A supramolecular tightening dropsordry containing pelvic floor muscle repair composition and a method for preparing the same

Abstract

The present application relates to the technical field of health care products, in particular to a pelvic floor muscle repairing composition containing supermolecular tightening Dropsordry and a preparation method thereof, which comprises the following components in mass fraction: 10-14 parts of protein composition, 20-24 parts of chitosan modified Dropsordry, 5-7 parts of freeze-dried powder of Lactobacillus bulgaricus, 10-12 parts of hyaluronic acid, 6-8 parts of glycine and 15-17 parts of proline; each part of the protein composition comprises 20-30 parts of myosin, 30-34 parts of actin, 5-10 parts of hydrolyzed collagen and 240-280 parts of deionized water. The pelvic floor muscle repairing composition can reduce drug toxicity, reduce the amount of drug administration, continuously supplement the required elements of the body during treatment, and quickly relieve the constipation problem caused by the functional damage of the pelvic floor muscle.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of health products, and particularly relates to a pelvic floor muscle repair composition containing supermolecular tightening Dropsordry and a preparation method thereof. BACKGROUND

[0002] Pelvic floor dysfunction refers to a series of non-organic diseases caused by weakness, defects and dysfunction of the pelvic floor support structure. Pelvic floor dysfunction is closely related to age, obesity, pregnancy and childbirth, chronic diseases and other factors, and seriously affects the quality of life of patients. According to epidemiological surveys, about 45% of married and childbearing women have different degrees of pelvic floor dysfunction, among which urinary incontinence and pelvic organ prolapse are the most common clinical manifestations. The causes of pelvic floor dysfunction are diverse, including but not limited to pregnancy and childbirth, changes in hormone levels, aging, obesity, chronic cough, constipation, etc. Pregnancy and childbirth are independent risk factors for pelvic floor dysfunction. Chronic stretching of the uterus during pregnancy and compression of the pelvic floor during childbirth can cause damage to the pelvic floor muscles and connective tissue. In addition, the decrease in estrogen levels also affects the collagen metabolism of the pelvic floor connective tissue, leading to weakening of the pelvic floor support structure. The main clinical manifestations of pelvic floor dysfunction include urinary incontinence, pelvic organ prolapse, sexual dysfunction, chronic pelvic pain, fecal incontinence and constipation, etc. These symptoms not only affect the daily life of patients, but also can lead to psychological problems such as social phobia and depression.

[0003] The diagnosis of pelvic floor dysfunction mainly relies on medical history inquiry, routine examination and pelvic floor muscle function assessment. In terms of treatment, pelvic floor rehabilitation is the first choice of first-line measures, including non-surgical treatment methods such as pelvic floor muscle exercise, biofeedback and electrical stimulation. For patients with severe pelvic floor dysfunction, surgical treatment may be needed. Drug therapy plays an auxiliary role in the treatment of pelvic floor dysfunction, mainly aiming at the relief of symptoms and the improvement of quality of life.

[0004] With the deepening of the understanding of pelvic floor dysfunction, patients' demand for improving the quality of life is increasing, and the demand for drug treatment as part of the comprehensive treatment of pelvic floor muscle dysfunction is also increasing. Drug treatment plays an important role in relieving symptoms and improving the quality of life, especially when non-surgical treatment is ineffective or the patient is not suitable for surgical treatment. Although drug treatment shows potential in the treatment of pelvic floor dysfunction, its side effects and safety are still the focus of research, and some drugs may cause local or systemic side effects, such as hormone replacement therapy, which may increase the risk of breast cancer and endometrial cancer. Therefore, researchers are looking for safer drug alternatives, as well as developing drug delivery routes and dosage regimens that can minimize side effects, while the development of new drugs is mainly focused on selective estrogen receptor modulators and drugs targeting specific pathophysiological mechanisms, which provide new ideas and more treatment options for drug treatment of pelvic floor dysfunction.

[0005] For example, CN111671911B discloses a drug composition for treating pelvic floor dysfunction and its preparation method, which is composed of buspirone, polyethylene glycol 4000, microcrystalline cellulose, 6% PVP ethanol solution and micro-powder silica gel. The composition has significant synergistic effect by specific drug matching, significantly reduces the toxic and side effects of the two drugs on the human body, reduces the pressure of anal sphincter muscle, especially the anal sphincter muscle, reduces the anal rectal maximum voluntary contraction pressure and anal rectal resting pressure, improves the efficacy of the drug composition, and has broad medical application prospect.

[0006] For example, CN112156173A discloses a gel for treating pelvic floor muscle dysfunction and its preparation method, which is prepared from zedoary extract, soy isoflavones, frankincense acid, fibronectin, hydrolyzed collagen and excipients. The composition can promote the contraction of pelvic floor muscle tissue, repair muscle nerves and improve muscle tension, achieving the treatment of pelvic floor dysfunction and significantly improving stress urinary incontinence caused by pelvic floor injury.

[0007] However, the above-mentioned disclosed patents do not mention and solve the problem of how to reduce drug toxicity, without affecting the re-pregnancy of pregnant women and without damaging the physiological function of women.

[0008] Therefore, it is urgent to develop a pelvic floor muscle repair composition that can reduce drug toxicity, reduce drug dosage, continuously supplement the body's needs during treatment, and quickly relieve constipation caused by pelvic floor muscle dysfunction. SUMMARY

[0009] The main purpose of the present application is to provide a supramolecular tightening Dropsordry-containing pelvic floor muscle repair composition and a preparation method thereof, which has a sustained-release effect, can reduce drug toxicity and reduce the amount of drug administration, can continuously supplement the body's required elements during treatment, and can quickly relieve constipation problems caused by pelvic floor muscle function damage.

[0010] To achieve the above-mentioned purpose, the technical scheme adopted by the present application is as follows:

[0011] In one aspect, the present application provides a supramolecular tightening Dropsordry-containing pelvic floor muscle repair composition, which comprises the following components by mass fraction: 10-14 parts of a protein composition, 20-24 parts of a chitosan modified Dropsordry, 5-7 parts of a freeze-dried powder of Lactobacillus bulgaricus, 10-12 parts of hyaluronic acid, 6-8 parts of glycine, 15-17 parts of proline, and 240-280 parts of deionized water; each part of the protein composition comprises 20-30 parts of myosin, 30-34 parts of actin, and 5-10 parts of hydrolyzed collagen.

[0012] Pelvic floor muscle repair is one of the important medical measures to maintain women's health. At present, there are various drugs for pelvic floor muscle repair. Although drug therapy shows potential in the treatment of female pelvic floor dysfunction, its side effects and safety are still the focus of research. Some drugs may cause local or systemic side effects, such as hormone replacement therapy, which may increase the risk of breast cancer and endometrial cancer. Therefore, researchers are looking for safer drug alternatives, as well as developing drug administration routes and dosage regimens that can minimize side effects.

[0013] The pelvic floor muscle repair composition of the present application is prepared by using a protein composition, a chitosan modified Dropsordry, a freeze-dried powder of Lactobacillus bulgaricus, hyaluronic acid, glycine, and proline as main components. Through the synergistic effect of various components, it can not only repair the pelvic floor muscle, but also have a sustained-release effect to reduce the amount of drug administration and reduce biological toxicity. At the same time, it can continuously supplement the body's required elements during treatment, and can quickly relieve constipation problems caused by pelvic floor muscle function damage.

[0014] In some embodiments, the preparation method of the freeze-dried powder of Lactobacillus bulgaricus comprises the following steps:

[0015] A1, inoculate the Lactobacillus bulgaricus activation culture into the MRS culture medium at an inoculation amount of 1-2% of the volume of the MRS culture medium, activate the culture at 30-37℃ for 18-24h, and obtain a seed bacterial solution;

[0016] A2, inoculate the seed bacteria liquid in step A1 into a fermentation tank containing sterilized fermentation medium, control the pH to be 4-6, ferment at 35-40℃ for 30-60h, add fermentation accelerant at 10-15h after the start of fermentation to continue fermentation, cool down to 25-30℃ after the end of fermentation, then centrifuge to obtain bacterial slurry;

[0017] A3, mix the bacterial slurry in step A3 with a protective agent, control the pH of the mixture to be 4.5-6.5, freeze-dry the mixture to obtain Lactobacillus bulgaricus freeze-dried powder.

[0018] The Lactobacillus bulgaricus freeze-dried powder of the present application is prepared by a specific preparation method and conditions, and can obtain Lactobacillus bulgaricus freeze-dried powder with higher viable count. The conventional preparation method is difficult to obtain Lactobacillus bulgaricus freeze-dried powder with the same viable count as the present application. The reason may be that the specific fermentation accelerant used in the present application not only can improve the fermentation rate of the bacterial liquid, but also can play a surface active role to reduce the amount of foam generated during fermentation, thereby improving the viable count of Lactobacillus bulgaricus freeze-dried powder.

[0019] In some embodiments, the Lactobacillus bulgaricus is preserved in China Center for Type Culture Collection, the preservation date is December 11, 2022, and the biological preservation number is CCTCC S2022087.

[0020] The specific biological preservation number of Lactobacillus bulgaricus used in the present application is more suitable for the pelvic floor muscle repair composition of the present application, so that the composition can promote intestinal digestion and relieve pelvic floor muscle pressure during repair, thereby improving the pelvic floor muscle repair efficiency.

[0021] In some embodiments, the fermentation medium comprises the following raw materials, based on 100% of the total mass:

[0022] Casein peptone 1-3%, corn starch 1-2%, glucose 2-4%, soybean protein powder 0.5-1%, yeast extract powder 1-3%, beef extract powder 0.5-1%, potassium dihydrogen phosphate 0.05-0.1%, diammonium hydrogen citrate 0.05-0.1%, magnesium sulfate 0.01-0.1%, vitamin E 0.5-0.8%, defoaming agent 0.1-0.5%, and the balance is water; the pH of the fermentation medium is 5-6.

[0023] In some embodiments, the fermentation accelerant comprises the following components by mass: 0.5-0.9 parts of calcium myristate, 3-5 parts of sodium 2-hydroxyvalerate, 0.05-0.09 parts of magnesium chloride, and 20-30 parts of Span-80.

[0024] The present application uses specific components in a certain ratio as a fermentation accelerant, which can improve the fermentation efficiency of the bacterial liquid and reduce the fermentation time.

[0025] In some embodiments, the preparation method of the chitosan modified Dropsordry includes the following steps:

[0026] B1, dichlorosulfoxide is cooled to -5-0℃, humic acid is added thereto and the temperature is controlled, after the addition is completed, the temperature is increased to room temperature, constant temperature stirring is carried out for 2-4h, after the reaction is completed, it is quenched, dried by reduced pressure concentration to obtain acyl chloride humic acid;

[0027] B2, the acyl chloride humic acid in step B1, chitosan and acetone are mixed, sodium bicarbonate is added, the temperature is increased to 50-60℃, constant temperature stirring is carried out for 10-12h, after the reaction is completed, filtration is added, acetone is washed, and the humic acid grafted modified chitosan is obtained;

[0028] B3, under the atmosphere of inert protective gas, 4, 4'-dibromobenzophenone is mixed with chloroform, zinc is added thereto, stirring is carried out at room temperature for 1-3h, after the reaction is completed, filtration is carried out, the filtrate is dried by reduced pressure concentration to obtain compound A;

[0029] B4, compound A in step B3 and silver nitrate are mixed, dichloromethane is added, stirring is carried out at room temperature for 26-30h in the dark, after the reaction is completed, filtration is carried out, and drying is carried out to obtain compound B;

[0030] B5, compound B in step B4, humic acid grafted modified chitosan in step B2 and Dropsordry are mixed, an ethanol and dichloromethane mixed solution is added, stirring is carried out at room temperature for 18-20h, after the reaction is completed, n-heptane is added until no precipitate is generated, after filtration, -10--5℃ ethanol is used for washing, and drying is carried out to obtain chitosan modified Dropsordry.

[0031] Dropsordry in Spain is a mature drug, in which the main components are soybean isoflavones and pumpkin seed extract, which has antimuscarinic receptor effect, reduces the expression of bladder wall receptors M2 and M3, reduces the spontaneous contraction of the detrusor muscle, promotes collagen synthesis, promotes the growth and repair of muscle tissue, improves the atrophy of the bladder and urethra, balances androgens, binds androgen receptors-synthesis metabolism, and strengthens the function of the pelvic floor muscles.

[0032] It is known that genistein in soybean isoflavones belongs to low toxicity substances, and it can cause false pregnancy, stillbirth, abortion and infertility in animals in experiments, and the development trend of the pharmaceutical field is to research low toxicity drugs, the application adopts Dropsordry, compound B and chitosan for combination modification, can form a stable supramolecular gel to release effective components, reduce the intake of toxic substances per unit time, reduce the risk of disease and reduce toxicity, in addition, a large amount of Zn element is contained, which can improve the body immunity of pregnant women, and further improve the muscle contraction ability near the uterus, reduce the dependence on the pelvic floor muscle, thereby improving the repair efficiency of the composition on the pelvic floor muscle.

[0033] In some embodiments, in step B2, the mass ratio of the acyl chloride humic acid and chitosan is (1-3):1.

[0034] Preferably, in step B2, the mass ratio of the acyl chloride humic acid and chitosan is 2:1.

[0035] In some embodiments, in step B3, the molar ratio of the 4,4'-dibromobenzophenone to zinc is 1:(1.2-1.5).

[0036] Preferably, in step B3, the molar ratio of the 4,4'-dibromobenzophenone to zinc is 1:1.3.

[0037] In some embodiments, in step B5, the mass ratio of the compound B, humic acid grafted chitosan and Dropsordry is (0.2-0.5):1:(3-5).

[0038] Preferably, in step B5, the mass ratio of the compound B, humic acid grafted chitosan and Dropsordry is 0.3:1:4.

[0039] Another aspect of the application provides a preparation method of a pelvic floor muscle repair composition containing supramolecular tightening Dropsordry, comprising the following steps:

[0040] S1, uniformly mixing a protein composition, hyaluronic acid, glycine and proline to obtain material A;

[0041] S2, mixing chitosan modified Dropsordry and deionized water to form a supramolecular gel;

[0042] S3, under continuous stirring, adding the material A in step S1 and freeze-dried powder of Lactobacillus bulgaricus to the supramolecular gel in step S2 to obtain a pelvic floor muscle repair composition containing supramolecular tightening Dropsordry.

[0043] Compared with the prior art, the application has the following beneficial effects:

[0044] (1) The present application is prepared by adopting protein composition, chitosan modified Dropsordry, Lactobacillus bulgaricus freeze-dried powder, hyaluronic acid, glycine and proline as main components, through the synergistic effect of various components, it can not only play a sustained-release role to reduce the amount of drug application and reduce biological toxicity on the basis of repairing the pelvic floor muscle, but also can continuously supplement the body's required elements during treatment, and quickly relieve the constipation problem caused by pelvic floor muscle function damage.

[0045] (2) The preparation of the Lactobacillus bulgaricus freeze-dried powder of the present application adopts specific fermentation accelerators, which can not only improve the fermentation rate of the bacterial solution, but also play a surface activity role to reduce the amount of foam generated during fermentation, thereby improving the number of viable bacteria of the Lactobacillus bulgaricus freeze-dried powder; in addition, the Lactobacillus bulgaricus with a specific biological preservation number is more suitable for the pelvic floor muscle repair composition of the present application, so that the composition can promote intestinal digestion and relieve pelvic floor muscle pressure during repair, further improving the pelvic floor muscle repair efficiency.

[0046] (3) The chitosan modified Dropsordry of the present application adopts Dropsordry, compound B and chitosan for combination modification, which can form a stable supramolecular gel to release effective components, reduce the intake of toxic substances per unit time, reduce the risk of disease and reduce toxicity; in addition, it contains a large amount of Zn element, which can improve the body immunity of pregnant women, and further improve the muscle contraction ability near the uterus, reduce the dependence on the pelvic floor muscle, thereby improving the repair efficiency of the composition on the pelvic floor muscle. DETAILED DESCRIPTION

[0047] The various exemplary embodiments of the present application will now be described in detail, which should not be considered as limiting the present application, but should be understood as a more detailed description of certain aspects, characteristics and embodiments of the present application.

[0048] It should be understood that the terms described in the present application are only for describing the specific embodiments, and are not used to limit the present application. In addition, for the numerical range in the present application, it should be understood that each intermediate value between the upper limit and the lower limit of the range is also specifically disclosed. Each smaller range between any stated value or intermediate value within the stated range and any other stated value or intermediate value within the stated range is also included in the present application. The upper limit and the lower limit of these smaller ranges can be independently included or excluded from the range.

[0049] All technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application pertains unless otherwise specifically defined herein. Although preferred methods and materials are described herein, any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present application. Numerous modifications and variations of the present application are possible in light of the above teachings and, therefore, it is to be understood that within the scope of the present application, the application can be practiced otherwise than specifically described. Other implementations of this application will be apparent to those skilled in the art from consideration of the specification and practice of the application disclosed herein. The examples and embodiments described herein are exemplary only and are not intended to be limiting.

[0050] It should be noted that the "centrifugation", "filtration", "stirring" and other operations described in the present application are routine operations for those skilled in the art, which can be selected by the actual operation.

[0051] MRS medium was purchased from Shenzhen Dotpoint Biotechnology Co., Ltd.; the activated culture of Lactobacillus bulgaricus with biological preservation numbers CCTCC S2022087 and CCTCC CB 20082294 can be obtained through commercial channels; commercially available fermentation accelerators were purchased from Hebei Runbu Biotechnology Co., Ltd.; Dropsordry was purchased from Macklin Co., Ltd.

[0052] Preparation Example 1

[0053] The method for preparing the Lactobacillus bulgaricus freeze-dried powder comprises the following steps:

[0054] A1, inoculate the activated culture of Lactobacillus bulgaricus with biological preservation number CCTCC S2022087 into MRS medium at an inoculation amount of 1% of the volume of the MRS medium, activate and culture at 34℃ for 21h to obtain a seed bacterial solution;

[0055] A2, inoculate the seed bacterial solution in step A1 into a fermentation tank containing sterilized fermentation medium, control the pH to be 5, and ferment at 37℃ for 50h; at 12h after the start of fermentation, add a fermentation accelerator at a volume of 0.3% of the fermentation medium to continue fermentation, and after the end of fermentation, cool to 27℃, then centrifuge to obtain bacterial slurry;

[0056] A3, mix the bacterial slurry in step A3 with a protective agent to make the pH of the mixture be 5, freeze-dry the mixture to obtain Lactobacillus bulgaricus freeze-dried powder with a bacterial count of 1.5×10 11 cfu / g.

[0057] The protective agent comprises the following components in parts by mass: 130 parts of trehalose, 120 parts of sorbitol, 30 parts of milk protein, 1 part of K2HPO4·3H2O, 0.5 parts of Na2HPO4·12H2O, 0.7 parts of histidine, 0.8 parts of arginine, and 0.8 parts of lysine.

[0058] The fermentation accelerator comprises the following components in parts by mass: 0.7 parts of calcium myristate, 4 parts of sodium 2-hydroxyvalerate, 0.05-0.09 parts of magnesium chloride, and 20-30 parts of Span-80.

[0059] The fermentation medium comprises the following raw materials in total 100% by mass:

[0060] Casein peptone 2%, corn starch 1.5%, glucose 3%, soybean protein powder 0.7%, yeast extract powder 2%, beef extract powder 0.7%, potassium dihydrogen phosphate 0.08%, diammonium hydrogen citrate 0.07%, magnesium sulfate 0.07%, vitamin E 0.7%, Tween-80 0.3%, and the balance is water; the pH of the fermentation medium is 5.

[0061] Preparation Example 2

[0062] The preparation method of the freeze-dried powder of Lactobacillus bulgaricus is the same as that in Preparation Example 1, except that the preservation number of the Lactobacillus bulgaricus is CCTCC CB 20082294.

[0063] Preparation Example 3

[0064] The preparation method of the freeze-dried powder of Lactobacillus bulgaricus is the same as that in Preparation Example 1, except that a commercially available fermentation accelerator is used instead of the fermentation accelerator in Preparation Example 1.

[0065] Preparation Example 4

[0066] The preparation method of the chitosan modified Dropsordry comprises the following steps:

[0067] B1, 300 mL of dichlorosulfide is cooled to -3 ℃, 5 g of humic acid is added and the temperature is controlled, after the addition is completed, the temperature is raised to room temperature, constant temperature stirring is carried out for 3 h, after the reaction is completed, it is quenched, and dried under reduced pressure to obtain acylated humic acid;

[0068] B2, 5 g of the acylated humic acid in step B1, 2.5 g of chitosan and 300 mL of acetone are mixed, 1 g of sodium bicarbonate is added, the temperature is raised to 55 ℃, constant temperature stirring is carried out for 11 h, after the reaction is completed, it is filtered, and washed with acetone to obtain humic acid grafted modified chitosan;

[0069] B3, 10 mmol of 4,4'-dibromobenzophenone was mixed with 50 mL of chloroform under N2 atmosphere, 13 mmol of zinc was added, stirred at room temperature for 2 h, after the reaction was completed, the filtrate was concentrated under reduced pressure and dried to obtain compound A;

[0070] B4, 10 mmol of compound A in step B3 was mixed with 100 mol of silver nitrate, 120 mL of dichloromethane was added, stirred at room temperature for 28 h in the dark, after the reaction was completed, the filtrate was dried to obtain compound B;

[0071] B5, 1.2 g of compound B in step B4, 4 g of humic acid grafted modified chitosan in step B2 and 5.6 g of Dropsordry were mixed, 100 mL of a mixed solution of ethanol and dichloromethane was added, stirred at room temperature for 19 h, after the reaction was completed, n-heptane was added until no precipitate was produced, after filtration, washed with -3℃ ethanol, and dried to obtain chitosan modified Dropsordry.

[0072] Preparation Example 5

[0073] The preparation method of chitosan modified Dropsordry was the same as in Preparation Example 5, except that the acylchlorinated humic acid was 1.8 g.

[0074] Preparation Example 6

[0075] The preparation method of chitosan modified Dropsordry was the same as in Preparation Example 5, except that the acylchlorinated humic acid was 8.2 g.

[0076] Preparation Example 7

[0077] The preparation method of chitosan modified Dropsordry was the same as in Preparation Example 5, except that the zinc was 6 mmol.

[0078] Preparation Example 8

[0079] The preparation method of chitosan modified Dropsordry was the same as in Preparation Example 5, except that the compound B was 2.5 g.

[0080] Example 1

[0081] A pelvic floor muscle repair composition containing supermolecular tightening Dropsordry, comprising the following components in mass parts: 12 parts of a protein composition, 22 parts of chitosan modified Dropsordry, 6 parts of Lactobacillus bulgaricus freeze-dried powder, 11 parts of hyaluronic acid, 7 parts of glycine, 16 parts of proline, and 260 parts of deionized water; each part of the protein composition comprises 25 parts of myosin, 32 parts of actin, and 7 parts of hydrolyzed collagen.

[0082] The chitosan modified Dropsordry is prepared according to Preparation Example 4; and the Lactobacillus bulgaricus freeze-dried powder is prepared according to Preparation Example 1.

[0083] The preparation method of the pelvic floor muscle repair composition containing supermolecular tightening Dropsordry comprises the following steps:

[0084] S1, uniformly mixing a protein composition, hyaluronic acid, glycine, and proline to obtain material A;

[0085] S2, mixing chitosan modified Dropsordry and deionized water to form a supermolecular gel;

[0086] S3, under continuous stirring, adding material A and Lactobacillus bulgaricus freeze-dried powder in step S1 to the supermolecular gel in step S2 to obtain the pelvic floor muscle repair composition containing supermolecular tightening Dropsordry.

[0087] Example 2

[0088] A pelvic floor muscle repair composition containing supermolecular tightening Dropsordry, comprising the following components in mass parts: 10 parts of a protein composition, 20 parts of chitosan modified Dropsordry, 5 parts of Lactobacillus bulgaricus freeze-dried powder, 10 parts of hyaluronic acid, 6 parts of glycine, 15 parts of proline, and 240 parts of deionized water; each part of the protein composition comprises 20 parts of myosin, 30 parts of actin, and 5 parts of hydrolyzed collagen.

[0089] The chitosan modified Dropsordry is prepared according to Preparation Example 4; and the Lactobacillus bulgaricus freeze-dried powder is prepared according to Preparation Example 1.

[0090] The preparation method of the pelvic floor muscle repair composition containing supermolecular tightening Dropsordry comprises the following steps:

[0091] S1, uniformly mixing a protein composition, hyaluronic acid, glycine, and proline to obtain material A;

[0092] S2, mixing chitosan modified Dropsordry and deionized water to form a supermolecular gel;

[0093] S3, under continuous stirring, the supramolecular gel in step S2 is added with the material A in step S1 and the freeze-dried powder of Lactobacillus bulgaricus to obtain the pelvic floor muscle repair composition containing supramolecular tightening Dropsordry.

[0094] Example 3

[0095] A pelvic floor muscle repair composition containing supramolecular tightening Dropsordry, comprising the following components by mass fraction: 14 parts of a protein composition, 220 parts of chitosan modified Dropsordry, 7 parts of freeze-dried powder of Lactobacillus bulgaricus, 12 parts of hyaluronic acid, 8 parts of glycine, 17 parts of proline and 280 parts of deionized water; each part of the protein composition comprises 30 parts of myosin, 34 parts of actin and 10 parts of hydrolyzed collagen.

[0096] The chitosan modified Dropsordry is prepared according to Preparation Example 4; and the freeze-dried powder of Lactobacillus bulgaricus is prepared according to Preparation Example 1.

[0097] A preparation method of a pelvic floor muscle repair composition containing supramolecular tightening Dropsordry, comprising the following steps:

[0098] S1, mixing a protein composition, hyaluronic acid, glycine and proline uniformly to obtain a material A;

[0099] S2, mixing chitosan modified Dropsordry with deionized water to form a supramolecular gel;

[0100] S3, under continuous stirring, the supramolecular gel in step S2 is added with the material A in step S1 and the freeze-dried powder of Lactobacillus bulgaricus to obtain the pelvic floor muscle repair composition containing supramolecular tightening Dropsordry.

[0101] Example 4

[0102] A pelvic floor muscle repair composition containing supramolecular tightening Dropsordry and a preparation method thereof, the specific implementation manner is the same as that in Embodiment 1, except that the freeze-dried powder of Lactobacillus bulgaricus is prepared according to Preparation Example 2.

[0103] Example 5

[0104] A pelvic floor muscle repair composition containing supramolecular tightening Dropsordry and a preparation method thereof, the specific implementation manner is the same as that in Embodiment 1, except that the freeze-dried powder of Lactobacillus bulgaricus is prepared according to Preparation Example 3.

[0105] Example 6

[0106] A pelvic floor repair composition containing supramolecular tightening Dropsordry and a method of preparing the same, the detailed description of which is the same as that of Example 1, except that the chitosan-modified Dropsordry is prepared according to Preparation Example 5.

[0107] Example 7

[0108] A pelvic floor repair composition containing supramolecular tightening Dropsordry and a method of preparing the same, the detailed description of which is the same as that of Example 1, except that the chitosan-modified Dropsordry is prepared according to Preparation Example 6.

[0109] Example 8

[0110] A pelvic floor repair composition containing supramolecular tightening Dropsordry and a method of preparing the same, the detailed description of which is the same as that of Example 1, except that the chitosan-modified Dropsordry is prepared according to Preparation Example 7.

[0111] Example 9

[0112] A pelvic floor repair composition containing supramolecular tightening Dropsordry and a method of preparing the same, the detailed description of which is the same as that of Example 1, except that the chitosan-modified Dropsordry is prepared according to Preparation Example 8.

[0113] Comparative Example 1

[0114] A pelvic floor repair composition containing supramolecular tightening Dropsordry and a method of preparing the same, the detailed description of which is the same as that of Example 1, except that Dropsordry is used instead of chitosan-modified Dropsordry.

[0115] Comparative Example 2

[0116] A pelvic floor repair composition containing supramolecular tightening Dropsordry and a method of preparing the same, the detailed description of which is the same as that of Example 1, except that the Lactobacillus bulgaricus freeze-dried powder is not added.

[0117] Performance test:

[0118] (1) Sustained-release performance: 10 mL of the pelvic floor repair composition was added to 8 mL of a buffer solution with pH = 6.86, 3 mL of the supernatant was taken every 1 h and 3 mL of the buffer solution with pH = 6.86 was added, and the UV absorbance of hyaluronic acid was measured to calculate the sustained-release rate according to the following formula:

[0119] Sustained-release rate (%) = 3∑C i-1 + 5C i / m x 100

[0120] wherein C i-1 is the concentration of the measured hyaluronic acid in the slow-release liquid at the (i-1)th time, C i is the concentration of the measured hyaluronic acid in the slow-release liquid at the ith time, and m is the total mass of the hyaluronic acid in the composition.

[0121] (2) Effect on intestinal function: Referring to the description of CN111671911B "A pharmaceutical composition for treating pelvic floor dyskinesia and constipation and a preparation method thereof", paragraph

[0077] , the average time for the first black stool to be discharged was calculated for each group of 10 mice, and the longer the time, the worse the intestinal function and the greater the degree of constipation.

[0122] The pelvic floor muscle repair compositions of each example and comparative example were tested according to the above methods (1) to (2), and the test results are shown in Table 1.

[0123] Table 1

[0124]

[0125]

[0126] According to the data in Table 1, the compositions of Examples 1-3 have good sustained-release performance and certain therapeutic effect on constipation. Example 4 has a decreased effect on the intestinal tract due to the use of Lactobacillus bulgaricus with different preservation numbers, which is not conducive to reducing the load of the pelvic floor muscles. Example 5 has a decreased effect on the intestinal tract due to the use of commercially available fermentation accelerators instead of the fermentation accelerators prepared in Preparation Example 1, which leads to a decrease in the number of viable bacteria in the freeze-dried powder and is not conducive to reducing the load of the pelvic floor muscles. Example 6 has a decreased effect on the intestinal tract due to the change in the mass ratio of acylated humic acid and chitosan, which leads to an excessively fast drug release rate, and the soy isoflavones may cause certain toxic effects on the body and are not conducive to reducing drug intake to achieve long-term therapeutic effect. Example 7 has an excessively slow drug release rate due to the change in the mass ratio of acylated humic acid and chitosan, which affects the time of black stool discharge in mice. Example 8 has an excessively fast drug release rate due to the change in the molar ratio of 4,4'-dibromodiphenylmethanone and zinc, which leads to a decrease in the crosslinking density of the formed gel, and the soy isoflavones may cause certain toxic effects on the body and are not conducive to reducing drug intake to achieve long-term therapeutic effect. Example 9 has an excessively slow drug release rate due to the change in the mass ratio of Compound B, humic acid grafted chitosan and Dropsordry, which leads to an excessively large crosslinking density of the formed gel and affects the time of black stool discharge in mice, and the short-term therapeutic effect is poor. Comparative Example 1 has poor sustained-release performance due to the use of Dropsordry instead of chitosan modified Dropsordry, and the soy isoflavones may cause certain toxic effects on the body and are not conducive to reducing drug intake to achieve long-term therapeutic effect. Comparative Example 2 has a weakened effect on the intestinal tract due to the absence of Lactobacillus bulgaricus freeze-dried powder, which is not conducive to reducing the load of the pelvic floor muscles.

[0127] (3) Small-scale experiment for treatment of pelvic floor muscle damage: three female volunteers aged 40-50 years old and suffering from urinary incontinence caused by pelvic floor muscle problems were selected, and 550 mL of the composition prepared in Example 1 was taken on an empty stomach every morning, once every other day for 3 weeks. During the period, the leakage of urine was tested by jumping rope every week, and the original diet condition was not changed during the taking period.

[0128] The composition prepared in Example 1 was tested according to method (3), and the test results are shown in Table 2.

[0129] Table 2

[0130]

[0131]

[0132] According to the data in Table 2, the composition prepared in Example 1 has a certain effect on the repair of the pelvic floor muscles.

[0133] Finally, it should be noted that the above detailed description is merely illustrative of the technical solutions of the present application and is not limiting, and although the present application has been described in detail with reference to the examples, it should be understood by those skilled in the art that the technical solutions of the present application can be modified or equivalently replaced without departing from the spirit and scope of the technical solutions of the present application, and all should be covered in the scope of the claims of the present application.

Claims

1. A pelvic floor muscle repair composition containing supramolecular firming Dropsordry, characterized in that, The product comprises the following components by weight: 10-14 parts protein composition, 20-24 parts chitosan-modified Dropsordry, 5-7 parts Lactobacillus bulgaricus lyophilized powder, 10-12 parts hyaluronic acid, 6-8 parts glycine, 15-17 parts proline, and 240-280 parts deionized water; each part of the protein composition comprises 20-30 parts myosin, 30-34 parts actin, and 5-10 parts hydrolyzed collagen. The Lactobacillus bulgaricus is deposited at the China Center for Type Culture Collection on December 11, 2022, with accession number CCTCC S2022087. The preparation method of the chitosan-modified Dropsordry includes the following steps: B1. Cool thionyl chloride to -5~0℃, add humic acid while controlling the temperature, and after the addition is complete, heat to room temperature and stir at a constant temperature for 2~4 hours. After the reaction is completed, quench the reaction, concentrate and dry under reduced pressure to obtain acyl humic acid. B2. Mix the acyl humic acid, chitosan and acetone from step B1, add sodium bicarbonate, heat to 50~60℃ and stir for 10~12h. After the reaction is complete, add and filter, wash with acetone to obtain humic acid grafted modified chitosan. B3. Under an inert protective gas atmosphere, 4,4'-dibromobenzophenone was mixed with chloroform, zinc was added, and the mixture was stirred at room temperature for 1-3 hours. After the reaction was completed, the mixture was filtered, and the filtrate was concentrated and dried under reduced pressure to obtain compound A. B4. Mix compound A from step B3 with silver nitrate, add dichloromethane, stir at room temperature in the dark for 26-30 h, filter after the reaction is complete, and dry to obtain compound B; B5. Mix compound B from step B4, humic acid-grafted modified chitosan from step B2, and Dropsordry. Add a mixed solution of ethanol and dichloromethane and stir at room temperature for 18-20 hours. After stirring, add n-heptane until no precipitate is formed. Filter and wash with ethanol at -10 to -5℃. Dry to obtain chitosan-modified Dropsordry.

2. The pelvic floor muscle repair composition containing supramolecular firming Dropsordry according to claim 1, characterized in that, The method for preparing the Lactobacillus bulgaricus lyophilized powder includes the following steps: A1. Inoculate the activated culture of Lactobacillus bulgaricus into MRS medium at an inoculation rate of 1-2% of the medium volume, and activate it at 30-37℃ for 18-24 hours to obtain seed culture. A2. Inoculate the seed culture from step A1 into a fermenter containing sterilized fermentation medium, adjust the pH to 4-6, and ferment at 35-40℃ for 30-60 hours. 10-15 hours after the start of fermentation, add a fermentation promoter to continue fermentation. After the fermentation is completed, cool down to 25-30℃ and then centrifuge to obtain the bacterial sludge. A3. Mix the bacterial sludge from step A3 with the preservative to make the pH of the mixture 4.5~6.5, and freeze-dry the mixture to obtain Lactobacillus bulgaricus freeze-dried powder; The fermentation promoter comprises the following components in parts by weight: 0.5-0.9 parts calcium myristate, 3-5 parts sodium 2-hydroxyvalerate, 0.05-0.09 parts magnesium chloride, and 20-30 parts Span-80.

3. The pelvic floor muscle repair composition containing supramolecular firming Dropsordry according to claim 2, characterized in that, The fermentation medium, by 100% of its total mass, comprises the following raw materials: The fermentation medium contains 1-3% casein peptone, 1-2% corn starch, 2-4% glucose, 0.5-1% soybean protein powder, 1-3% yeast extract, 0.5-1% beef extract, 0.05-0.1% potassium dihydrogen phosphate, 0.05-0.1% diammonium hydrogen citrate, 0.01-0.1% magnesium sulfate, 0.5-0.8% vitamin E, 0.1-0.5% defoamer, and the balance being water; the pH of the fermentation medium is 5-6.

4. The pelvic floor muscle repair composition containing supramolecular firming Dropsordry according to claim 1, characterized in that, In step B2, the mass ratio of acyl humic acid to chitosan is (1~3):

1.

5. The pelvic floor muscle repair composition containing supramolecular firming Dropsordry according to claim 1, characterized in that, In step B3, the molar ratio of 4,4'-dibromobenzophenone to zinc is 1:(1.2~1.5).

6. The pelvic floor muscle repair composition containing supramolecular firming Dropsordry according to claim 1, characterized in that, In step B5, the mass ratio of compound B, humic acid-grafted modified chitosan, and Dropsordry is (0.2~0.5):1:(3~5).

7. A method for preparing a pelvic floor muscle repair composition containing supramolecular firming Dropsordry as described in any one of claims 1 to 6, characterized in that, Includes the following steps: S1. Mix the protein composition, hyaluronic acid, glycine, and proline evenly to obtain material A; S2. Chitosan-modified Dropsordry is mixed with deionized water to form a supramolecular gel. S3. Under continuous stirring, add material A from step S1 and Lactobacillus bulgaricus lyophilized powder to the supramolecular gel in step S2 to obtain a pelvic floor muscle repair composition containing supramolecular firming Dropsordry.

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