A fumigant, its preparation method and uses

Abstract

The present invention relates to a fumigant, which is made from traditional Chinese medicine raw materials in the following weight ratios: 20 - 40 parts by weight of Astragalus membranaceus, 15 - 25 parts by weight of Paederia scandens, 10 - 20 parts by weight of Rubia cordifolia, 10 - 20 parts by weight of Cnidium monnieri, 5 - 15 parts by weight of Pueraria lobata, 5 - 15 parts by weight of Corydalis yanhusuo, 5 - 15 parts by weight of Cimicifuga foetida, 5 - 15 parts by weight of Citrus reticulata Blanco, 5 - 15 parts by weight of Vaccaria segetalis, 5 - 15 parts by weight of Eucommia ulmoides, 3 - 8 parts by weight of Eugenia caryophyllata, 3 - 8 parts by weight of Foeniculum vulgare Mill. This fumigant can be used for the treatment of pelvic floor dysfunction diseases, especially for improving stress urinary incontinence in menopausal women.

Description

Technical Field

[0001] The present invention belongs to the technical field of fumigants, and particularly relates to a fumigant, a preparation method thereof and uses thereof. Background Art

[0002] Female pelvic floor dysfunction (PFD) occurs frequently in women with a history of childbirth and middle-aged and elderly women, and is mainly caused by the special physiological structure during pregnancy or damage to the pelvic floor support structure. If pelvic floor rehabilitation intervention and treatment are not carried out in a timely manner, it will lead to symptoms such as uterine prolapse, stress urinary incontinence, and chronic pelvic pain. Among them, stress urinary incontinence (SUI) refers to the involuntary leakage of urine caused by a sudden increase in abdominal pressure, but not caused by the detrusor contraction pressure or the bladder wall tension pressure on the urine. According to epidemiological investigations, the incidence of stress urinary incontinence in adult women is about 19%, and the incidence of this disease increases with age. It can be seen that women in menopause are the main population affected by this disease. Since patients with this disease have no enuresis under normal conditions, and urine automatically flows out when the abdominal pressure suddenly increases, women with stress urinary incontinence often experience involuntary urine leakage when the abdominal pressure increases during coughing, sneezing, laughing or exercising, etc., which not only brings inconvenience to the daily life and social interaction of patients, but also brings great mental pressure to patients. It is the most common situation in female pelvic floor dysfunction diseases, seriously affecting the physical health and quality of life of patients. If stress urinary incontinence is not actively treated, the situation of urinary incontinence will not disappear naturally, but will only become more and more serious.

[0003] Female stress urinary incontinence is mainly affected by age or childbirth. The main reason for parturients with this disease is that when the pelvic floor muscles and the posterior urethra of the bladder neck are relaxed, the posterior urethra of the bladder neck gradually moves downward, changing the angle of the bladder neck and the urethra. When the abdominal pressure suddenly increases, the pressure in the bladder will exceed the pressure in the posterior urethra, resulting in stress urinary incontinence. Another part of the parturients suffer from this disease due to nerve damage to the pelvic floor muscles or levator ani muscles caused by childbirth. For menopausal, perimenopausal or elderly women, it is mainly due to the decrease in estrogen secretion, atrophy of the urethral mucosa and submucosal blood vessels, resulting in the loss of the closing function of the urethral mucosa and the weakening of the tension of the intrinsic urethral sphincter or other pelvic floor muscles, making them prone to stress urinary incontinence. For the treatment of early PFD or SUI, non-surgical therapies such as bladder training, pelvic floor muscle exercise, traditional Chinese medicine acupuncture, electrostimulation biofeedback therapy, injection of urethral surrounding fillers, and adrenergic agonists are usually used clinically. When the patient's condition develops to a severe degree, the pelvic floor dysfunction will gradually worsen, and finally only surgical treatment can be performed, such as retropubic bladder urethral suspension; tension-free vaginal midurethral sling surgery, etc. The above intervention methods can, to a certain extent, improve the nerve conduction function by enhancing the tension and contractility of the pelvic floor muscles, thereby enhancing the sphincter muscle strength of the patient and providing a stable support force for the pelvic floor. This method can achieve certain therapeutic effects, but it has little effect on disease control and the prognosis of patients. Therefore, early diagnosis and active acceptance of pelvic floor function recovery treatment can effectively improve the prognosis and reduce the impact of this disease on the physical and mental health of patients.

[0004] Although there are also some methods in the prior art that use traditional Chinese medicine for oral and external use, such as taking decoctions like Buzhong Yiqi Decoction with modifications orally, or treating the disease by means of moxibustion, acupuncture, acupoint application, etc., they mainly focus on the prevention and treatment of postpartum stress urinary incontinence and are mainly used to repair the damage of pelvic floor muscles and nerves caused by childbirth. For example, ZL201310732318.4 discloses a drug for treating senile urinary incontinence, which is composed of the following raw materials in parts by weight: 3-6 parts of Taxillus chinensis, 4-8 parts of Chinese yam, 3-6 parts of Atractylodes macrocephala, 3-6 parts of Cornus officinalis, 2-6 parts of Cortex Moutan, 3-5 parts of Alpinia oxyphylla, 2-4 parts of Ootheca Mantidis, 2-4 parts of Rosa laevigata, 3-5 parts of Poria cocos, 4-6 parts of Astragalus membranaceus, 2-5 parts of Rubus idaeus, 2-5 parts of Glycyrrhiza uralensis, 2-5 parts of Psoralea corylifolia, 2-5 parts of Salvia miltiorrhiza, 1-3 parts of Cimicifuga foetida, 5-7 parts of Euryale ferox. The preparation method of the drug is to select and wash various raw materials, dry them at low temperature, crush them through a 100-mesh sieve, sterilize them, fill them into capsules and use them orally. The literature also gives a traditional Chinese medicine fumigation solution for treating urinary incontinence in parturients, which is obtained by extracting 30 g of Astragalus membranaceus and calcined Dragon Bone, and 15 g of Codonopsis pilosula, Atractylodes macrocephala, Chinese yam, Rubus idaeus, Cuscuta chinensis, Euryale ferox, Ootheca Mantidis, and 10 g of Cimicifuga foetida and roasted Glycyrrhiza uralensis (Observation on the Prevention and Treatment of Postpartum Stress Urinary Incontinence by Traditional Chinese Medicine Fumigation Combined with Pelvic Floor Rehabilitation, Wang Huili, Liu Jianfen, Ye Yuemei, Cheng Xiaoyan; Chinese Journal of Traditional Chinese Medicine Science and Technology, Vol. 29, No. 3, May 2022, pp. 451-452). However, for menopausal patients, due to the decrease in estrogen content, atrophy of the pelvic floor muscles, insufficient blood supply to the pelvic floor structure and tissue, indirectly leading to electrophysiological abnormalities of muscle fibers, resulting in atrophy and weakened muscle strength of the pelvic floor muscles, which are the most important factors causing high abdominal pressure and then stress urinary incontinence. It is not suitable to achieve satisfactory results by simply surgery, exercise, or the prevention and treatment methods for postpartum stress urinary incontinence. Although clinically, continuous administration of a small amount of estrogen is used as an adjuvant treatment for stress urinary incontinence in menopausal women, which has an excitatory effect on the smooth muscle in the urethral wall to increase urethral pressure and relieve urinary incontinence symptoms. However, due to the possible occurrence of endometrial cancer, breast cancer and ovarian cancer caused by long-term use of estrogen, the use of the above methods is greatly restricted. And excluding the influence of multiple births or trauma, the muscle strength of the pelvic floor muscles of most postpartum stress urinary incontinence patients can gradually decrease with the recovery of the body after childbirth; but for menopausal stress urinary incontinence patients, if no means are taken to intervene in the disease process in time, the longer the disease duration, the more severe the symptoms will be with the further decrease of hormone levels. It can be seen that the pathogenic causes, treatment methods, and prognosis of postpartum stress urinary incontinence and menopausal women's stress urinary incontinence are different, and targeted therapeutic drugs are needed.Oral medications have difficulty reaching the pelvic floor area. Existing fumigation medications lack strong specificity and have a slow onset of action, taking about 1 - 2 months to become effective. This makes it difficult for many patients to adhere to the treatment because they cannot quickly see the effects, resulting in poor drug compliance. Currently, there is still a lack of safe, effective, and fast-acting medications and methods for improving climacteric pelvic floor dysfunctional diseases, especially for the prevention and treatment of stress urinary incontinence in menopausal women. Summary of the Invention

[0005] Through long-term practice by the inventors, a fumigant with a fast onset of action and definite effects for improving climacteric pelvic floor dysfunction, especially for improving stress urinary incontinence in menopausal women, has been invented.

[0006] The technical solution of the present invention is as follows:

[0007] The present invention provides a fumigant, which is made from raw materials in the following weight ratios:

[0008]

[0009] Preferably, the fumigant is made from raw materials in the following weight ratios:

[0010]

[0011] More preferably, the fumigant is made from raw materials in the following weight ratios:

[0012]

[0013] The preparation method of the fumigant includes the following steps:

[0014] (1) Weigh dried tangerine peel, chicken manure vine, cnidium fruit, clove, and fennel in doses, mix them, crush them into coarse powder, soak with 2 times the amount of water for 0.5 hour, and then use steam distillation for 2 hours to extract volatile oil. Collect the volatile oil and store it separately. Add water to decoct the medicinal residues twice, each time adding 1 - 5 times the amount of water based on the weight of the above 5 kinds of medicinal materials, decocting for 1 hour each time. Combine the decoction liquids, filter, and obtain traditional Chinese medicine extract A for standby;

[0015] (2) Weigh astragalus root, kudzu root, madder, Corydalis yanhusuo, rhizoma cimicifugae, semen vaccariae, and eucommia ulmoides in doses, crush them into coarse powder, decoct and extract with water twice, each time adding 3 - 8 times the amount of water based on the weight of the above 7 kinds of medicinal materials, decocting for 1.5 hours each time. Combine the decoction liquids, filter, and obtain traditional Chinese medicine extract B for standby;

[0016] (3) Take the volatile oil prepared in step (1), and include it with hydroxypropyl-β-cyclodextrin to obtain an aqueous solution of volatile oil inclusion compound;

[0017] (4) Add the Chinese medicine extract A obtained in step (1) and the Chinese medicine extract B obtained in step (2) to the aqueous solution of the volatile oil inclusion complex obtained in step (3), mix well, and prepare an extraction solution with a crude drug concentration of 0.2 g / ml, thus obtaining the fumigant.

[0018] In the preparation method of the fumigant:

[0019] In step (1), the amounts of water added twice are respectively 3 times the weight of the medicinal materials to be extracted; in step (2), the amounts of water added twice are respectively 5 times the weight of the medicinal materials to be extracted.

[0020] The method for preparing the aqueous solution of the volatile oil inclusion complex in step (3) includes the following steps:

[0021] Measure the volume of the volatile oil prepared in step (1), and prepare a volatile oil ethanol solution with 5 times the volume of the volatile oil; weigh hydroxypropyl-β-cyclodextrin for inclusion at a feeding ratio of 7:1 (g / mL), add the hydroxypropyl-β-cyclodextrin to distilled water with a volume 70 times that of the volatile oil, heat to dissolve, cool down, and obtain an aqueous solution of hydroxypropyl-β-cyclodextrin; under the stirring condition at a constant temperature of 50 °C, slowly add the volatile oil:ethanol solution to the aqueous solution of hydroxypropyl-β-cyclodextrin, and stir for 1 hour to obtain the aqueous solution of the volatile oil inclusion complex.

[0022] The application of the aforementioned fumigant in the preparation of a fumigant for treating pelvic floor dysfunctional diseases, wherein the pelvic floor dysfunctional disease is stress urinary incontinence in menopausal women.

[0023] The raw material drugs used in this application have the following effects.

[0024] Astragalus membranaceus, sweet in taste, slightly warm in nature, attributing to the spleen and lung meridians, having the effects of invigorating qi and ascending yang, strengthening the exterior and consolidating the superficial resistance, promoting diuresis and alleviating edema, and promoting granulation and generating muscle.

[0025] Paederia scandens, sweet, sour and neutral in taste, attributing to the liver, gallbladder, spleen and stomach meridians, having the effects of promoting digestion and strengthening the stomach, resolving phlegm and relieving cough, activating blood circulation and dissipating stasis, and dispelling wind and relieving pain.

[0026] Rubia cordifolia, bitter in taste, cold in nature, attributing to the heart and liver meridians, having the effects of promoting blood circulation to remove stasis, dredging meridians and relieving pain, clearing the heart and removing vexation, and cooling blood and eliminating carbuncles.

[0027] Cnidium monnieri, pungent and bitter in taste, warm in nature, attributing to the kidney meridian, having the effects of killing parasites and relieving itching, warming the kidney and strengthening yang, and drying dampness and expelling wind.

[0028] Pueraria lobata, sweet and pungent in taste, cool in nature, attributing to the spleen and stomach meridians, having the effects of relieving muscle spasms and reducing fever, promoting eruption, promoting fluid production and quenching thirst, and ascending yang and stopping diarrhea.

[0029] Corydalis yanhusuo, pungent and bitter in taste, warm in nature, attributing to the liver and spleen meridians, having the effects of promoting blood circulation; dissipating stasis; regulating qi; relieving pain.

[0030] Cimicifuga foetida L., pungent and slightly sweet in taste, slightly cold in nature, has the effects of dispersing wind and promoting eruption, clearing heat and detoxifying, lifting yang and raising prolapse.

[0031] Tangerine peel, pungent in taste, warm in nature, belonging to the spleen and lung meridians, has the effects of promoting qi circulation and strengthening the spleen, descending adverse qi and resolving phlegm.

[0032] Vaccaria segetalis (Neck.) Garcke, bitter in taste, neutral in nature, belonging to the liver and stomach meridians, has the effects of promoting blood circulation and dredging meridians, promoting lactation and detumescence, and promoting diuresis and relieving stranguria.

[0033] Eucommia ulmoides Oliv., pungent in taste, neutral in nature, entering the liver and kidney meridians, has the effects of tonifying the liver and kidney, strengthening tendons and bones, and lowering blood pressure.

[0034] Flos Caryophylli, pungent in taste, warm in nature, belonging to the spleen, stomach, liver and kidney meridians, has the effects of warming the middle-jiao to dispel cold and regulating qi to relieve pain.

[0035] Fructus Foeniculi, pungent in taste, warm in nature, belonging to the liver, kidney, bladder and stomach meridians, has the effects of warming the kidney and liver, regulating qi to relieve pain, and harmonizing the stomach.

[0036] Beneficial effects

[0037] The main TCM pathogenesis of stress urinary incontinence in menopausal women is mainly caused by reasons such as aging: kidney deficiency and liver depression, qi and blood obstruction forming blood stasis; spleen and stomach weakness, gathering dampness to form phlegm; phlegm and blood stasis interacting and binding, blocking the collaterals, deficiency and excess mingling, affecting the pelvic floor muscle tension and local blood circulation, resulting in bladder incontinence. In the drug prescription of this application, Astragalus membranaceus (Fisch.) Bunge replenishes qi and lifts yang, Cnidium monnieri (L.) Cusson nourishes the kidney yang, Paederia scandens (Lour.) Merr. dispels wind and resolves phlegm, and Rubia cordifolia L. promotes blood circulation to remove blood stasis. These four drugs together are the monarch drugs. Pueraria lobata (Willd.) Ohwi, Cimicifuga foetida L., and Tangerine peel assist Astragalus membranaceus (Fisch.) Bunge to strengthen the effects of lifting yang and promoting qi circulation; Corydalis yanhusuo W. T. Wang ex Z. Y. Su & C. Y. Wu assists Cnidium monnieri (L.) Cusson and Paederia scandens (Lour.) Merr. to strengthen the effects of soothing the liver and promoting blood circulation. These four drugs together are the minister drugs. Vaccaria segetalis (Neck.) Garcke and Eucommia ulmoides Oliv. are used as adjuvants to dredge collaterals and tonify the kidney, and Flos Caryophylli and Fructus Foeniculi are used as guides to warm the middle-jiao to dispel cold. This formula mainly focuses on replenishing qi and lifting yang, and in this application, the method of separately extracting volatile oil is used. Although it increases the content of active ingredients of the drug and can promote the absorption of active ingredients. However, in actual research, it is found that simply increasing the absorption of drugs for replenishing qi and yang and providing warm drugs is instead not conducive to the recovery of the condition of urinary incontinence in menopausal syndrome, slows down the cell proliferation rate, and is not conducive to the accumulation of collagen in the cell interstitium. After adding Rubia cordifolia L. for cooling blood and promoting blood circulation, it neutralizes the overly dry and warm property of the whole formula, not only enhances the proliferative effect of the fumigation solution on fibroblasts, promotes the accumulation of the content of type I and type III collagen in the cell matrix, and can promote the recovery of stress urinary incontinence in menopausal women faster and better. The above-mentioned formula combined with fumigation, hip bath, and exercises such as anal lifting and contraction, the skin and mucous membranes can directly absorb the drug, can improve the local blood circulation, increase the sensitivity of nerves, promote muscle recovery, and slow down muscle atrophy. For stress urinary incontinence in menopausal women, the fumigation agent of this application combined with pelvic floor rehabilitation muscle exercise is significantly superior to simple exercise in terms of the urinary incontinence rating, urine leakage volume, and the impact of urine leakage symptoms on daily life of the patients treated with the fumigation agent of this application after one month, and has a fast onset and stable effect. Detailed implementation mode

[0038] Various traditional Chinese medicine herbs were purchased from Yifeng Pharmacy, and hydroxypropyl-β-cyclodextrin was purchased from Zhiyuan Biotechnology Company.

[0039] Example 1

[0040] Weigh 30g of Astragalus membranaceus, 20g of Paederia scandens, 15g of Pueraria lobata, 15g of Cnidium monnieri, 10g of Rubia cordifolia, 10g of Corydalis yanhusuo, 10g of Cimicifuga foetida, 10g of Citrus reticulata Blanco, 10g of Vaccaria segetalis, 10g of Eucommia ulmoides, 5g of Eugenia caryophyllata, and 5g of Foeniculum vulgare Mill. respectively for standby.

[0041] Weigh Citrus reticulata Blanco, Paederia scandens, Cnidium monnieri, Eugenia caryophyllata, and Foeniculum vulgare Mill. according to the dosage, mix them, crush them into coarse powder, soak them in 2 times of water for 0.5 hours, then use steam distillation for 2 hours to extract volatile oil, and collect the volatile oil for storage in another container; decoct the medicinal residues with water twice, each time adding 3 times of water with the weight of the above 5 kinds of medicinal materials, decocting for 1 hour each time, combine the decoction liquid, filter it, and obtain traditional Chinese medicine extract A for standby.

[0042] Weigh Astragalus membranaceus, Pueraria lobata, Rubia cordifolia, Corydalis yanhusuo, Cimicifuga foetida, Vaccaria segetalis, and Eucommia ulmoides according to the dosage, crush them into coarse powder, decoct and extract them with water twice, each time adding 5 times of water with the weight of the above 7 kinds of medicinal materials, decocting for 1.5 hours each time, combine the decoction liquid, filter it, and obtain traditional Chinese medicine extract B for standby.

[0043] Measure the volume of the volatile oil obtained by steam distillation extraction and preparation, and prepare a volatile oil ethanol solution with 5 times the volume of the volatile oil. Weigh hydroxypropyl-β-cyclodextrin for inclusion according to the feeding ratio of 7:1 (g / ML), add hydroxypropyl-β-cyclodextrin to distilled water with 70 times the volume of the volatile oil, heat to dissolve it, and cool down to obtain a hydroxypropyl-β-cyclodextrin aqueous solution. Under the condition of stirring at a constant temperature of 50°C, slowly add the volatile oil:ethanol solution to the hydroxypropyl-β-cyclodextrin aqueous solution and stir for 1 hour to obtain a volatile oil inclusion aqueous solution.

[0044] Add traditional Chinese medicine extract A and traditional Chinese medicine extract B to the volatile oil inclusion aqueous solution, mix them evenly, and make an extraction solution with a crude drug concentration of 0.2g / ml, that is, the fumigant of Example 1 is obtained.

[0045] Example 2

[0046] Weigh 20g of Astragalus membranaceus, 20g of Paederia scandens, 20g of Rubia cordifolia, 20g of Cnidium monnieri, 15g of Pueraria lobata, 15g of Corydalis yanhusuo, 15g of Cimicifuga foetida, 15g of Citrus reticulata Blanco, 15g of Vaccaria segetalis, 15g of Eucommia ulmoides, 8g of Eugenia caryophyllata, and 8g of Foeniculum vulgare Mill. respectively for standby.

[0047] Weigh dried tangerine peel, paederia scandens, cnidium monnieri, clove, and fennel according to the dosage, mix them, and crush them into coarse powder. After soaking in 2 times the amount of water for 0.5 hours, perform steam distillation for 2 hours to extract the volatile oil, and collect and store the volatile oil separately; decoct the medicinal residues twice with water, each time adding 1 time the weight of the above 5 kinds of medicinal materials of water, decocting for 1 hour each time, combine the decoction liquids, filter, and obtain the traditional Chinese medicine extract A for standby.

[0048] Weigh astragalus membranaceus, pueraria lobata, rubia cordifolia, rhizoma corydalis, rhizoma cimicifugae, vaccaria segetalis, and eucommia ulmoides according to the dosage, crush them into coarse powder, and perform water decoction extraction twice, each time adding 8 times the weight of the above 7 kinds of medicinal materials of water, decocting for 1.5 hours each time, combine the decoction liquids, filter, and obtain the traditional Chinese medicine extract B for standby.

[0049] Measure the volume of the volatile oil obtained by steam distillation extraction and preparation, and prepare a volatile oil ethanol solution with 5 times the volume of the volatile oil of ethanol. Weigh hydroxypropyl-β-cyclodextrin for inclusion at a feeding ratio of 7:1 (g / ML), add hydroxypropyl-β-cyclodextrin to 70 times the volume of the volatile oil of distilled water, heat to dissolve, cool down, and obtain a hydroxypropyl-β-cyclodextrin aqueous solution. Under the stirring condition at a constant temperature of 50 °C, slowly add the volatile oil:ethanol solution to the hydroxypropyl-β-cyclodextrin aqueous solution and stir for 1 hour to obtain a volatile oil inclusion complex aqueous solution.

[0050] Add the traditional Chinese medicine extract A and the traditional Chinese medicine extract B to the volatile oil inclusion complex aqueous solution, mix well, and prepare an extraction solution with a crude drug concentration of 0.2 g / ml, thus obtaining the fumigant of Example 2.

[0051] Example 3

[0052] Weigh 35 g of astragalus membranaceus, 23 g of paederia scandens, 12 g of rubia cordifolia, 12 g of cnidium monnieri, 13 g of pueraria lobata, 13 g of rhizoma corydalis, 13 g of rhizoma cimicifugae, 13 g of dried tangerine peel, 13 g of vaccaria segetalis, 7 g of eucommia ulmoides, 4 g of clove, and 4 g of fennel for standby.

[0053] Weigh dried tangerine peel, paederia scandens, cnidium monnieri, clove, and fennel according to the dosage, mix them, and crush them into coarse powder. After soaking in 2 times the amount of water for 0.5 hours, perform steam distillation for 2 hours to extract the volatile oil, and collect and store the volatile oil separately; decoct the medicinal residues twice with water, each time adding 3 times the weight of the above 5 kinds of medicinal materials of water, decocting for 1 hour each time, combine the decoction liquids, filter, and obtain the traditional Chinese medicine extract A for standby.

[0054] Weigh astragalus membranaceus, pueraria lobata, rubia cordifolia, rhizoma corydalis, rhizoma cimicifugae, vaccaria segetalis, and eucommia ulmoides according to the dosage, crush them into coarse powder, and perform water decoction extraction twice, each time adding 4 times the weight of the above 7 kinds of medicinal materials of water, decocting for 1.5 hours each time, combine the decoction liquids, filter, and obtain the traditional Chinese medicine extract B for standby.

[0055] Measure the volume of the volatile oil obtained by steam distillation extraction, and prepare an ethanol solution of the volatile oil by taking ethanol with a volume 5 times that of the volatile oil. Weigh hydroxypropyl-β-cyclodextrin for inclusion at a feeding ratio of 7:1 (g / mL). Add the hydroxypropyl-β-cyclodextrin to distilled water with a volume 70 times that of the volatile oil, heat to dissolve it, and then cool down to obtain an aqueous solution of hydroxypropyl-β-cyclodextrin. Under the condition of stirring at a constant temperature of 50 °C, slowly add the volatile oil:ethanol solution to the aqueous solution of hydroxypropyl-β-cyclodextrin and stir for 1 hour to obtain an aqueous solution of the volatile oil inclusion complex.

[0056] Add both traditional Chinese medicine extract A and traditional Chinese medicine extract B to the aqueous solution of the volatile oil inclusion complex, mix well, and prepare an extraction solution with a crude drug concentration of 0.2 g / mL to obtain the fumigant of Example 3.

[0057] Comparative Example 1

[0058] Weigh 30 g of Astragalus membranaceus, 20 g of Paederia scandens, 15 g of Rubia cordifolia, 15 g of Cnidium monnieri, 10 g of Pueraria lobata, 10 g of Corydalis yanhusuo, 10 g of Cimicifuga foetida, 10 g of Citrus reticulata Blanco, 10 g of Vaccaria segetalis, 10 g of Eucommia ulmoides, 5 g of Eugenia caryophyllata, and 5 g of Foeniculum vulgare Mill. Mix all the medicinal materials, crush them into coarse powder, extract by boiling with water twice, with the water addition amount each time being 5 times the total weight of the raw medicinal materials, boil for 1.5 hours each time, combine the decoctions, filter, and prepare an extraction solution with a crude drug concentration of 0.2 g / mL to obtain the fumigant of Comparative Example 1.

[0059] Comparative Example 2

[0060] Subtract Rubia cordifolia from the raw materials of Example 1, and the remaining raw materials and preparation process are exactly the same as those of Example 1, and finally obtain the fumigant of Comparative Example 2.

[0061] Comparative Example 3

[0062] For the drug for treating senile urinary incontinence according to ZL201310732318.4, weigh 10 g of Taxillus chinensis, 15 g of Dioscorea opposita Thunb., 10 g of Atractylodes macrocephala Koidz., 10 g of Cornus officinalis Sieb. et Zucc., 10 g of Paeonia suffruticosa Andr., 10 g of Alpinia oxyphylla Miq., 7.5 g of Ootheca mantidis, 7.5 g of Rosa laevigata Michx., 10 g of Poria cocos (Schw.) Wolf, 12.5 g of Astragalus membranaceus (Fisch.) Bunge, 7.5 g of Rubus idaeus L., 5 g of Glycyrrhiza uralensis Fisch., 7.5 g of Psoralea corylifolia L., 10 g of Salvia miltiorrhiza Bunge, 2.5 g of Cimicifuga foetida L., and 15 g of Euryale ferox Salisb. Mix all the medicinal materials, crush them into coarse powder, extract by boiling with water twice, with the water addition amount each time being 5 times the total weight of all the raw medicinal materials, boil for 1.5 hours each time, combine the decoctions, filter, and prepare an extraction solution with a crude drug concentration of 0.2 g / mL to obtain the fumigant of Comparative Example 1.

[0063] Cell experiment

[0064] (1) Isolate and culture rat pubococcygeal myofibroblasts (RPMF)

[0065] Experimental animals: Five female SD rats were selected, raised for 2 weeks, with an average body weight of (200 ± 30) g, provided by the Experimental Animal Center of China Pharmaceutical University, and all were in good health after physical examination.

[0066] Cell culture medium: DMEM + 10% FBS + 100 μg / mL streptomycin + 100 U / mL penicillin + 0.2% MEM Non-Essential Amino Acids (100×) + 0.2% MEM Vitamin Solution (100×) + 2 mM Sodium Pyruvate + 0.4% MEM Amino Acids (50×).

[0067] Culture conditions: Constant temperature at 37°C, with a CO2 content of 5% and an O2 content of 2% (tri-gas incubator).

[0068] After sacrificing SD female rats by air embolism in the auricular marginal vein, soak them in ethanol for 5 minutes, take 3 - 5 mm of the pubococcygeal muscle and the surrounding fascia, place it in phosphate-buffered saline (PBS) containing penicillin and streptomycin, immediately enter the laminar flow hood, wash it and place it in a DMEM culture dish containing 10% fetal bovine serum. Use ophthalmic scissors to cut the tissue into pieces of 1 mm × 1 mm × 1 mm size, rinse it twice with PBS, add 1 ml of 0.1% type II collagenase and culture medium to 5 ml, shake it in a constant temperature shaker at 37°C for 3 - 4 hours, mix it every 30 minutes, filter it through a 200-mesh sieve, centrifuge it for 5 minutes (1000 r / min), discard the supernatant, take the lower precipitate and place it in a culture flask, add culture medium to 5 ml, resuspend it, and a small number of round cells can be seen under the microscope. Change the medium every day. When the cells grow to 90%, perform cell passage. When passing to the 3 - 4th generation by differential adhesion method, relatively pure fibroblasts are obtained and passaged to the 10th generation. The cells are used for cytospin, and immunohistochemical staining of vimentin and cytokeratin is performed. Under an inverted microscope, it is observed that most fibroblasts are mainly spindle-shaped. After immunohistochemical staining, vimentin is positive, brown granules are located in the cytoplasm, the nucleus is blue, single nucleus, and cytokeratin staining is negative. After identification, the cells express vimentin and are fibroblasts, and the cell purity is more than 90%.

[0069] The RPMF cells cultured and grown to more than 80% were rinsed 2 - 3 times with PBS, 1 mL of 0.25% trypsin was added and digested at room temperature for 30 - 60 s. The cell wall was gently tapped to make the cells detached. 2 times the volume of medium containing fetal bovine serum was added to terminate the digestion. It was pipetted evenly with a pipette gun, centrifuged at 1000 r / min for 8 min, the supernatant was removed, and an appropriate amount of complete medium was added to the precipitate to make a single cell suspension. The concentration was adjusted, and the RPMF in the logarithmic growth phase with good condition was used for the experiment.

[0070] (2) Effects of different fumigants on the proliferation activity of rat pubococcygeus muscle fibroblasts

[0071] Take the RPMF in the logarithmic growth phase and inoculate them at 1×10 5 cells / mL into multiple 96 - well culture plates, 90 μL was inoculated in each well, placed in a cell culture incubator at 37 °C and 5% CO2 for culture, the O2 content was 2%, taken out after 24 h, and it was observed under the microscope that all the cells had adhered. The grouping was as follows: blank control group (containing an equal amount of DMEM culture medium), negative control group (containing an equal amount of cell resuspension) and 6 experimental groups. The experimental groups were respectively added with the fumigants of Example 1 - 3 and Comparative Example 1 - 3, 10 μL / well, so that their final concentrations were 100 μg / mL respectively. The blank control group and the negative control group were added with an equal amount of DMEM culture medium in each well. Each sample was set with at least 3 replicates. After shaking evenly, it was placed in a standard incubator for continuous culture. After 24, 48, and 72 h of drug administration, 20 μL of MTT was added to each well, and after continuous culture for 4 h, 150 μL of DMSO was added to each well. After shaking evenly, the absorbance value (OD) of each well was measured at a wavelength of 490 nm with an enzyme - linked immunosorbent assay (ELISA) reader. The effects of different fumigants on the OD values of RPMF cells at different time periods are shown in Table 1.

[0072] Statistical analysis was performed using SPSS 17.0 statistical software. Each detected data was expressed as , t - test was performed on the data of each group, and analysis of variance was performed on the time factor. A difference with P < 0.05 was considered statistically significant.

[0073] Table 1 Effects of different fumigants with the same concentration on the OD values of RPMF cells at different time periods

[0074] Group Concentration (μg / mL) 24h 48h 72h Negative control group 0 0.797±0.054 0.533±0.085 0.357±0.013 Blank control group 0 0.205±0.012** 0.263±0.029** 0.281±0.011* Example 1 100 0.987±0.074** 0.843±0.114** 0.919±0.074** Example 2 100 0.957±0.026** 0.903±0.083** 0.799±0.015** Example 3 100 1.063±0.040** 0.776±0.049** 0.831±0.038** Comparative example 1 100 0.927±0.018** 0.746±0.018** 0.716±0.027** Comparative example 2 100 0.868±0.053▲ 0.679±0.033* 0.612±0.093** Comparative example 3 100 0.975±0.025** 0.803±0.016** 0.615±0.017**

[0075] Note: Compared with the negative control group, *P < 0.05, **P < 0.01, ▲P > 0.05.

[0076] As shown in Table 1, there was a statistically significant difference between the blank control group (DMEM medium containing 10% inactivated fetal bovine serum) and the negative control group (P<0.001), indicating that the OD value of the negative control group was much larger than that of the blank control group. It can be considered that the solvent had a relatively small effect on RPMF cells, but the influence of the solvent in the blank group on the experimental results still needed to be excluded.

[0077] The proliferation survival rate of RPMF cells with different fumigation solutions was calculated according to the following formula:

[0078] Survival rate (%) = (OD 实验组 - OD 空白对照组 ) / (OD 阴性对照组 - OD 空白对照组 ) × 100%. The results of the proliferation survival rate of cells in each experimental group are shown in Table 2.

[0079] Table 2 Effects of different fumigation solutions with the same concentration on the proliferation survival rate of RPMF cells at different time lengths

[0080]

[0081]

[0082] Note: Compared with the negative control group, *P<0.05, **P<0.01, ▲P>0.05.

[0083] (3) Immunohistochemical detection of the expression of type I collagen and type III collagen in cells of each group at 72 h of culture

[0084] Cells of the negative control group and each experimental group were collected separately, and the total cellular proteins and total RNAs were extracted. The expression of type I and type III collagens in cells of each group was detected by Western blot experiment, and the relative expression levels of type I collagen and type III collagen mRNAs were detected by PCR. The primer sequences were designed according to the gene sequences of type I and type III collagens in GenBank, as shown in Table 3 specifically.

[0085] Table 3 Table of primer sequences for collagen

[0086]

[0087] The experimental results showed that the expression levels of type I collagen and type III collagen, as well as the relative expression levels of mRNAs, in the cells of Examples 1 - 3 cultured for 72 h were higher than those of the negative control group and Comparative Examples 1 - 3, and the difference was statistically significant (P = 0.000), while there was no obvious difference in the expression levels of type I collagen and type III collagen among Examples 1 - 3.

[0088] (4) Experimental results:

[0089] Fibroblasts are the main effector cells for synthesizing extracellular matrix in the pelvic floor. The main histological and pathological features of the occurrence of PFD or SUI are as follows: the mechanical resistance of connective tissues such as pelvic floor muscles, fascia, and ligaments decreases. The above changes are due to the abnormal metabolism of extracellular matrix in pelvic floor tissues, especially the decrease in the synthesis of type I / III collagen and the increase in degradation by fibroblasts. Therefore, the proliferative ability of fibroblasts in pelvic floor tissues and the content of type I / III collagen in cells are considered important pathological factors affecting the onset and recovery of PFD or SUI. Therefore, when treating pelvic floor dysfunctional diseases, such as stress urinary incontinence in menopausal women, whether the therapeutic drug can relieve or even reverse the decrease in the synthesis of type I / III collagen and the increase in degradation by fibroblasts; and even achieve the effect of promoting the proliferation of fibroblasts in pelvic floor tissues is one of the key factors for the drug to effectively improve PFD and SUI.

[0090] Under the microscope, it can be seen that when RPMF cells grow normally, they are long spindle-shaped, the cell nucleus is nearly round, single nucleus, with obvious polarity, and each cell has 3-5 antennae, and the boundary is clear. When the density of RPMF cells is high, there are more connections between cell bodies, the boundary is not obvious, and the cells are arranged in a vortex shape. When the fumigation liquids of Comparative Examples 1 and 3 act on RPMF cells at a concentration of 100 μg / mL, the cell morphology is not much different from that of the negative control group, the cell growth rate is slightly faster than that of the negative control group, or is only faster than that of the negative control group at 72 hours. When the fumigation liquid of Comparative Example 2 acts on RPMF cells at a concentration of 100 μg / mL, the cell growth rate is always lower than that of the negative control group. During the whole process from 24h to 72h when the fumigation liquids of Examples 1-3 act on RPMF cells at a concentration of 100 μg / mL, the proliferation rate of RPMF cells is always faster than that of the negative control group, and the cell density is significantly higher after 48h, and the cells are arranged in a vortex shape. It can be seen that compared with the fumigation liquids of Comparative Examples 1-3, the fumigation liquids of Examples 1-3 have a significant difference in the proliferation of RPMF cells. The fumigation liquid of the present application has the effect of promoting the proliferation of these fibroblasts in RPMF cells, and can promote the synthesis of type I / III collagen in cells and increase the content of type I / III collagen, while the fumigation liquids of Comparative Examples 1-3 cannot achieve the above effects. And the fumigation liquid of Example 1 has a better proliferation effect on fibroblasts than the fumigation liquids of Examples 2-3. Thus, it can be seen that the fumigation liquid formula prepared by the method of the present application has a better effect on improving the recovery and proliferation of fibroblasts in the pelvic floor muscles.

[0091] Clinical experiment

[0092] A total of 60 female patients with symptoms of stress urinary incontinence during menopause were recruited, including 30 patients with grade I and 30 patients with grade II stress urinary incontinence. The diagnostic criteria refer to: "Resident Standardized Training Knowledge Manual", edited by Zeng Xiandong and Zhang Man, published by Liaoning Science and Technology Publishing House, the first edition, the first printing, in May 2019, pages 532 - 533. The selected patients need to meet the following requirements simultaneously:

[0093] 1) Women aged 45 - 55 years old, who have been menopausal for more than 1 year.

[0094] 2) Have symptoms of grade I or II stress urinary incontinence for more than 3 months.

[0095] Grading of stress urinary incontinence:

[0096] Grade I: Urinary incontinence only occurs under severe stress, such as coughing, sneezing or jogging.

[0097] Grade II: Urinary incontinence occurs under moderate stress, such as rapid movement, going up and down stairs.

[0098] Grade III: Urinary incontinence occurs under mild stress, such as occurring immediately when standing, and urine can be controlled in the supine position.

[0099] 3) Good compliance, informed consent, and willing to accept clinical data observation.

[0100] Other exclusion criteria:

[0101] (1) Those with breast malignant tumors, uterine malignant tumors, ovarian tumors; those with severe breast hyperplasia, endometrial polyps;

[0102] (2) Patients with severe cardiovascular and cerebrovascular diseases, those with severe heart, liver, and kidney insufficiency; patients with hyperthyroidism;

[0103] (3) Those who have taken estrogen or progesterone drugs within the past 2 months,

[0104] (4) Those who have taken traditional Chinese medicine decoctions or proprietary Chinese medicines for treating menopausal syndrome or urinary incontinence within the past month.

[0105] The patients with grade I and grade II stress urinary incontinence were randomly divided into 5 groups, with 6 people in each group. Then, 1 group of grade I urinary incontinence patients and 1 group of grade II urinary incontinence patients were randomly combined into a new experimental group, with 12 people in each group, for a total of 5 groups. Each patient carried out routine Kegel exercises 3 times a day, each time for 3 minutes, to improve the strength of the pelvic floor muscles. On the basis of Kegel exercises for each group, fumigation treatment was carried out respectively, each time for 30 minutes. When performing fumigation treatment, it was carried out once a day. One week was a cycle, with fumigation for 5 days a week and 2 days of rest. Treatment and observation were carried out continuously for 6 weeks. Among them, the control group was fumigated with distilled water, and the other four groups were fumigated with the fumigants of Example 1 and Comparative Examples 1 - 3 respectively.

[0106] Equipment used: Kangjingxing intelligent fumigation machine, ZD-168, 2.6 liters. Sit-type steam sauna box. Timer.

[0107] Fumigation liquid used: Distilled water, fumigants of Example 1 and Comparative Examples 1-3.

[0108] Usage method: After the patient washes the vulva with warm water, sit in the steam sauna box, seal the part below the abdomen with a fumigation cover, raise the steam outlet of the fumigation machine into the steam sauna box, pay attention to keeping a certain distance between the steam outlet and the patient to prevent scalding, and perform steam bath for the lower body. Each time, add 2L of fumigation liquid into a container, heat it to boiling, and prevent the hot air from escaping for fumigation.

[0109] Evaluate the degree of stress urinary incontinence grade I or II symptoms every 7 days and record the urine leakage amount of the patient in the urine pad test once. The test method of the urine pad test is as follows: Before the test, take a dry urine pad, weigh it and record. Then empty the urine in the bladder, place it under the body, weigh the urine pad in advance, drink 500 mL of water, finish drinking it within 15 minutes, go up and down 5 floors of stairs once, walk for half an hour, do 10 abdominal pressure actions and squats continuously, listen to the sound of running water for 1 minute, take off the urine pad after 1 hour and weigh it (1 g = 1 mL), and calculate the total urine volume. All data are incorporated into the SPSS 20.0 software system for comparison and calculation of the test value. The urine leakage amount in the urine pad test is subjected to a t-test. When P < 0.05, the comparison difference is statistically significant.

[0110] Efficacy criteria

[0111] Cure: The symptoms of stress urinary incontinence completely disappear, and there is no urine leakage phenomenon in the urine pad test.

[0112] Marked effect: Stress urinary incontinence grade II changes to grade I.

[0113] Effective: The rating of stress urinary incontinence does not change, but the urine leakage phenomenon in the urine pad test improves.

[0114] Ineffective: The rating of stress urinary incontinence and the urine leakage condition in the urine pad test remain unchanged or worsen.

[0115] Experimental results

[0116] Table 4 Record of the results of treating menopausal stress urinary incontinence in the control group

[0117] Cured (persons) Markedly effective (persons) Effective (persons) Ineffective (persons) Week 1 0 0 2 10 Week 2 0 0 2 10 Week 3 0 0 4 8 Week 4 0 3 3 6 Week 5 1 2 3 6 Week 6 2 3 1 6

[0118] Table 5 Record of the results of treating menopausal stress urinary incontinence with the fumigant of Example 1

[0119] Cured (persons) Markedly effective (persons) Effective (persons) Ineffective (persons) Week 1 0 2 4 6 Week 2 3 5 1 3 Week 3 5 3 2 2 Week 4 7 2 1 2 Week 5 7 2 2 1 Week 6 7 2 2 1

[0120] Table 6 Record of the results of treating menopausal stress urinary incontinence with the fumigant of Comparative Example 1

[0121] Cured (persons) Markedly effective (persons) Effective (persons) Ineffective (persons) Week 1 0 0 5 7 Week 2 0 0 5 7 Week 3 2 1 4 5 Week 4 4 4 0 4 Week 5 5 3 1 3 Week 6 6 2 2 2

[0122] Table 7 Record of the Results of the Fumigant in Comparative Example 2 for Treating Menopausal Stress Urinary Incontinence

[0123] Cured (persons) Markedly effective (persons) Effective (persons) Ineffective (persons) Week 1 0 0 3 9 Week 2 0 0 4 8 Week 3 1 1 3 7 Week 4 1 3 3 5 Week 5 2 2 3 5 Week 6 2 3 2 5

[0124] Table 8 Record of the Results of the Fumigant in Comparative Example 3 for Treating Menopausal Stress Urinary Incontinence

[0125]

[0126]

[0127] In addition, during the follow-up 3 months after the end of the experiment, 1 cured patient in the control group had a recurrence, while none of the cured patients treated with the fumigant had a recurrence.

[0128] As can be seen from the above results, the fumigant in Example 1 has the best treatment effect. Not only are there the most people whose urinary incontinence and urine leakage conditions are improved in the first week, but also the number of people with grade II urinary incontinence who start to improve and are finally cured is the largest and fastest. It can also be seen from the above experiment that the treatment effects of Comparative Example 1 and Comparative Example 3 are similar, but Comparative Example 1 is better than Comparative Example 3, and the treatment effect of Comparative Example 2 is worse, but still better than the control group. The treatment effect of the control group using only Kegel exercises to treat menopausal stress urinary incontinence is very ordinary. Not only does it take effect slowly, but after 6 weeks, half of the subjects still have no improvement in their urinary incontinence symptoms, and there is also a risk of recurrence. It can be seen that the compatibility and preparation process of the fumigant in this application are reasonable, and it is a product with a fast onset, good effect, safe use, and high compliance for treating menopausal stress urinary incontinence.

Claims

1. A fumigant for treating stress urinary incontinence in menopausal women, characterized in that: It is made from the raw materials with the following weight ratios: Astragalus membranaceus 20 - 40 parts by weight; Paederia scandens 15 - 25 parts by weight; Rubia cordifolia 10 - 20 parts by weight; Cnidium monnieri 10 - 20 parts by weight: Pueraria lobata 5 - 15 parts by weight; Corydalis yanhusuo 5 - 15 parts by weight; Cimicifuga foetida 5 - 15 parts by weight; Citrus reticulata Blanco 5 - 15 parts by weight; Vaccaria segetalis 5 - 15 parts by weight; Eucommia ulmoides 5 - 15 parts by weight; Syzygium aromaticum 2 - 8 parts by weight; Foeniculum vulgare Mill. 2 - 8 parts by weight.

2. The fumigant according to claim 1, wherein: It is made from the raw materials with the following weight ratios: Astragalus membranaceus 25 - 35 parts by weight; Paederia scandens 17 - 23 parts by weight; Rubia cordifolia 12 - 18 parts by weight; Cnidium monnieri 12 - 18 parts by weight; Pueraria lobata 7 - 13 parts by weight; Corydalis yanhusuo 7 - 13 parts by weight; Cimicifuga foetida 7 - 13 parts by weight; Citrus reticulata Blanco 7 - 13 parts by weight; Vaccaria segetalis 7 - 13 parts by weight; Eucommia ulmoides 7 - 13 parts by weight; Syzygium aromaticum 4 - 6 parts by weight; Foeniculum vulgare Mill. 4 - 6 parts by weight.

3. The fumigant according to claim 1, wherein: It is made from the raw materials with the following weight ratios: Astragalus membranaceus 30 parts by weight; Paederia scandens 20 parts by weight; Rubia cordifolia 15 parts by weight; Cnidium monnieri 15 parts by weight; Pueraria lobata 10 parts by weight; Corydalis yanhusuo 10 parts by weight; Cimicifuga foetida 10 parts by weight; Citrus reticulata Blanco 10 parts by weight; Vaccaria segetalis 10 parts by weight; Eucommia ulmoides 10 parts by weight; Syzygium aromaticum 5 parts by weight; Foeniculum vulgare Mill. 5 parts by weight.

4. A method for preparing the fumigant according to any one of claims 1 to 3, characterized in that: It includes the following steps: (1) Weigh Citrus reticulata Blanco, Paederia scandens, Cnidium monnieri, Syzygium aromaticum, and Foeniculum vulgare Mill. according to the dosage, mix them, crush them into coarse powder, soak them in 2 times the amount of water for 0.5 hours, then use steam distillation for 2 hours to extract the volatile oil, and collect the volatile oil for storage in another container; decoct the medicinal residues with water twice, each time adding 1 - 5 times the amount of water of the weight of the above 5 kinds of medicinal materials, decocting for 1 hour each time, combine the decoction liquids, filter, to obtain the traditional Chinese medicine extract A for standby; (2) Weigh Astragalus membranaceus, Pueraria lobata, Rubia cordifolia, Corydalis yanhusuo, Cimicifuga foetida, Vaccaria segetalis, and Eucommia ulmoides according to the dosage, crush them into coarse powder, extract them with water by decoction twice, each time adding 3 - 8 times the amount of water of the weight of the above 7 kinds of medicinal materials, decocting for 1.5 hours each time, combine the decoction liquids, filter, to obtain the traditional Chinese medicine extract B for standby; (3) Take the volatile oil prepared in step (1), and include it with hydroxypropyl - β - cyclodextrin to obtain an aqueous solution of the volatile oil inclusion complex; (4) Add the traditional Chinese medicine extract A obtained in step (1) and the traditional Chinese medicine extract B obtained in step (2) into the aqueous solution of the volatile oil inclusion complex obtained in step (3), mix well, and make an extraction solution with a crude drug concentration of 0.2 g / ml, then the fumigant is obtained.

5. The preparation method of the fumigant according to claim 4, characterized in that: In step (1), the water addition amounts for the two - time water decoction are respectively 3 times the weight of the extracted medicinal materials; in step (2), the two - time water addition amounts are respectively 5 times the weight of the extracted medicinal materials.

6. The preparation method of the fumigant according to claim 5, characterized in that: Step (3) includes the following steps: Measure the volume of the volatile oil prepared in step (1), and prepare a volatile oil ethanol solution with 5 times the volume of the volatile oil; Weigh hydroxypropyl - β - cyclodextrin for inclusion at a feeding ratio of 7:1 in g / ml, add hydroxypropyl - β - cyclodextrin to 70 times the volume of the volatile oil of distilled water, heat to dissolve it, and cool down to obtain an aqueous solution of hydroxypropyl - β - cyclodextrin; Under the condition of stirring at a constant temperature of 50 °C, the volatile oil ethanol solution was slowly added to the hydroxypropyl-β-cyclodextrin aqueous solution and stirred for 1 hour to obtain the aqueous solution of the volatile oil inclusion compound.

7. Use of the fumigant according to any one of claims 1-3 or the fumigant prepared by the preparation method according to any one of claims 4-6 in the preparation of a fumigant for treating stress urinary incontinence in menopausal women.

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