A pharmaceutical composition containing cistanche deserticola components and its uses

Abstract

The present invention provides a pharmaceutical composition for preventing and treating male infertility. The pharmaceutical composition contains acteoside and echinacoside, wherein the concentration ratio of acteoside to echinacoside is 1 to 4 or 1 / 3. The pharmaceutical composition has the effects of improving male reproductive damage and protecting spermatocytes from the toxicity of environmental estrogen BPA.

Description

Technical Field

[0001] The present invention relates to the field of pharmaceutical technology, and particularly relates to a pharmaceutical composition containing cistanche deserticola components and its uses. Background Art

[0002] Cistanche deserticola is the dried fleshy stem with scales of the plant Cistanche deserticola in the Orobanchaceae family. It has a long medicinal history and is mainly produced in Inner Mongolia, Xinjiang and other places in China. It is a unique sandy endangered medicinal material in the arid regions of northwest China and is known as the "ginseng in the desert". Its dried fleshy stem was first recorded in "Shennong Ben Cao Jing" for medicinal use. The medicinal material of cistanche deserticola is sweet, salty and warm in nature. Modern research shows that the active components of cistanche deserticola are mainly phenyl ethanol glycosides, polysaccharides (oligosaccharides) and iridoids. Among them, phenyl ethanol glycoside components have various effects such as antibacterial, anti-inflammatory, hepatoprotective, antioxidant, immune regulation, memory enhancement and anti-aging, and are the characteristic active components of cistanche plants. There are many phenyl ethanol glycoside components isolated from cistanche plants, reaching dozens of them. Typical ones are acteoside and verbascoside, and these two components are used as one of the quality control indicators for cistanche deserticola in the pharmacopoeia.

[0003] In recent years, the research, development and utilization of cistanche deserticola have attracted extensive attention from traditional Chinese medicine researchers and have become one of the research hotspots in the field of traditional Chinese medicine. The research contents are mostly about the isolation and identification of its chemical components, the identification of active components of cistanche deserticola of different varieties in different regions, the research and determination of the quality standards of traditional Chinese medicinal materials, and the pharmacological effects of total glycoside active components, etc. In vivo experiments mostly use pathological models induced by chemical drugs or toxic traditional Chinese medicine components to study the protective effects of cistanche deserticola on corresponding organs and diseases; there is no research on the effect on the key link in the process of sperm development - the proliferation and differentiation of spermatocytes. At the same time, the pharmacological research on verbascoside and acteoside only stays at the determination of quality standards, and there is no conclusion on their action ratio and concentration.

[0004] Bisphenol A (BPA) is a type of environmental estrogen that people often come into contact with in production and life. It is often used as the packaging material for food and beverages, the inner lining of metal cans, dental sealants and additives for other products. It can enter the organism through the food chain and interact with estrogen receptors (ER), interfering with the production, release, transportation, metabolism and other links of normal hormones in the organism, thus affecting the reproductive, nervous, immune and other functions of organisms and having strong "carcinogenic, teratogenic and mutagenic" effects on organisms.

[0005] In this experiment, bisphenol A was selected as the in vitro modeling agent to construct a spermatocyte damage model, explore the protective effects and concentration ratios of phenyl ethanol glycoside active components verbascoside and acteoside of cistanche deserticola on spermatocytes, improve its treatment plan, and provide a traditional Chinese medicine solution for solving the currently common and intractable global fertility problem. Summary of the Invention

[0006] Based on this, the present invention provides a pharmaceutical composition for preventing and treating male infertility, which comprises acteoside and cichoric acid, wherein the concentration ratio of acteoside to cichoric acid is 1 to 4 or 1 / 3.

[0007] Further, the concentration ratio of acteoside to cichoric acid is about 1.

[0008] Further, the concentration is molar concentration, equivalent concentration, mass-volume concentration or mass percentage concentration.

[0009] Further, the pharmaceutical composition further comprises one or more drugs and / or extracts for preventing and treating male infertility.

[0010] Further, the drug is selected from one or more of the following: follicle-stimulating hormone, clomiphene, letrozole, imipramine and levocarnitine.

[0011] Another aspect of the present invention provides a pharmaceutical preparation comprising the above pharmaceutical composition and a pharmaceutically acceptable excipient.

[0012] Further, the excipient is selected from one or more of the following: inclusion agent, flavoring agent, sustained-release agent, diluent, lubricant, disintegrant, suspending agent, light-blocking agent, dispersant, plasticizer, wetting agent, binder and antioxidant.

[0013] Further, the dosage form of the pharmaceutical preparation is an oral preparation, capsule, powder, sustained-release preparation, injection, granule, pill, controlled-release preparation or targeted preparation.

[0014] Another aspect of the present invention provides the use of the above pharmaceutical composition or the above pharmaceutical preparation in the preparation of a drug for preventing and treating male infertility.

[0015] Further, the male infertility is male infertility with decreased spermatocyte viability.

[0016] Further, the male infertility is oligoasthenospermia.

[0017] Further, the male infertility is drug-induced male infertility.

[0018] Further, the drug is an environmental estrogen such as bisphenol A.

[0019] Further, the concentration of bisphenol A is ≥160 μM.

[0020] Advantages of the present invention:

[0021] In this invention, by administering acteoside and cichoric acid at different concentrations to mouse GC-2 spermatocytes, it is found that both can promote the proliferation of spermatocytes in normal growth environments and in environments poisoned by bisphenol A (BPA). Moreover, when the ratio of the two is 1:1, the effect is the best, suggesting that the active components of cistanche phenyl ethanol glycosides have the effect of improving male reproductive damage and protecting spermatocytes from the toxicity of environmental estrogen BPA. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] In order to more clearly illustrate the technical solutions in the embodiments of the present invention, the following will briefly introduce the drawings required for the description of the embodiments. Obviously, the drawings in the following description are only some embodiments of the present invention. For those of ordinary skill in the art, other drawings can also be obtained based on these drawings without exceeding the scope of protection required by the present invention.

[0023] Figure 1 It is a schematic diagram of the results of the promoting proliferation effect of acteoside and cichoric acid on spermatocytes.

[0024] Among them, compared with the normal group, * P < 0.05, ** P < 0.01, *** P < 0.001.

[0025] Figure 2 It is a schematic diagram of the results of the effect of bisphenol A (BPA) on the activity of spermatocytes. Among them, compared with the normal group, * P < 0.05, *** P < 0.001.

[0026] Figure 3 It is a schematic diagram of the results of the effect of acteoside and cichoric acid on the proliferation activity of spermatocytes in a BPA-toxic environment. Among them, compared with the 180 μM BPA group, * P < 0.05, *** P < 0.001.

[0027] Figure 4 It is a schematic diagram of the results of the effect of the combined application of acteoside and cichoric acid at a ratio of 1:1 on the proliferation activity of spermatocytes in a BPA-toxic environment. Among them, compared with the 180 μM BPA group, *** P < 0.001. For pairwise comparisons, # P < 0.05.

[0028] Figure 5 It is a schematic diagram of the results of the effect of the combined application of acteoside and cichoric acid at different ratios on the proliferation activity of spermatocytes in a BPA-toxic environment. Among them, compared with the 180 μM BPA group, * P < 0.05, ** P < 0.01,*** P < 0.001. When comparing two groups pairwise, # P < 0.05, ## P < 0.01. Detailed implementation manners

[0029] Next, the technical solutions in the embodiments of the present invention will be clearly and completely described in conjunction with the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, rather than all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative efforts fall within the scope of protection of the present invention.

[0030] Unless otherwise stated, all technical and scientific terms and abbreviations used herein have the meanings commonly understood by those of ordinary skill in the art in the field of the present invention or the field where the term is applied. Although any ratios, temperatures, substances or materials similar to or equivalent to those disclosed herein may be used in the implementation process of the present invention, the preferred ratios, temperatures, substances or materials are described herein. The present invention includes multiple aspects, features and implementation manners, and such multiple aspects, features and implementation manners can be combined and modified in any desired manner. These and other aspects, features and implementation manners of the present invention become obvious when referring to the rest of the present invention including the following detailed description. Unless the context clearly indicates, as used herein, the singular form of a word includes the plural, and vice versa. Therefore, "a", "an" and "the" generally include the plural of the corresponding terms. Similarly, the term "for example", especially when placed in front of a series of terms, is only illustrative and explanatory and should not be regarded as exclusive or comprehensive.

[0031] In the present invention, the terms "comprising" and "including" are synonymous. The terms "including", "comprising", "having", "containing" or any other modification thereof used herein are intended to cover non-exclusive inclusion. For example, a composition, drug or pharmaceutical excipient containing the listed elements does not have to be limited to those elements listed in the present invention, but may include other elements not explicitly listed in the composition, drug or pharmaceutical excipient. Similarly, the term "for example", especially when placed in front of a series of terms, is only illustrative and explanatory and should not be regarded as exclusive or comprehensive.

[0032] As described in the background art section, there are no relevant reports on the effects of acteoside and cichoric acid on the key link in the process of sperm development - the proliferation and differentiation of spermatocytes. To solve the above problems, the present invention provides a pharmaceutical composition for preventing and treating male infertility, which comprises acteoside and cichoric acid, and the concentration ratio of the acteoside to the cichoric acid is 1 to 4 or 1 / 3.

[0033] In the present invention, when a concentration ratio, molar ratio, content, concentration, time, or other value or parameter is expressed as a range, a preferred range, or a range defined by a series of upper preferred values and lower preferred values, it should be understood that all ranges formed by any pairing of any range upper limit or preferred value with any range lower limit or preferred value are specifically disclosed, regardless of whether the range is disclosed individually. For example, when the range "1 to 4" is disclosed, the described range should be interpreted as including ranges "1 to 2", "1 to 3", "1 to 4", "2 to 3", "2 to 4", and "3 to 4", etc. When a numerical range is described herein, unless otherwise specified, the range is intended to include its end values and all integers, fractions, and numerical values with 1 to 2 decimal places within the range, such as 1, 1.05, 1.1, 1.15, 1.2, 1.25, 1.3, 1.35, 1.4, 1.45, 1.5, 1.55, 1.6, 1.65, 1.7, 1.75, 1.8, 1.85, 1.9, 1.95, 2, 2.05, 2.1, 2.15, 2.2, 2.25, 2.3, 2.35, 2.4, 2.45, 2.5, 2.55, 2.6, 2.65, 2.7, 2.75, 2.8, 2.85, 2.9, 2.95, 3, 3.05, 3.1, 3.15, 3.2, 3.25, 3.3, 3.35, 3.4, 3.45, 3.5, 3.55, 3.6, 3.65, 3.7, 3.75, 3.8, 3.85, 3.9, 3.95, and 4, and within the above range, the technical effects of the present invention can be achieved.

[0034] In the present invention, the term "prevention and treatment" includes "treatment" and / or "prevention", and the terms "treatment (of)" and "treatment (ly)" should be understood accordingly. Specifically, the term "treatment" includes alleviating, inhibiting, or improving the symptoms or conditions of a disease; inhibiting the occurrence of complications; improving or preventing potential metabolic syndrome; inhibiting the occurrence of a disease or symptoms, such as controlling the development of a disease or condition; alleviating a disease or symptoms; reducing a disease or symptoms; causing a disease or symptoms to subside; alleviating the complications caused by a disease or symptoms, or preventing or treating the signs caused by a disease or symptoms. As used herein, after administration of a pharmaceutical composition or a pharmaceutical preparation, a certain disease, symptom, or condition can be improved, especially the severity thereof can be improved, the onset can be delayed, the progression of the disease can be slowed down, or the duration of the disease can be reduced. Whether administered regularly or temporarily, continuously or intermittently, it can be attributed to or related to the situation of administration.

[0035] In a preferred embodiment, the concentration ratio of verbascoside to echinacoside is about 1.

[0036] In the present invention, "about" refers to a value within the range of ±5% of a specific value. For example, "about 1" includes ±5% of 1, or from 0.95 to 1.05.

[0037] In a preferred embodiment, the concentration ratio of verbascoside to echinacoside is 0.95 to 1.05.

[0038] In a preferred embodiment, the concentration is molar concentration, normal concentration, mass - volume concentration or mass percentage concentration.

[0039] Among them, the concentration of a solution expressed by the number of moles of solute contained in 1 liter of the solution is called molar concentration, represented by the symbol mol; the concentration of a solution expressed by the number of gram equivalents of solute contained in 1 liter of the solution is called normal concentration, represented by the symbol N; the concentration expressed by the mass number of solute contained in a unit volume (1 cubic meter or 1 liter) of the solution is called mass - volume concentration, represented by the symbol g / m 3 or mg / L; the concentration of a solution expressed by the percentage of the mass of the solute in the total mass of the solution is called mass percentage concentration, represented by the symbol %.

[0040] In a preferred embodiment, the concentration is molar concentration.

[0041] In the actual production and application process, for the pharmaceutical compositions obtained with different normal concentration ratios, mass - volume concentration ratios or mass percentage concentration ratios between verbascoside and echinacoside, as long as the molar concentration ratio between the two obtained through conversion falls within the above - mentioned molar concentration ratio range of the present invention, they all fall within the protection scope of the present invention.

[0042] In a preferred embodiment, the pharmaceutical composition further comprises one or more drugs and / or extracts for preventing and treating male infertility.

[0043] In a preferred embodiment, the drug is selected from one or more of the following: follicle - stimulating hormone, clomiphene, letrozole, imipramine and levocarnitine.

[0044] Another aspect of the present invention provides a pharmaceutical preparation comprising the above - mentioned pharmaceutical composition and a pharmaceutically acceptable excipient.

[0045] In a preferred embodiment, the excipient is selected from one or more of the following: inclusion agent, flavoring agent, sustained - release agent, diluent, lubricant, disintegrant, suspending agent, light - shielding agent, dispersant, plasticizer, wetting agent, binder and antioxidant.

[0046] In a preferred embodiment, the dosage form of the pharmaceutical preparation is an oral preparation, capsule, powder, sustained - release preparation, injection, granule, pill, controlled - release preparation or targeted preparation.

[0047] The above-mentioned drugs, excipients and dosage forms of the present invention are only illustrative, and are not limited to those listed above during actual application. As long as they can be used in combination with the pharmaceutical composition of the present invention, they are all within the protection scope of the present invention.

[0048] Another aspect of the present invention provides the use of the above-mentioned pharmaceutical composition or the above-mentioned pharmaceutical preparation in the preparation of a drug for preventing and treating male infertility.

[0049] According to another aspect of the present invention, there is provided the above-mentioned pharmaceutical composition or the above-mentioned pharmaceutical preparation for preventing and treating male infertility in a subject.

[0050] According to another aspect of the present invention, there is provided a method for preventing and treating male infertility in a subject, comprising administering an effective amount of the above-mentioned pharmaceutical composition or the above-mentioned pharmaceutical preparation to the subject.

[0051] In a preferred embodiment, the male infertility is male infertility with decreased spermatocyte viability.

[0052] In a preferred embodiment, the male infertility is oligoasthenospermia.

[0053] In a preferred embodiment, the male infertility is drug-induced male infertility.

[0054] In a preferred embodiment, the drug is bisphenol A.

[0055] In a preferred embodiment, the drug is an environmental estrogen such as bisphenol A.

[0056] In a preferred embodiment, the concentration of bisphenol A ≥ 160 μM.

[0057] Within the above concentration range of bisphenol A, bisphenol A exhibits a toxic effect of inhibiting the proliferation of spermatocytes.

[0058] In the present invention, the term "subject" refers to any animal that can benefit from the above-mentioned pharmaceutical composition or the above-mentioned pharmaceutical preparation of the present invention, such as a human, bird, cow, canine, horse, feline, goat, wolf, rodent, sheep or swine, but is not limited to these examples. Mammals other than humans can be advantageously used as subjects representing the above-mentioned disease models. Preferably, the subject is a human.

[0059] In the present invention, the term "effective amount" means the amount of the pharmaceutical composition or pharmaceutical preparation of the present invention that achieves the following purposes: ① treating or preventing a specific disease, disorder or condition, ② attenuating, alleviating or eliminating one or more symptoms of the specific disease, disorder or condition, or ③ preventing or delaying the onset of one or more symptoms of the specific disease, disorder or condition described herein. The amount effective for this purpose will depend on, for example, the mode of administration, the stage and severity of the disease being treated, the individual body weight and overall health status, and the judgment of the prescribing physician. The dose can be administered once a week, or once every two days or every day, or even several times a day. The dose unit can be administered over a short period (e.g., several weeks to several months) or a longer period (several months to several years).

[0060] It should be noted that, without conflict, the embodiments in the present invention and the features in the embodiments can be combined with each other. The present invention will be described in detail below in conjunction with the embodiments.

[0061] The following further describes the present invention in detail with reference to specific embodiments, and these embodiments should not be construed as limiting the scope of the present invention claimed.

[0062] Example 1

[0063] Proliferation-promoting effect of acteoside and echinacoside on spermatocytes

[0064] 1 Materials

[0065] 1.1 Cells

[0066] GC-2 mouse spermatocytes (purchased from ATCC).

[0067] 1.2 Drugs and reagents

[0068] Fetal bovine serum, penicillin / streptomycin, DMEM medium, 0.25% trypsin, acteoside, echinacoside, MTT.

[0069] 2 Methods

[0070] 2.1 Cell culture

[0071] GC-2 cells were cultured in DMEM medium containing 10% fetal bovine serum and 1% penicillin / streptomycin in an incubator at 37°C with 5% CO2 and saturated humidity. The culture medium was changed every other day, and the cell growth status was observed under an inverted microscope. When the cells grew and fused into a dense monolayer (coverage rate reached 70%-80%), they were routinely digested and passaged with 0.25% trypsin, and passaged at a ratio of 1:5 or 1:6 every 2-3 days.

[0072] 2.2 MTT colorimetric assay

[0073] In this experiment, different concentrations of acteoside and cistanoside A (100 nmol / L, 1 μmol / L, 10 μmol / L, 100 μmol / L) were used to intervene in GC-2 cells for 24 h to observe the cell proliferation effect.

[0074] The specific steps are as follows:

[0075] (1) Collect GC-2 cells in the logarithmic growth phase, prepare a single-cell suspension with DMEM medium containing 10% fetal bovine serum, and inoculate it into a 96-well plate at a cell density of 8000 cells / well, with a volume of 180 μL per well;

[0076] (2) After the cells adhered, add 20 μL of acteoside and cistanoside A with corresponding 10-fold concentrations to each group, and set 6 replicate wells in each group. The final volume in each well is the same, all 200 μL;

[0077] (3) After 24 h of drug intervention, add 20 μL of MTT (5 mg / mL) to each well, continue to incubate in a 5% CO2, 37 °C cell culture incubator for 4 h, terminate the culture, aspirate the liquid in the wells, add 150 μL of DMSO to each well, and oscillate for 3 min to fully dissolve the crystals;

[0078] (4) Colorimetry: Use an enzyme-linked immunosorbent assay (ELISA) reader to measure the absorbance value (OD value) of each well at a wavelength of 490 nm, statistically analyze the optical density values of each group, and draw a cell growth trend graph with time as the abscissa and OD value as the ordinate.

[0079] 3 Results

[0080] As Figure 1 shown, 10 μM acteoside and 1 μM cistanoside A can significantly promote the proliferation of spermatocytes (P < 0.001, 0.05), indicating that the active components of cistanche phenylethanoid glycosides have a promoting effect on the growth of normal spermatocytes and a positive impact on the male reproductive system.

[0081] Example 2

[0082] Effect of acteoside and cistanoside A on the proliferation ability of spermatocytes poisoned by bisphenol A (BPA)

[0083] 1 Materials

[0084] Same as Example 1.

[0085] 2 Methods

[0086] 2.1 Cell culture

[0087] Same as Example 1.

[0088] 2.2 MTT colorimetric experiment

[0089] In this experiment, GC-2 cells were intervened with different concentrations of bisphenol A (100 μM, 125 μM, 150 μM, 180 μM, 200 μM, 220 μM, 240 μM, 250 μM, BPA) for 24 h to observe the inhibitory effect of bisphenol A on spermatocyte proliferation.

[0090] Different concentrations of acteoside (5 μM, 10 μM, 20 μM, 30 μM, 40 μM, 50 μM, 60 μM) and cichoric acid (5 μM, 10 μM, 15 μM, 20 μM, 30 μM, 40 μM, 60 μM) were selected and co-administered with an appropriate concentration of BPA to explore their effect on promoting spermatocyte proliferation under the toxic environment of BPA.

[0091] Different ratios of acteoside:cichoric acid were selected to explore the effect of different ratios on spermatocyte proliferation under the toxic environment of an appropriate concentration of BPA.

[0092] The specific steps are as follows:

[0093] (1) GC-2 cells in the logarithmic growth phase were collected, and a single-cell suspension was prepared with DMEM medium containing 10% fetal bovine serum. The cells were seeded into a 96-well plate at a density of 8000 cells / well, and the volume per well was 180 μL.

[0094] (2) After the cells adhered, semi-media change was used for drug administration. 90 μL of the culture medium was aspirated from each group of cells, and then 90 μL of bisphenol A (BPA) at twice the final concentration of the corresponding cells was added according to the grouping. After 45 min of intervention, 90 μL of the drug solution was aspirated according to the grouping, and 90 μL of acteoside and cichoric acid at twice the final concentration of the corresponding cells were added. Each group had 6 replicate wells. The final volume in each well was the same, all 180 μL.

[0095] (3) After 24 h of drug intervention, 20 μL of MTT (5 mg / mL) was added to each well, and the cells were further incubated in a 5% CO2, 37 °C cell culture incubator for 4 h. The culture was terminated, the liquid in the wells was aspirated, and 150 μL of DMSO was added to each well and shaken for 3 min to fully dissolve the crystals.

[0096] (4) Colorimetric assay: The absorbance value (OD value) of each well was measured at a wavelength of 490 nm using an enzyme-linked immunosorbent assay (ELISA) reader. The optical density values of each group were statistically analyzed, and a cell growth trend graph was plotted with time as the abscissa and the OD value as the ordinate.

[0097] 3 Results

[0098] 3.1 High-concentration bisphenol A is toxic to spermatocytes

[0099] As Figure 2As shown, low concentrations of bisphenol A (BPA, 100, 125, 150 μM) have a promoting effect on the proliferative activity of spermatocytes, significantly increasing the number of spermatocytes (P < 0.001); when the concentration rises above 180 μM, the toxic effect of BPA on spermatocytes begins to appear (P < 0.05, 0.001), and 180 μM BPA was selected to construct the subsequent poisoning model.

[0100] 3.2 Low doses of acteoside and cichoric acid can improve the proliferative activity of spermatocytes in the BPA-toxic environment

[0101] As Figure 3 shown, in the BPA-toxic environment, low doses of acteoside (10 μM) and cichoric acid (5 μM) can significantly improve the proliferative activity of spermatocytes (P < 0.001, 0.05), and the proliferative-promoting effects of the two on spermatocytes show a dose-dependent trend in the range of 0 - 60 μM. Therefore, a 10 μM drug intervention group with significant differences from the 180 μM BPA group was further selected for subsequent experiments.

[0102] 3.3 The combined application of acteoside and cichoric acid has a promoting effect on the viability of spermatocytes in the BPA-toxic environment

[0103] As Figure 4 shown, the combined application of 5 μM and 10 μM acteoside and cichoric acid is more significant in promoting the proliferation of spermatocytes under BPA toxicity compared with the single application of 10 μM and 20 μM acteoside and cichoric acid (P < 0.05). Since the two components can show a significant promoting effect of combined administration when administered alone at 10 μM, the subsequent total concentration of the two-drug combination was selected as 10 μM as the standard to explore the effect of different compatibility ratios of the two components on the proliferation of spermatocytes.

[0104] Among them, in Figure 4 for example, "180 + 20 + 0" represents 180 μM BPA, 20 μM acteoside, and 0 μM cichoric acid; "180 + 0 + 10" represents 180 μM BPA, 0 μM acteoside, and 10 μM cichoric acid.

[0105] 3.4 Effects of different compatibility ratios of acteoside and cichoric acid on the viability of spermatocytes in the BPA-toxic environment

[0106] As Figure 5As shown, the ratio of acteoside to cistanoside A was 1:1 (i.e., 5 μM:5 μM), 4:1 (i.e., 8 μM:2 μM), 3:2 (i.e., 6 μM:4 μM), 3:1 (i.e., 7.5 μM:2.5 μM), 1:3 (i.e., 2.5 μM:7.5 μM). Compared with the 180 μM BPA group, the promoting effects on spermatocytes proliferation were all very significant (P < 0.05, 0.01, 0.001). The combined application of the two drugs at a ratio of 1:1 had the most significant promoting effect on the viability of spermatocytes in the BPA-toxic environment (P < 0.001), and the effect was better than that of 10 μM acteoside and 10 μM cistanoside A (P < 0.05, 0.01). At the same time, the combined application of 4:1 and 1:3 was also better than the 10 μM cistanoside A group to a certain extent, but there was no significant difference.

[0107] In addition, when the ratio of the two was 1:4 (i.e., 2 μM:8 μM) and 2:3 (i.e., 4 μM:6 μM), the promoting effects on the viability of spermatocytes were weaker compared with 10 μM acteoside and 10 μM cistanoside A (P < 0.05, 0.05; 0.01, 0.05. To ensure the cleanliness of the figure, they are not shown in the figure), and there was no significant difference compared with the 180 μM BPA group.

[0108] 4 Conclusions

[0109] The experiment showed that the environmental estrogen bisphenol A (BPA) showed estrogenic effects of promoting proliferation on spermatocytes at low concentrations, and toxic effects of inhibiting proliferation on spermatocytes at high concentrations, and there was a dose-dependence, which could be used to establish a spermatocyte damage model in vitro. Subsequently, different concentrations of acteoside and cistanoside A were administered in the BPA-toxic environment. The results showed that low doses of acteoside (10 μM) and cistanoside A (5 μM) could significantly improve the proliferation viability of spermatocytes (P < 0.001, 0.05). The two had a certain degree of protective effect on BPA-induced spermatocyte damage, suggesting that phenylethanoid glycosides from Cistanche deserticola have the effects of protecting spermatogenic cells and improving male reproductive ability. Further, in the BPA-toxic environment, different ratios of acteoside and cistanoside A were administered in combination. The results showed that the 1:1 compatibility of the two was the optimal concentration, and the effect was the most significant compared with the administration of acteoside and cistanoside A alone.

[0110] The above are only examples of the present invention, and common general knowledge such as specific features well known in the solution is not described in detail here. It should be pointed out that for those skilled in the art, without departing from the premise of the present invention, several modifications and improvements can be made, which should also be regarded as the protection scope of the present invention, and these will not affect the implementation effect of the present invention and the practicality of the patent. The protection scope required by the present invention should be subject to the content of its claims, and the specific implementation manners described in the specification can be used to explain the content of the claims.

[0111] The embodiments of the present invention have been introduced in detail above. Specific examples are used herein to elaborate on the principles and implementation manners of the present invention. The description of the above embodiments is only used to help understand the method and its core idea of the present invention. At the same time, any changes or deformations made by those skilled in the art based on the idea of the present invention, within the specific implementation manners and application scope of the present invention, fall within the protection scope of the present invention. In summary, the content of this specification should not be construed as a limitation to the present invention.

Claims

1. Use of a pharmaceutical composition composed of acteoside and echinacoside in the preparation of a drug for preventing and treating male infertility with decreased spermatocyte viability, characterized in that, The molar concentration ratio between the acteoside and the echinacoside is 1:

1.

2. Use of a pharmaceutical preparation in the manufacture of a medicament for preventing and treating male infertility caused by decreased spermatocyte viability, characterized in that, The preparation contains a pharmaceutical composition and pharmaceutically acceptable excipients, wherein the pharmaceutical composition consists of acteoside and echinacoside, and the molar concentration ratio between the acteoside and the echinacoside is 1:

1.

3. The use according to claim 2, characterized in that, The excipients are selected from one or more of the following: inclusion agent, flavoring agent, sustained-release agent, diluent, lubricant, disintegrant, suspending agent, light-blocking agent, dispersing agent, plasticizer, wetting agent, binder and antioxidant.

4. The use according to claim 3, characterized in that, The dosage form of the pharmaceutical preparation is an oral preparation, a sustained-release preparation, an injection, a controlled-release preparation or a targeted preparation.

5. The use according to claim 4, characterized in that, The oral preparation is a capsule, a powder, a granule or a pill.

6. The use according to any one of claims 1 to 5, characterized in that, The male infertility is oligospermia and asthenospermia.

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