changed daily. After passage for 3 times, the cell morphology is stable, and subsequent experiments can be carried out.
2. Adhesion test:
(1) Digesting the cultured HT-29 cells, and then suspending the cells in DMEM medium to adjust the cell concentration to 1 × 10 6 cfu/mL。
(2) Inoculating into 24-well cell culture plate containing sterile cell slide, inoculating 1mL per well, and culturing at 37 deg.C with 5% CO 2 Culturing in a concentration incubator until the cells form a fully differentiated monolayer.
(3) The medium was discarded and washed three times with sterile PBS.
(4) Inoculating activated Streptococcus thermophilus JYST-26 and LGG strain in MRS liquid culture medium, and culturingCulturing for 16h, centrifuging the fermentation liquid at 4000r/min to collect thallus, and adjusting the concentration of the bacteria liquid to 10 with PBS buffer solution 8 cfu/mL。
(5) Adding the prepared bacterial solution into 24-well plate, adding 1mL per well, and at 37 deg.C, and containing 5% CO 2 Incubating in an incubator with concentration for 1h.
(6) Unbound cells were precipitated, washed 5 times with PBS, and fixed with methanol for 10min.
(7) The slide was taken out for gram staining and counted under a microscope.
3. And (4) analyzing results:
as shown in FIG. 1, the adhesion amount is expressed by the total number of bacteria/100 cells in twenty random visual fields, and the adhesion rate of Streptococcus thermophilus JYST-26 is significantly higher compared with that of the LGG strain, which indicates that the Streptococcus thermophilus JYST-26 has stronger intestinal colonization ability.
Example 4
The in-vitro oxalic acid decomposition capacity of streptococcus thermophilus JYST-26 is detected by the following detection method:
1. culturing:
inoculating Streptococcus thermophilus JYST-26 in MRS liquid culture medium for activation culture for 16h, and inoculating 5% fermentation liquid in MRS-sodium oxalate culture medium for culture for 16h after culture. Meanwhile, a blank MRS-sodium oxalate culture medium which is not inoculated with streptococcus thermophilus JYST-26 is used as a control group.
2. Sample treatment:
and (3) respectively centrifuging the fermentation liquor and the blank MRS-sodium oxalate culture medium in the step (1) at 4 ℃ and 12000r/min, taking supernatant, and filtering the supernatant through a 0.2-micron filter membrane for later use.
3. And (3) detecting the concentration of oxalic acid:
the measurement is carried out by using an ion chromatograph, the ambient temperature is 25 ℃, the carrier gas is nitrogen, the pressure is 0.2MPa, the maximum pressure of a pump is set to be 21MPa, the flow rate is 1mL/min, and the sample injection volume is 100 mu L.
Using Dionex IonPac AS23 (4X 250 mm) anion protection column with high hydrophilicity and high column capacity, ionPac AG11 analytical column (4X 50 mm) anion protection column; the eluent was 16mmol/L sodium carbonate and 4mmol/L sodium bicarbonate.
Preparing 1000mg/L oxalic acid solution, filtering with 0.2 μm filter membrane, and diluting with ultrapure water to obtain standard solutions with concentrations of 16, 8, 4, 2, 1, 0.5, and 0.25 mg/L.
Filtering the sample with 0.2 μm filter membrane, diluting with ultrapure water 100 times, passing through On-Guard H column and C 18 And (5) pretreating the small column and then carrying out sample injection analysis.
4. And (4) analyzing results:
as shown in figure 2, the streptococcus thermophilus JYST-26 has strong oxalate degradation capability and certain potential in the aspects of reducing oxalate and improving kidney stones.
Example 5
The effect of streptococcus thermophilus JYST-26 on inhibiting the formation of calcium oxalate stones in Drosophila Marsderi is verified, and the test method comprises the following steps:
1. culturing female fruit flies:
w1118 Drosophila (purchased from Blomington Drosophila Stock Center) were fed to Drosophila tubes containing normal standard food, 20 flies/tube, 10 males and females each, at 25 ℃, 50% humidity, 12 hours day and night alternating climatic chamber for culture, and eclosion female adults were collected after 10 days.
2. Grouping tests:
(1) Control group: w1118 female drosophila melanogaster born on day 1 were fed by normal culture.
(2) Model group: w1118 female fruit flies born for 1 day were placed in chow containing 1% by mass sodium oxalate.
(3) Low dose group: w1118 female fruit flies born for 1 day were placed in chow containing 1% by weight sodium oxalate and the powder of Streptococcus thermophilus JYST-26 of example 2 was added to the chow to 10 7 cfu/g。
The medium dose group: w1118 female fruit flies born for 1 day were placed in a diet containing 1% by mass of sodium oxalate, and powder of JYST-26 of Streptococcus thermophilus of example 2 was added to the diet to 10% 8 cfu/g。
High dose group: w1118 female fruit flies born for 1 day were placed in a diet containing 1% by mass of sodium oxalate, and powder of JYST-26 of Streptococcus thermophilus of example 2 was added to the diet to 10% 9 cfu/g。
3. And (4) analyzing results:
dissect the fruit fly Marfan tube after one week, observe the calculus formation under the polarized light microscope, and count the calculus area. The results are shown in fig. 3, and it can be seen that the low dose group has a significant effect of inhibiting calculus formation (p < 0.05) compared with the model group, while the medium dose group and the high dose group have no calculus formation, indicating that the addition amount of streptococcus thermophilus JYST-26 in the medium dose group can achieve the effect of completely inhibiting calculus formation.
Example 6
The dissolving effect of streptococcus thermophilus JYST-26 on stones in Drosophila Marsderi is verified, and the test method comprises the following steps:
1. culturing female fruit flies:
w1118 Drosophila (purchased from Blomington Drosophila Stock Center) were fed to Drosophila tubes containing normal standard food at 20/tube, male and female each 10, incubated at 25 deg.C, 50% humidity, 12 hours day and night in alternating climatic chambers, and eclosion female adults were collected after 10 days.
2. Grouping tests:
(1) Control group: w1118 female drosophila melanogaster born on day 1 were fed by normal culture.
(2) Model group: w1118 female fruit flies born for 1 day were placed in chow containing 1% by mass sodium oxalate.
(3) Low dose group: placing W1118 female fruit flies born for 1 day into food containing 1% by mass of sodium oxalate, dissecting the fruit flies one week later, determining that the molding is successful, and transferring to food containing 1% by mass of sodium oxalate and 10% by mass of sodium oxalate 7 cfu/g of the food of Streptococcus thermophilus JYST-26 powder of example 2.
(4) The medium dose group: the same low dose group was operated, and the content of Streptococcus thermophilus JYST-26 in the food was changed to 10 8 cfu/g。
(5) High dose group: the same low dose group was operated, and the content of Streptococcus thermophilus JYST-26 in the food was changed to 10 9 cfu/g。
3. And (4) analyzing results:
dissecting the fruit fly Marfan tube after one week of culture, observing the calculus formation under a polarized light microscope, and counting the calculus area. As shown in FIG. 4, it can be seen that the addition of Streptococcus thermophilus JYST-26 results in a significant decrease in fruit fly Marek's tube stones (p < 0.05); as the addition amount of the streptococcus thermophilus JYST-26 is increased, stones in the Marek's tube are reduced, and the effect of dissolving stones of the streptococcus thermophilus JYST-26 strain is proved to be certain.
Example 7
The effect of streptococcus thermophilus JYST-26 on inhibiting the formation of rat calculus is verified, and the test method is as follows:
1. rat culture:
SPF grade male Sprague-Dawley rats (purchased from Guangdong provincial medical laboratory animal center) were selected at 39 and weighed at 180-220g. After 1 week of acclimation, the rats were randomly divided into 3 groups of 13 animals each and the body weight records were weighed.
2. Grouping tests:
(1) Control group: the mouse food is eaten, and the distilled water is used for drinking water.
(2) Model group: 5% L-Hydroxyproline (HLP) is added into rat food, and distilled water is used for drinking water.
(3) JYST-26 group: gavage 2X 10 daily on the basis of model group 8 cfu/kg of JYST-26 powder of Streptococcus thermophilus of example 2.
All rats were fed free diet and cultured at 25 ℃ for 12h day-night cycle.
3. And (3) rat index detection:
(1) Weight change%: after 28 days of the experiment, the rats were weighed and the weight change was calculated, weight change = (weight after experiment-weight before experiment)/weight before experiment x 100%.
(2) Urine volume mL: the rats were then placed in metabolic cages and urine was collected over 24h and urine volume was measured.
(3) Oxalic acid mg in urine: the method for detecting oxalic acid in example 4 was used to detect the oxalic acid content in urine samples. And calculating the amount of oxalic acid in 24h, wherein the amount of oxalic acid in 24h = the concentration of oxalic acid x the total amount of urine in 24h.
(4) Weight of kidney g: rats were anesthetized by intraperitoneal injection of 10% chloral hydrate (0.35 mL/100 g), the abdominal cavity was opened, the kidneys were washed in situ with 4 ℃ physiological saline through the abdominal aorta, the kidneys were harvested after cleaning, the water content was drained through sterile filter paper, and the rats were weighed.
(5) Degree of stone formation: <xnotran> , , , , , (+ + +), (+ +), (+), (±), (-), (-) , (+ + +), (+ +), (+), (±) . </xnotran>
4. And (4) analyzing results:
the effects of Streptococcus thermophilus JYST-26 on rat body weight, urine volume, kidney weight and urinary oxalic acid are shown in Table 1. As can be seen, for the four indexes of rat weight, urine volume, kidney weight and urinary oxalic acid, JYST-26 group has no significant difference (p > 0.05) compared with the control group, and the model group has significant difference (p < 0.01) compared with the control group. Shows that the streptococcus thermophilus JYST-26 has better inhibiting effect on the formation of the rat calculus.
TABLE 1 JYST-26 Effect on rat body weight, urine volume, kidney weight, urooxalic acid
The evaluation table of the degree of renal tissue calculi in rats is shown in Table 2. As can be seen, the lithogenesis rate of the JYST-26 group is obviously reduced compared with that of the model group (p is less than 0.01), which indicates that the JYST-26 of streptococcus thermophilus has obvious inhibition effect on the formation of the rat calculus.
TABLE 2 evaluation table of degree of renal tissue of rat
Although the present invention has been described in detail by referring to the drawings in connection with the preferred embodiments, the present invention is not limited thereto. Various equivalent modifications or substitutions can be made on the embodiments of the present invention by those skilled in the art without departing from the spirit and scope of the present invention, and these modifications or substitutions are within the scope of the present invention/any person skilled in the art can easily conceive of the changes or substitutions within the technical scope of the present invention.
Claims (8)
1. The Streptococcus thermophilus JYST-26 is characterized in that Streptococcus thermophilus JYST-26 is preserved in China general microbiological culture Collection center on 27.06.2019 at the position of No.3 of No.1 Homey of Xilu-Chen of the sunward area in Beijing, the preservation number is CGMCC NO.18045, and the 16S rRNA sequence of the Streptococcus thermophilus JYST-26 is shown in SEQ ID NO. 3.
2. Use of streptococcus thermophilus JYST-26 according to claim 1 in the preparation of a product for reducing oxalic acid and improving kidney stone.
3. An oxalic acid reducing and kidney stone improving product prepared from streptococcus thermophilus JYST-26 as claimed in claim 1, wherein the preparation method of the oxalic acid reducing and kidney stone improving product comprises the following steps:
(1) The streptococcus thermophilus JYST-26 is firstly activated on an MRS plate culture medium;
(2) Inoculating the activated streptococcus thermophilus JYST-26 to an MRS liquid culture medium for culture to obtain a bacterial liquid;
(3) Centrifugally collecting thalli by using bacterial liquid, washing by using sterile normal saline, and then suspending in 15wt% of recovered skim milk to obtain suspension;
(4) Adjusting the concentration of the suspension to 1.0-2.0 × 10 10 cfu/mL to obtain bacterial suspension, and freeze-drying the bacterial suspension to obtain the product for reducing oxalic acid and improving kidney stone.
4. The product for reducing oxalic acid and improving kidney stones according to claim 3, wherein the raw materials of the MRS plate culture medium of step (1) comprise: 10g of peptone, 5g of beef powder, 5g of sodium acetate trihydrate, 2g of dipotassium phosphate heptahydrate, 1mL of tween 80, 0.05g of manganese sulfate tetrahydrate, 2g of triammonium citrate, 20g of glucose, 0.2g of magnesium sulfate heptahydrate, 15g of agar and 1000mL of distilled water;
the preparation method comprises mixing the above raw materials, stirring at natural pH, sterilizing at 121 deg.C and 0.1MPa for 20min, pouring the sterilized culture medium into a plate, and cooling.
5. The product for reducing oxalic acid and improving kidney stone according to claim 3, wherein the raw materials of the MRS liquid culture medium in the step (2) comprise: 10g of peptone, 5g of beef powder, 5g of sodium acetate trihydrate, 2g of dipotassium phosphate heptahydrate, 80 mL of tween, 0.05g of manganese sulfate tetrahydrate, 2g of triammonium citrate, 20g of glucose, 0.2g of magnesium sulfate heptahydrate and 1000mL of distilled water;
the preparation method comprises mixing the above materials, adjusting pH to 6.8, stirring, and sterilizing at 121 deg.C and 0.1MPa for 20 min.
6. The product for reducing oxalic acid and improving kidney stone according to claim 3, wherein the culture condition in step (2) is 37 ℃ and 24 hours.
7. The product of claim 3, further comprising glucose.
8. The product for reducing oxalic acid and improving kidney stone of claim 3, wherein the count of the mycelia of Streptococcus thermophilus JYST-26 in the product for reducing oxalic acid and improving kidney stone is 10 10 cfu/g。
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| CN117100774A (en) * | 2023-10-25 | 2023-11-24 | 山东中科嘉亿生物工程有限公司 | Application of lactobacillus acidophilus JYLA-16 in preparation of products for treating gall-stone |
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| US20020061292A1 (en) * | 1999-05-28 | 2002-05-23 | Claudio De Simone | Dietary or pharmaceutical composition for use for the prevention or treatment of hyperoxaluria |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN117100774A (en) * | 2023-10-25 | 2023-11-24 | 山东中科嘉亿生物工程有限公司 | Application of lactobacillus acidophilus JYLA-16 in preparation of products for treating gall-stone |
| CN117100774B (en) * | 2023-10-25 | 2024-02-13 | 山东中科嘉亿生物工程有限公司 | Application of lactobacillus acidophilus JYLA-16 in preparation of products for treating gall-stone |
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