Sperm-producing soup as well as preparation method, application and product thereof

Abstract

The invention belongs to the technical field of traditional Chinese medicines, and particularly relates to sperm-producing decoction as well as a preparation method, application and a product thereof. The sperm-producing decoction is prepared from the following raw materials in parts by weight: 5 to 15 parts of radix salviae miltiorrhizae, 5 to 15 parts of rhizoma chuanxiong, 5 to 15 parts of radix angelicae sinensis, 10 to 25 parts of herba epimedii, 10 to 25 parts of fructus cnidii, 10 to 25 parts of semen cuscutae, 5 to 15 parts of fructus rubi and 5 to 15 parts of fructus schisandrae chinensis. The spermatogenic decoction can improve testis injury caused by an anti-inflammatory and anti-immune drug tripterygium wilfordii hormone, improve oxidative injury of testis tissues, regulate the expression level of antioxidant stress related proteins Nrf2 and HO-1, improve the phenomena of testis diminishment, seminiferous tubule damage, sperm number reduction, sperm motility weakening, testis spermatogenic cell apoptosis and the like, and can be used for preparing the anti-inflammatory and anti-immune drug. And the expression level of apoptosis-related proteins Bcl-2 and Bax of the testicular spermatogenic cells is regulated.

Description

Technical Field

[0001] The invention belongs to the technical field of traditional Chinese medicine, and particularly relates to a shengjing decoction, a preparation method thereof, an application thereof, and a product thereof. Background Art

[0002] Triptolide (TP), also known as triptolide or triptolide alcohol, is an epoxyditerpene lactone compound extracted from the roots, leaves, flowers, and fruits of the Celastraceae plant Tripterygium wilfordii. It exhibits anti-inflammatory and immunosuppressive effects and is widely used clinically to treat rheumatoid arthritis, inflammation, and tumors. However, when used to treat inflammatory conditions in the human body, TP poses a risk of accumulation, which can severely damage the reproductive system. Overdose of TP can lead to adverse reactions such as testicular atrophy, hormonal imbalances, and oligospermia.

[0003] Previous studies have shown that TP-induced oxidative stress and apoptosis are the primary mechanisms of its reproductive toxicity, primarily inducing ROS accumulation in testicular tissue, leading to spermatogenesis impairment and testicular damage. Therefore, male spermatogenesis impairment caused by TP-induced oxidative stress may be restored by activating intracellular antioxidant levels.

[0004] Patent publication number CN119236024A discloses the use of Shugan Bushen Yulin Decoction in the preparation of a drug for treating hypothyroidism-related asthenospermia. The decoction comprises 12-18g of Tribulus terrestris, 12-18g of Curcuma aromatica, 12-18g of Lycium barbarum, 12-18g of Cuscuta seed, 12-18g of Chuanxiong, 8-12g of antler glue (melted), 8-12g of Rubus idaeus, 8-12g of Astragalus complanatus, and 8-12g of Schisandra chinensis. The decoction can repair testicular oxidative stress damage caused by hypothyroidism, but does not specifically alleviate TP-induced testicular oxidative stress damage and hypospermatogenesis.

[0005] A journal article (Yin Jinlong, 2013) disclosed the effects of Wuzi Yanzong compound on oxidative stress damage and cell apoptosis of testicular Sertoli cells. Wuzi Yanzong compound is composed of 16g of wolfberry fruit, 16g of dodder seed, 8g of raspberry, 4g of plantain seed, and 2g of schisandra chinensis. The study found that Wuzi Yanzong compound can improve the growth activity of testicular SC in the oxidative damage model group, increase the intracellular SOD level, reduce the MDA content, reduce cell apoptosis, and improve cell survival rate, but it did not specifically alleviate TP-induced testicular oxidative stress damage and spermatogenesis impairment.

[0006] A journal article (Bi Mingshuai, 2015) reported on research progress on the effects of antioxidant Chinese herbal medicines on testicular spermatogenesis. Among them, an aqueous extract of Cuscuta chinensis, containing flavonoids, inhibited oxidative damage and apoptosis in testicular cells through its antioxidant effects. Other Chinese herbs, such as Salvia miltiorrhiza, also contain flavonoids. Salvia miltiorrhiza extracts, containing phenolic acids, can reduce apoptosis in spermatogenic cells. Chinese herbs such as Angelica sinensis and Chuanxiong also contain phenolic acids. However, further research is needed on the antioxidant mechanisms of the active ingredients and the synergistic effects of multiple components in Chinese herbal antioxidants to provide theoretical and data support for the use of TCM antioxidants in the treatment of male infertility.

[0007] The invention patent with publication number CN1686207A discloses a Chinese patent medicine for treating male infertility, which is composed of the following raw materials in the following weight ratio: 6-20g of ginseng, 10-20g of deer antler, 10-30g of seahorse, 6-30g of Batian, 10-30g of Schisandra chinensis, 60-200g of wolfberry, 10-20g of Ligustrum lucidum, 100-200g of Cuscuta australis, 10-20g of Rosa laevigata, 10-40g of Rubus idaeus, 10-30g of Plantago seed, 10-30g of Angelica sinensis, 10-30g of Rehmannia glutinosa, 10-30g of Cistanche deserticola, 20-100g of Epimedium, 10-30g of Cnidium monnieri, 10-30g of Allium sibiricum seed, and 10-30g of Chuanxiong fruit. This invention can improve the quality of men's sexual life, accelerate sperm formation and improve fertility, but it does not specifically alleviate TP-induced testicular oxidative stress damage and spermatogenesis impairment.

[0008] The invention patent with publication number CN105902759A discloses a traditional Chinese medicine composition for treating male and female infertility, which is composed of the following raw materials in the following weight ratios: 6-9 parts of ginseng, 30 parts of raw Astragalus, 15-30 parts of Chinese angelica, 10-15 parts of Atractylodes macrocephala, 10 parts of Poria, 20-30 parts of fried yam, 15 parts of wolfberry, 15-30 parts of Cuscuta australis, 10 parts of Morinda officinalis, 10-30 parts of Rhizoma Cyperi, 10 parts of Polygonum multiflorum, 10 parts of Dipsacus asper, 10 parts of scorched Eucommia, 10 parts of Cnidium monnieri, 3 parts of fried Artemisia argyi, 1.5 parts of cinnamon twig, 1.5 parts of fennel, 30 parts of Leonurus japonicus, 15 parts of fried white peony root, 15 parts of Cyperus rotundus, 6 parts of roasted licorice, 10 parts of antler glue, 10 parts of Schisandra chinensis, and 6 parts of raspberry. It can be used to treat male sperm defects, such as low sperm count, deformity, poor motility, azoospermia, etc., but it does not specifically alleviate TP-induced testicular oxidative stress damage and spermatogenesis impairment.

[0009] In summary, the current treatment methods for male spermatogenesis disorders still need to be further improved. Summary of the Invention

[0010] The present invention addresses the problems of the prior art and provides a Shengjing Decoction, its preparation method, application, and product. The present invention utilizes a combination of eight herbs: Danshen (Salvia miltiorrhiza), Chuanxiong (Lotus Root), Angelica sinensis (Angelica Sinensis), Epimedium (Epiphyllum Nobilis), Cnidium monnieri (Fructus Cnidium), Cuscuta chinensis (Dodder Seed), Rubus idaeus (Russica idaeus), and Schisandra chinensis (Schisandra chinensis). The decoction can improve testicular damage caused by triptolide, improve spermatogenesis, and alleviate the effects of TP on male reproductive capacity. This method provides a reference for preserving fertility during the clinical use of TP to treat inflammatory diseases.

[0011] To achieve the above object, the technical solution adopted by the present invention is as follows:

[0012] In a first aspect, the present invention provides a raw material soup comprising the following raw materials:

[0013] 5-15 parts of Salvia miltiorrhiza, 5-15 parts of Chuanxiong, 5-15 parts of Angelica sinensis, 10-25 parts of Epimedium, 10-25 parts of Cnidium monnieri, 10-25 parts of Cuscuta chinensis, 5-15 parts of Rubus idaeus and 5-15 parts of Schisandra chinensis.

[0014] Preferably, the raw material soup is composed of the following raw materials:

[0015] 7-12 parts of Salvia miltiorrhiza, 7-12 parts of Chuanxiong, 7-12 parts of Angelica sinensis, 15-20 parts of Epimedium, 15-20 parts of Cnidium monnieri, 15-20 parts of Cuscuta chinensis, 7-12 parts of Rubus idaeus and 7-12 parts of Schisandra chinensis.

[0016] More preferably, the raw material soup is composed of the following qualities:

[0017] 9 parts of Salvia miltiorrhiza, 9 parts of Chuanxiong, 9 parts of Angelica sinensis, 18 parts of Epimedium, 18 parts of Cnidium monnieri, 18 parts of Cuscuta australis, 9 parts of Rubus idaeus and 9 parts of Schisandra chinensis.

[0018] Preferably, the mass ratio of Epimedium brevicornum, Cnidium monnieri and Cuscuta chinensis is 1:1-2:1-2.

[0019] Further preferably, the mass ratio of Epimedium brevicornum, Cnidium monnieri and Cuscuta chinensis is 1:1:1.

[0020] In a second aspect, the present invention further provides a method for preparing the Shengjing Decoction described in the above technical solution, comprising:

[0021] The raw materials are mixed, added with water and boiled to obtain the sperm-producing soup.

[0022] Preferably, the raw materials are mixed according to the formula ratio, and water is added for decoction. The material-liquid ratio is preferably 100-120g:0.8-1L, and the decoction time is 0.5-1h to obtain the Shengjing Decoction. The concentration of the Shengjing Decoction is 90-100g / L.

[0023] In a third aspect, the present invention further provides a use of the Shengjing Decoction described in the above technical solution or the Shengjing Decoction prepared by the preparation method described in the above technical solution in the preparation of a drug for improving testicular damage caused by triptolide.

[0024] Preferably, the testicular damage includes oxidative damage to testicular tissue.

[0025] Preferably, the proteins related to testicular tissue oxidative damage are Nrf2 and HO-1 proteins.

[0026] Preferably, the testicular tissue oxidative damage includes testicular shrinkage, testicular germ cell vacuolation, seminiferous tubule damage, decreased sperm count, weakened sperm motility, and testicular spermatogenic cell apoptosis; the proteins related to testicular spermatogenic cell apoptosis are preferably Bcl-2 and Bax proteins.

[0027] In a fourth aspect, the present invention provides a product comprising: the Shengjing Decoction described in the above technical solution or the Shengjing Decoction prepared by the preparation method described in the above technical solution and auxiliary materials.

[0028] The pharmaceutical composition is in an oral dosage form or a non-oral dosage form.

[0029] The dosage form of the product can be any one commonly used in the art, without limitation herein, and can be tablets, capsules, powders, pills, granules, solutions, suspensions, syrups, injections, suppositories, inhalants or sprays.

[0030] The excipients are commonly used pharmaceutical excipients in the art, including but not limited to emulsifiers, solubilizers, adhesives, disintegrants, fillers, lubricants, pH regulators, stabilizers, glidants, capsule shell agents, and coating agents.

[0031] In the product, the Shengjing Decoction can be mixed with auxiliary materials to prepare different dosage forms; or the Shengjing Decoction can be concentrated and dried and then mixed with auxiliary materials to prepare different dosage forms.

[0032] The product can be used in the preparation of a medicament for restoring fertility in human males or male animals.

[0033] In a fifth aspect, the present invention provides a product comprising: the Shengjing Decoction described in the above technical solution or the Shengjing Decoction prepared by the preparation method described in the above technical solution and triptolide.

[0034] The Shengjing Decoction is used in combination with triptolide to prepare anti-tumor drugs.

[0035] The Shengjing decoction can alleviate the vacuolation of testicular germ cells induced by triptolide, which leads to the disruption of the spermatogenesis process, the shrinkage of testicles, the apoptosis of spermatogenic cells, the decrease in sperm motility parameters, the increase in the expression level of the anti-apoptotic factor Bcl-2, the decrease in the pro-apoptotic factor Bax; the increase in the expression level of the transcription factor Nrf2 and the decrease in the expression level of HO-1.

[0036] Compared with the prior art, the present invention has the following beneficial effects:

[0037] First, the Shengjing Decoction provided by the present invention is a combination of eight medicinal herbs, namely, Danshen, Chuanxiong, Angelica, Epimedium, Cnidium, Cuscuta, Rubus and Schisandra, with Epimedium, Cuscuta and Cnidium serving as monarch herbs, Rubus as ministerial herbs, Danshen, Chuanxiong and Angelica serving as adjuvant herbs, and Schisandra serving as a guiding herb. The decoction can improve testicular damage caused by the anti-inflammatory and anti-immune drug Tripterygium wilfordii hormone, improve testicular tissue oxidative damage, regulate the expression levels of anti-oxidative stress-related proteins Nrf2 and HO-1, improve phenomena such as testicular enlargement, testicular germ cell vacuolation, seminiferous tubule damage, decreased sperm count, weakened sperm motility, and testicular spermatogenic cell apoptosis, regulate the expression levels of testicular spermatogenic cell apoptosis-related proteins Bcl-2 and Bax, increase the expression level of the anti-apoptotic factor Bcl-2, and reduce the pro-apoptotic factor Bax; increase the expression level of the transcription factor Nrf2 and reduce the expression level of HO-1.

[0038] Secondly, male patients with rheumatic diseases or cancer who continuously use the drug triptolide will suffer testicular damage, impaired testicular spermatogenesis, and decreased reproductive capacity, including testicular shrinkage, destruction of germ cells in the seminiferous tubules, increased vacuolation, and significantly weakened sperm motility parameters. At the same time, testicular oxidative stress damage increases and the number of apoptotic cells increases significantly. Oral administration of the Shengjing Decoction of the present invention at the same time as the injection of triptolide can alleviate the damage of triptolide to testicular reproductive capacity, including improving oxidative stress damage, alleviating the incidence of apoptotic cells, protecting the testicles, and improving sperm quality. This provides a direction for the treatment of fertility in human men or male animals, and provides guidance or reference for patients with rheumatic diseases and tumors who want to protect their fertility during treatment. BRIEF DESCRIPTION OF THE DRAWINGS

[0039] Figure 1 The results of testicular size and sperm quality of each group of mice are shown in Figure 1, wherein A is the testicular morphology and size result of the Control group, TP group and TP+M group of mice, B is the testicular weight result of the Control group, TP group and TP+M group of mice, C is the sperm count and sperm motility result of the three groups of mice, D is the sperm morphology result of the Control group, TP group and TP+M group of mice, and E is a comparison of the testicular morphology and size of the mice in the Control group, TP group, Examples 1-3 and Comparative Examples 1-3 ( * P<0.05, **P < 0.01, *** P < 0.005, vs Control group; # P<0.05, ## P < 0.01, vs TP group);

[0040] Figure 2 Figures 2 and 3 are the testicular morphology and seminiferous tubules of mice in each group. A is the histological structure of the testicular cross-section of mice in the Control group, TP group, and TP+M group. The arrow in the TP group indicates a vacuolar structure. The scale bar is 50 μm. B is the histogram of the proportion of abnormal seminiferous tubules and the histogram of the diameter of seminiferous tubules in mice in the Control group, TP group, and TP+M group ( * P<0.05, ** P < 0.01, *** P < 0.005, vs Control group; ## P < 0.01, ### P < 0.005, vs TP group);

[0041] Figure 3 Figure 2 shows the histomorphological results of the epididymis of mice in each group. The arrows in the TP group indicate vacuolar structures. The scale bar is 50 μm. 20X represents 20 times the magnification, and 40X represents 40 times the magnification.

[0042] Figure 4 Figure 2 is the result of oxidative stress and apoptosis in the testes of male mice in each group, among which A is the result of ROS, MDA and GSH contents in the testes of mice in the Control group, TP group and TP+M group; B is the result of apoptotic cell ratio in each seminiferous tubule of mice in the Control group, TP group and TP+M group; C is the result of Bax, Bcl-2, Nrf2 and HO-1 protein expression levels in the testes of mice in the Control group, TP group and TP+M group ( * P<0.05, ** P < 0.01, *** P < 0.005, vs Control group; # P<0.05, ## P < 0.01, ### P<0.005, vs TP group). DETAILED DESCRIPTION

[0043] The present invention will be further described in detail below with reference to specific examples. The following examples are not intended to limit the present invention but are merely intended to illustrate the present invention. The experimental methods used in the following examples are generally based on conventional conditions unless otherwise specified. The materials and reagents used in the following examples are all commercially available unless otherwise specified. The following raw material sources are provided for illustrative purposes only:

[0044] Salvia miltiorrhiza: purchased from Hangzhou Huayi Pharmacy;

[0045] Chuanxiong: purchased from Hangzhou Huayi Pharmacy;

[0046] Angelica: purchased from Hangzhou Huayi Pharmacy;

[0047] Xianlingpi: purchased from Hangzhou Huayi Pharmacy;

[0048] Cnidium monnieri: purchased from Hangzhou Huayi Pharmacy;

[0049] Cuscuta seeds: purchased from Hangzhou Huayi Pharmacy;

[0050] Raspberries: purchased from Hangzhou Huayi Pharmacy;

[0051] Schisandra chinensis: purchased from Hangzhou Huayi Pharmacy.

[0052] Example 1

[0053] Shengjing Tang is composed of the following quality ingredients:

[0054] 9 parts of Salvia miltiorrhiza, 9 parts of Chuanxiong, 9 parts of Angelica sinensis, 18 parts of Epimedium, 18 parts of Cnidium monnieri, 18 parts of Cuscuta australis, 9 parts of Rubus idaeus and 9 parts of Schisandra chinensis.

[0055] Preparation method of Shengjing Decoction:

[0056] The conventional Chinese medicine boiling method is used to obtain the Chinese medicine liquid, specifically:

[0057] The raw materials were mixed according to the formula ratio, and water was added for decoction, with the material-liquid ratio being 120 g:1 L. The decoction was decocted for 1 hour to obtain the Shengjing decoction, and the concentration of the Shengjing decoction was 95 g / L.

[0058] Example 2

[0059] Shengjing Tang is composed of the following quality ingredients:

[0060] 5 parts of Salvia miltiorrhiza, 15 parts of Chuanxiong, 5 parts of Angelica sinensis, 10 parts of Epimedium, 25 parts of Cnidium monnieri, 10 parts of Cuscuta seeds, 15 parts of Rubus idaeus and 5 parts of Schisandra chinensis.

[0061] Shengjing decoction was prepared according to the method of Example 1, and the concentration of Shengjing decoction was 90 g / L.

[0062] Example 3

[0063] Shengjing Tang is composed of the following quality ingredients:

[0064] 15 parts of Salvia miltiorrhiza, 5 parts of Chuanxiong, 15 parts of Angelica sinensis, 25 parts of Epimedium, 10 parts of Cnidium monnieri, 25 parts of Cuscuta australis, 5 parts of Rubus idaeus and 15 parts of Schisandra chinensis.

[0065] Shengjing decoction was prepared according to the method of Example 1, and the concentration of Shengjing decoction was 100 g / L.

[0066] Comparative Example 1

[0067] The difference from Example 1 is that 18 parts of Xianlingpi are replaced by 18 parts of Cnidium monnieri, i.e., Shengjing Decoction, which is composed of the following raw materials by weight:

[0068] 9 parts of Salvia miltiorrhiza, 9 parts of Chuanxiong, 9 parts of Angelica sinensis, 36 parts of Cnidium monnieri, 18 parts of Cuscuta seeds, 9 parts of Rubus idaeus and 9 parts of Schisandra chinensis.

[0069] The conventional Chinese medicine boiling method is used to obtain the Chinese medicine liquid, specifically:

[0070] The raw materials were mixed according to the formula ratio, and water was added for decoction, with the material-liquid ratio being 120 g:1 L. The decoction was decocted for 1 hour to obtain the Shengjing decoction, and the concentration of the Shengjing decoction was 95 g / L.

[0071] The rest is the same as Example 1.

[0072] Comparative Example 2

[0073] The difference from Example 1 is that 18 parts of Xianlingpi are replaced by 18 parts of leek seeds, i.e., Shengjing Decoction, which is composed of the following raw materials by weight:

[0074] 9 parts of Salvia miltiorrhiza, 9 parts of Chuanxiong, 9 parts of Angelica sinensis, 18 parts of Allium tuberosum seeds, 18 parts of Cnidium monnieri, 18 parts of Cuscuta seeds, 9 parts of Rubus idaeus and 9 parts of Schisandra chinensis.

[0075] The raw materials were mixed according to the formula ratio, and water was added for decoction, with the material-liquid ratio being 120 g:1 L. The decoction was decocted for 1 hour to obtain the Shengjing decoction, and the concentration of the Shengjing decoction was 95 g / L.

[0076] The rest is the same as Example 1.

[0077] Comparative Example 3

[0078] The difference from Example 1 is that the mass ratio of Herba Epimedii, Fructus Cnidii and Fructus Cuscutae is changed to 3:1:1, i.e., Shengjing Decoction, which is composed of the following raw materials by weight:

[0079] 9 parts of Salvia miltiorrhiza, 9 parts of Chuanxiong, 9 parts of Angelica sinensis, 33 parts of Epimedium, 11 parts of Cnidium monnieri, 11 parts of Cuscuta australis, 9 parts of Rubus idaeus and 9 parts of Schisandra chinensis.

[0080] The raw materials were mixed according to the formula ratio, and water was added for decoction, with the material-liquid ratio being 120 g:1 L. The decoction was decocted for 1 hour to obtain the Shengjing decoction, and the concentration of the Shengjing decoction was 95 g / L.

[0081] The rest is the same as Example 1.

[0082] Test example

[0083] Effect of Shengjing Decoction on Testicular Damage Induced by Triptolide

[0084] 1. Experimental methods

[0085] 1.1 Experimental animals and drugs

[0086] Eight-week-old ICR male mice were provided by the Animal Experimentation Center of Sir Run Run Shaw Hospital, Zhejiang University. They were provided with free access to food and water, and maintained under controlled light conditions (12 h / day). The room temperature was maintained at 20–22°C. All experimental animals and related animal manipulations followed standard operating procedures and were reviewed by the Animal Ethics Center of Sir Run Run Shaw Hospital, Zhejiang University.

[0087] Take 10 g of the Shengjing Decoction prepared in Example 1-3 and Comparative Example 1-3, add 20 mL of normal saline to dissolve it, and use it for subsequent gavage treatment of mice at a rate of 20 μL / g.

[0088] 1.2 Administration

[0089] ICR male mice were randomly divided into 8 groups, with 8 mice in each group; the control group was injected with normal saline; the TP group was injected with 150 μ / kg of triptolide intraperitoneally; the TP group was injected with 150 μ / kg of triptolide intraperitoneally and gavaged with 20 μL / g of the Shengjing Decoction prepared in Example 1, which was recorded as the TP+M group. After continuous TP administration for 14 days, the administration was stopped, and the Chinese medicine gavage was continued for 28 consecutive days starting from the third day of administration, while adaptive feeding was performed at the same time. After 30 days, the mice were euthanized and the corresponding samples were collected; the intraperitoneal injection and gavage time and concentration of Example 2, Example 3 and Comparative Examples 1, 2, and 3 were consistent with those of the Example 1 group, and the testicles were collected for comparison of morphology and size and subsequent fertility experimental testing.

[0090] 1.3 Testicular size and sperm quality testing

[0091] The mice were euthanized, and the testicular and epididymal tissues were collected. The testicular tissues of each group were photographed and compared in size and weight.

[0092] Epididymal caudalis tissue was collected and placed in a preheated 37°C benchtop solution (purchased from Beijing Solebeau Technology Co., Ltd., product number T1420). Two to three small cuts were made and the tissue was placed in a 37°C incubator for 15 minutes. Twenty microliters of the epididymal mixture was added to 1 mL of benchtop solution and observed microscopically. Photos were taken and recorded. Computer-assisted sperm analysis (CASA) was used to measure sperm count, motility, and morphology.

[0093] 1.4 Examination of testicular and epididymal tissue morphology

[0094] Testicular and epididymal tissues of mice in each group were collected and immersed in 4% paraformaldehyde at 4°C for 48 hours. The tissues were then immersed in 50% ethanol at 4°C for 24 hours, 75% ethanol at 4°C for 24 hours, 80% ethanol at room temperature for 2 hours, 90% ethanol for 2 hours, 95% ethanol for 2 hours, 100% ethanol for 2 hours (repeated twice), and xylene for 2 hours (repeated twice). The tissues were then immersed in paraffin, embedded in paraffin, sectioned at 5 microns, and placed in a 60°C oven for 5 hours.

[0095] Paraffin sections of testicular and epididymal tissue from mice in different groups were randomly selected and dewaxed in xylene for 15 minutes twice. Sections were then treated in gradient ethanol (100%, 95%, 90%, 80%, and 70%) for 5 minutes, followed by treatment in distilled water for 2 minutes. Sections were stained with commercially available hematoxylin for 8 minutes, rinsed with running water for blueing, and then eosin was added to the epididymal tissue for 30 seconds. Sections were then treated with 80%, 90%, and 100% alcohol for 2 minutes. Finally, sections were cleared in xylene for 5 minutes and mounted with neutral gum. Sections of testicular and epididymal tissue from different groups were observed and photographed using a Leica microscope.

[0096] 1.5 Research on the intrinsic mechanism of male testicular spermatogenesis

[0097] Reactive oxygen species (ROS), glutathione (GSH), and malondialdehyde (MDA) are important markers of oxidative stress in vivo. We assessed testicular oxidative stress by measuring their levels in mouse testicular tissue. Following the manufacturer's instructions for the test kits, testicular homogenates were obtained from each group of mice and assayed for various oxidative stress markers.

[0098] To further understand the regulatory mechanism of triptolide and Shengjing Decoction on testicular spermatogenesis in mice, we dehydrated the paraffin sections of each group obtained in 1.4 and used the TUNEL apoptosis kit to detect the proportion of apoptotic cells in the testis.

[0099] 1.6 Fertility test of each group of mice

[0100] The fertility of each group of mice was tested by the mouse cage experiment, specifically:

[0101] Three male mice were randomly selected from each group, and each male mouse was caged with a 10-week-old ICR female mouse. The fertility of the female mice was counted and the fertility rate was calculated.

[0102] Fertility rate (%) = number of pups born in the experimental group / number of pups born in the control group

[0103] 2. Test results

[0104] 2.1 Testicular size and sperm quality test results

[0105] The results are as follows Figure 1 As shown, Figure 1 The figures are as follows: Figure A shows the testicular morphology and size of mice in the Control group, TP group, and TP+M group; Figure B shows the testicular weight of mice in the Control group, TP group, and TP+M group; Figure C shows the sperm count and sperm motility of the three groups of mice; Figure D shows the sperm morphology of mice in the Control group, TP group, and TP+M group; and Figure E shows a comparison of the testicular morphology and size of mice in the Control group, TP group, Examples 1-3, and Comparative Examples 1-3.

[0106] The results showed that the testicle size of mice injected with triptolide (TP group) was significantly smaller than that of the control group ( *** P<0.005). Compared with the TP group, the testicles of the TP+M group were significantly larger ( ## P < 0.01), indicating that the simultaneous administration of Shengjing Decoction (TP+M group) by gavage with triptolide can significantly alleviate the effect of triptolide on testicular size. After triptolide treatment, the number of sperm in the epididymis of mice was significantly reduced (TP group); compared with the TP group, the number of sperm was significantly increased after the simultaneous administration of Shengjing Decoction (TP+M group) by gavage with triptolide. After CASA system analysis, there was a significant difference in sperm count between the TP group and the TP+M group ( *** P<0.005, ## P<0.01); sperm motility analysis showed that after triptolide treatment of mice, sperm motility was significantly weakened ( *** P < 0.005,), while the sperm motility was significantly restored by gavage of Shengjing Decoction of Example 1 (TP+M group) while injecting triptolide ( ## P < 0.01). Compared with the TP group, the size of experimental examples 2 and 3 was significantly increased, while there was no significant difference between comparative examples 1, 2, and 3 and the TP group.

[0107] 2.2 Testicular and epididymal histomorphological examination results

[0108] The results are as follows Figure 2 and Figure 3 As shown, Figure 2 Figures 2 and 3 show the testicular morphology and seminiferous tubules of mice in the Control group, TP group, and TP+M group. Figure A shows the histomorphological structure of the testicular cross-sections of mice in the Control group, TP group, and TP+M group. Figure B shows the bar graph of the proportion of abnormal seminiferous tubules and the bar graph of the diameter of seminiferous tubules of mice in the Control group, TP group, and TP+M group. Figure 3 Figure 2 shows the histomorphological results of epididymis in mice in the Control group, TP group, and TP+M group.

[0109] like Figure 2 and Figure 3 The results showed that after treatment with triptolide, the testicular tissue morphology showed obvious vacuolar structures and spermatogenic cells were obviously absent ( Figure 2 The number of sperm in the epididymal duct was significantly reduced ( Figure 3 TP group in Figure A); after intraperitoneal treatment with triptolide and oral administration of Shengjing Decoction in Example 1, the structure of the seminiferous tubules of the testis was significantly restored, and spermatogenic cells at all levels showed normal morphology ( Figure 2 (TP+M group in Figure A); after intraperitoneal treatment with triptolide and oral administration of Shengjing decoction in control example 1, the seminiferous tubules showed multiple vacuoles, which had no significant difference from the TP group. Figure 3 Figure B).

[0110] Statistics of abnormal seminiferous tubules and diameter of seminiferous tubules in testis Figure 2 The analysis of Figure B) showed that there was a significant difference after treatment with triptolide ( *** P < 0.005), after intraperitoneal treatment with triptolide and intragastric administration of Shengjing Decoction of Example 1, abnormal seminiferous tubules were significantly reduced, and the diameter of the seminiferous tubules increased ( ### P<0.005).

[0111] 2.3 Intrinsic Mechanism of Male Testicular Spermatogenesis

[0112] The results are as follows Figure 4 As shown, Figure A shows the ROS, MDA and GSH contents in the testis of mice in the blank group, TP group and TP+M group; Figure B shows the proportion of apoptotic cells in each seminiferous tubule of mice in the blank group, TP group and TP+M group; and Figure C shows the expression levels of Bax, Bcl-2, Nrf2 and HO-1 proteins in the testis of mice in the blank group, TP group and TP+M group.

[0113] like Figure 4 As shown in Figure A, compared with the control group, after treatment with triptolide, the ROS content in the testis ( ** P<0.01) and MDA( **P<0.01) were significantly increased, but after the Shengjing Decoction of Example 1 was administered orally, the ROS content in the testis ( # P<0.05) and MDA( # P<0.05) levels were all down-regulated; similarly, after treatment with triptolide, the oxidative stress index GSH in the TP group was significantly decreased compared with the control group ( *** P < 0.005), when the Shengjing decoction of Example 1 was administered orally, the decreased GSH content was significantly increased ( ### P<0.005), indicating that the oxidative damage in testicular tissue was alleviated to a certain extent.

[0114] like Figure 4 As shown in middle B, compared with the control group, in each seminiferous tubule, triptolide caused a significant increase in the proportion of apoptotic cells in the testis ( *** P < 0.005), but after the simultaneous oral administration of Shengjing Decoction in Example 1, the proportion of apoptotic cells in each seminiferous tubule was significantly decreased ( ### P < 0.005, TP+M group), which shows that the Shengjing decoction of Example 1 can alleviate the cell damage caused by triptolide to a certain extent.

[0115] like Figure 4 As shown in Figure C, at the protein level, we found that compared with the control group, after the mice were treated with triptolide, the expression of Bcl-2 protein in the testicular tissue was significantly reduced, while the expression of Bax protein was significantly increased. The group treated with Shengjing Decoction of Example 1 (TP+M group) showed a significant increase in the expression of Bcl-2 protein in the testicular tissue ( ### P<0.005), while the expression of Bax protein was significantly decreased ( ## P < 0.01), indicating that the Shengjing Decoction of Example 1 has a certain inhibitory effect on cell apoptosis. At the same time, after treatment with triptolide, the expression of Nrf2 protein in the testicular tissue of mice was significantly reduced, while the expression of HO-1 protein was significantly increased. The Shengjing Decoction of Example 1 significantly increased the expression level of Nrf2 protein and significantly reduced the expression level of HO-1 protein ( ## P < 0.01), indicating that Shengjing Decoction in Example 1 has a certain antioxidant effect. Shengjing Decoction in Example 1 inhibits cell apoptosis by downregulating the expression of the pro-apoptotic protein Bax and upregulating the expression of the apoptosis-inhibiting protein Bcl-2. It also increases the expression of the antioxidant protein Nrf2 and reduces the expression level of HO-1, thereby resisting oxidative damage.

[0116] 2.4 Fertility test of mice in each group

[0117] The fertility results of mice in each group are shown in Table 1.

[0118] The results showed that after treatment with triptolide, the pregnancy rate of female mice was 0%. When the Shengjing Decoction of Example 1 was simultaneously administered orally, the fertility rate of female mice reached 60% compared with normal mice in the cage experiment. The fertility rates of Examples 2 and 3 were 58% and 62%, respectively, which were not significantly different from those of the Examples. The fertility rates of Comparative Examples 1-3 were all less than 10%, indicating that Shengjing Decoction can rescue the decreased fertility caused by triptolide.

[0119] Table 1 Fertility rate of mice in each group

[0120] Grouping fertility rate Control group 100％ Triptolide group (TP) 0％ Example 1 60％ Example 2 58％ Example 3 62％ Comparative Example 1 8％ Comparative Example 2 10％ Comparative Example 3 8％

[0121] Comprehensive analysis of the experimental results shows that the Shengjing Decoction of the present invention can protect testicular tissue by alleviating the oxidative stress damage of triptolide to testicular tissue.

[0122] Finally, it should be noted that the above content is only used to illustrate the technical solution of the present invention, rather than to limit the scope of protection of the present invention. Simple modifications or equivalent substitutions of the technical solution of the present invention by ordinary technicians in this field do not deviate from the essence and scope of the technical solution of the present invention.

Claims

1. A Shengjing Decoction, characterized in that: By weight, it is composed of the following raw materials: 5-15 parts of Salvia miltiorrhiza, 5-15 parts of Chuanxiong, 5-15 parts of Angelica sinensis, 10-25 parts of Epimedium, 10-25 parts of Cnidium monnieri, 10-25 parts of Cuscuta chinensis, 5-15 parts of Rubus idaeus and 5-15 parts of Schisandra chinensis.

2. The Shengjing Decoction according to claim 1, characterized in that: By weight, it is composed of the following raw materials: 9 parts of Salvia miltiorrhiza, 9 parts of Chuanxiong, 9 parts of Angelica sinensis, 18 parts of Epimedium, 18 parts of Cnidium monnieri, 18 parts of Cuscuta australis, 9 parts of Rubus idaeus and 9 parts of Schisandra chinensis.

3. The Shengjing Decoction according to claim 1, characterized in that The mass ratio of the herb Epimedium, Cnidium monnieri and Cuscuta australis is 1:1-2:1-2.

4. The method for preparing the Shengjing Decoction according to any one of claims 1 to 3, characterized in that: include: The raw materials are mixed, added with water and boiled to obtain the sperm-producing soup.

5. The preparation method according to claim 4, characterized in that: The decoction time is 0.5-1h; the concentration of the Shengjing decoction is 90-100g / L.

6. Use of the Shengjing Decoction according to any one of claims 1 to 3 or the Shengjing Decoction prepared by the preparation method according to any one of claims 4 to 5 in the preparation of a medicament for improving testicular damage caused by triptolide.

7. The use according to claim 6, characterized in that The dosage form of the medicine is pills, powders, granules or capsules.

8. The application according to claim 6, characterized in that: The testicular damage includes oxidative damage to testicular tissue.

9. The application according to claim 8, characterized in that: The proteins related to testicular tissue oxidative damage are Nrf2 and HO-1 proteins; testicular tissue oxidative damage includes testicular enlargement, testicular germ cell vacuolation, seminiferous tubule damage, decreased sperm count, weakened sperm motility and / or testicular spermatogenic cell apoptosis; the proteins related to testicular spermatogenic cell apoptosis are Bcl-2 and Bax proteins.

10. A product, characterized in that include: The Shengjing decoction according to any one of claims 1-3 or the Shengjing decoction prepared by the preparation method according to any one of claims 4-5.

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