Pine pollen compound fermentation composition as well as preparation method and application thereof

Abstract

The invention belongs to the technical field of traditional Chinese medicine, and relates to a pine pollen compound fermentation composition and a preparation method and application thereof, and the preparation method comprises the following steps: (1) water extraction; (2) enzymolysis; (3) sterilizing; (4) fermenting; (5) sterilizing; and (6) drying to obtain the pine pollen compound fermentation composition. The pine pollen compound fermentation composition disclosed by the invention can promote rapid dissolution of key active ingredients and enrich the content of active ingredients, namely flavone and polysaccharide, for improving benign prostatic hyperplasia in the raw materials, so that a foundation is laid for the benign prostatic hyperplasia improvement effect of a compound fermentation product; the inherent bitter taste of the pine pollen and other raw materials is effectively reduced, and the palatability of the product and the pleasure of consumers are improved; animal experiments prove that the traditional Chinese medicine composition has the beneficial effect of improving benign prostatic hyperplasia.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of traditional Chinese medicine, and relates to a pine pollen compound fermentation composition and a preparation method and application thereof. BACKGROUND

[0002] Benign prostatic hyperplasia is a common urological disease in middle-aged and elderly men, mainly manifested as symptoms such as frequent urination, urinary urgency, and dysuria, and severe cases can cause urinary retention, renal function damage and other complications. Although the existing treatment methods are effective, they are often accompanied by side effects or postoperative risks, which limits long-term use. Natural ingredients of "medicinal food" have a long history in traditional Chinese medicine for regulating prostate-related diseases. In recent years, they have received widespread attention due to their high safety and mild effects.

[0003] Traditional Chinese medicine fermentation technology can upgrade traditional Chinese medicine on the original basis, has the advantages of reducing toxicity and increasing efficacy, improving the content of active substances, improving flavor, and improving palatability. In addition, fermentation technology combined with the action of probiotics can produce a variety of metabolites beneficial to the human body. At present, there have been researches on applying fermentation technology to pollen products, which are mainly focused on product formula development, such as fruit wine, enzyme, fruit vinegar, and fermented tea made by combining pollen with medicinal and edible substances such as medlar and lotus leaf. Pine pollen itself has a bitter taste, and some traditional Chinese medicines and medicinal and edible substances in the pine pollen formula also have a strong medicinal taste and bitterness, which seriously affects the acceptance of pine pollen fermentation products by Chinese consumers. Therefore, how to maximize the content of active ingredients of pine pollen within the limited amount of addition, optimize the effective amount, and have an easily acceptable taste is a difficult problem to be solved. SUMMARY

[0004] The present application provides a pine pollen compound fermentation composition and a preparation method and application thereof in view of the deficiencies of the prior art, and the specific technical solutions are as follows: The first object of the present application is to provide a preparation method of a pine pollen compound fermentation composition, comprising the following steps: (1) water extraction: 10-15 parts of cistanche, 8-12 parts of astragalus, and 12-18 parts of dandelion are crushed and sieved, then mixed with 30-40 parts of pine pollen, and water is added for extraction to obtain a water extract; (2) enzyme hydrolysis: adding hemicellulase and pectinase to the water extract for enzyme hydrolysis to obtain an enzyme hydrolysis solution; (3) sterilization: sterilizing the enzyme hydrolysis solution to obtain a sterilized solution; (4) fermentation: inoculating lactobacillus rhamnosus and lactobacillus plantarum into the sterilized solution for fermentation to obtain a fermentation solution; (5) sterilization: sterilizing and cooling the fermentation solution to obtain a fermentation sterilized solution; (6) drying: freeze-drying the fermentation sterilization liquid to obtain the pine pollen compound fermentation composition.

[0005] The pine pollen compound fermentation composition has great superiority in improving benign prostatic hyperplasia, and has an excellent taste that is easy to accept, so the application range is very wide.

[0006] The pine pollen is the dry pollen of Pinaceae plants Pinus massoniana, Pinus tabulaeformis or several plants of the same genus, contains various chemical components, is rich in proteins, amino acids, vitamins, minerals, lipids, polypeptides, flavonoids, carotenes, auxins, polysaccharides and various bioactive substances, including 8 kinds of essential amino acids, 14 vitamins and 24 trace elements.

[0007] The Cistanche is the dry succulent stem with scale leaves of Cistanche deserticola Y.C.Ma or Cistanche tubulosa (Schenk) Woods, contains rich active ingredients, is rich in phenylethanoid glycosides, iridoid glycosides, polysaccharides, amino acids, trace elements, lipids, vitamins and various bioactive substances, including more than 15 kinds of amino acids and more than 20 kinds of trace elements such as iron, zinc and selenium; the Cistanche has the effects of tonifying kidney yang, benefiting essence and blood, moistening intestines and defecation, enhancing immunity and resisting fatigue.

[0008] The Astragalus is the dry root of Astragalus membranaceus (Fisch.) Bunge or Astragalus membranaceus (Fisch.) Bge. var. mongholicus (Regel) Hsiao, is rich in bioactive substances such as astragalosides, flavonoids and polysaccharides, and more than 20 kinds of amino acids and trace elements such as iron, zinc and selenium; the Astragalus has the effects of tonifying qi and consolidating the exterior, diuresis and removing toxin, pus discharge, and astringing and muscle regeneration.

[0009] The Taraxacum is the dry whole plant of Taraxacum mongolicum Hand.-Mazz., Taraxacum officinale F.H. Peral or several plants of the same genus, is rich in taraxacun sterols, choline, flavonoids, organic acids, vitamins, minerals and various bioactive substances; the Taraxacum has the effects of clearing heat and resolving toxins, removing swelling and resolving nodes, diuresis and relieving stranguria, and can be used for treating carbuncle, mastitis, internal abscess, stranguria and pain, and yellow jaundice.

[0010] Further, in step (1), the temperature of water extraction is 95-100℃, the extraction time is 1-3h, and the solid-liquid ratio is 1:8-12.

[0011] Further, in step (2), the total amount of added hemicellulase and pectinase is 1.0-1.5% (w / v), the volume ratio of hemicellulase to pectinase is (1-2):(1-2), the enzymatic hydrolysis temperature is 40-45℃, and the enzymatic hydrolysis time is 1-2h; preferably, the enzymatic hydrolysis temperature is 42℃, and the enzymatic hydrolysis time is 1.5h.

[0012] Further, in step (3), the sterilization condition is boiling water bath sterilization for 15-20min, cooling to room temperature, and then boiling water bath sterilization for 5-20min.

[0013] Further, in step (4), the total amount of inoculated Lactobacillus rhamnosus and Lactobacillus plantarum is 0.03-0.05% (w / v), the inoculation ratio of Lactobacillus rhamnosus to Lactobacillus plantarum is 1:1, the fermentation temperature is 37-45℃, and the fermentation time is 72-120h; preferably, the fermentation temperature is 40℃, and the fermentation time is 96h.

[0014] Further, in step (5), the sterilization condition is boiling water bath sterilization for 15-20min.

[0015] A second object of the present application is to provide a pine pollen compound fermentation composition prepared by the above preparation method.

[0016] A third object of the present application is to provide the use of the above pine pollen compound fermentation composition in the preparation of a drug for improving benign prostatic hyperplasia.

[0017] Further, the drug comprises a pharmaceutical excipient; the pharmaceutical excipient is selected from one or more than two combinations of wetting agents, emulsifying agents, preservatives, antioxidants, buffers, excipients, diluents, lubricants, bacteriostatic agents, suspending agents, suspending aids, solubilizers, thickening agents, stabilizers, sweeteners, and spices; preferably, the pharmaceutical excipient is at least one selected from lactose, mannose, starch, gum arabic, calcium phosphate, alginate, gelatin, calcium silicate, polyvinylpyrrolidone, cellulose, water, sugar syrup, methyl cellulose, methyl hydroxybenzoate, propyl hydroxybenzoate, magnesium stearate, and mineral oil.

[0018] Further, the dosage form of the drug is tablets, granules, oral liquids, capsules, or wine.

[0019] The present application has the following beneficial effects: (1) The preparation method of the pine pollen compound fermentation composition of the present application can promote the rapid dissolution of key active ingredients, enrich the active ingredients for improving benign prostatic hyperplasia in raw materials, and lay a foundation for the effect of the compound fermentation product on improving benign prostatic hyperplasia.

[0020] (2) The compound fermentation technology provided by the application effectively reduces the inherent bitter taste of pine pollen and other raw materials, and improves the palatability of the product and the consumer's degree of pleasure.

[0021] (3) The pine pollen compound fermentation composition of the application has beneficial effects on improving benign prostatic hyperplasia through animal experiments. BRIEF DESCRIPTION OF DRAWINGS

[0022] Figure 1 A graph of the effects of different compositions of the application on the prostate index of rats; Figure 2 The SOD level of rats gavaged with different compositions of the application; Figure 3 The GSH level of rats gavaged with different compositions of the application; Figure 4 The MDA level of rats gavaged with different compositions of the application. DETAILED DESCRIPTION

[0023] The principles and characteristics of the application will be described below in conjunction with the examples, which are only used to explain the application and not to limit the scope of the application.

[0024] The main material sources in the application are as follows: Pine pollen is provided by Yantai New Era Health Industry Co., Ltd.; Cistanche and Astragalus are provided by Hanzhong Natural Valley Biotechnology Co., Ltd.; Dandelion is provided by Beijing Baodexi Health Industry Co., Ltd.

[0025] Example 1 A preparation method of a pine pollen compound fermentation composition, comprising the following steps: (1) Water extraction: 15 parts of Cistanche, 10 parts of Astragalus, and 15 parts of dandelion are ground and sieved, then mixed with 40 parts of pine pollen, 10 times 100℃ hot water is added, stirred uniformly, extracted at 100℃ for 2h, and a water extract is obtained; (2) Enzymatic hydrolysis: 1.5% (w / v) of enzyme is added to the water extract, hemicellulase and pectinase are added, stirred at 42℃ for 1h, and an enzymatic hydrolysis solution is obtained; the volume ratio of hemicellulase to pectinase is 1:1, both are purchased from Novozymes (China) Investment Co., Ltd., and the enzyme activity is 100U / mg; (3) Sterilization: the enzymatic hydrolysis solution is boiled in a water bath for 15min to inactivate the enzyme, cooled to room temperature, then boiled in a water bath for 10min for sterilization, cooled to 40℃, and a sterilized solution is obtained; (4) Fermentation: inoculate Lactobacillus rhamnosus and Lactobacillus plantarum into the sterilized solution at an inoculation amount of 0.05% (w / v) and ferment at 45°C for 96h to obtain a fermentation liquor; the weight ratio of Lactobacillus rhamnosus and Lactobacillus plantarum is 1:1, both of which are purchased from Qingdao Nosan Biotechnology Co., Ltd. with the item number 20241208001; (5) Sterilization: sterilize the fermentation liquor in a boiling water bath for 20 min, cool to room temperature, and obtain a fermentation sterilized liquor; (6) Drying: freeze-dry the fermentation sterilized liquor, crush it, and obtain a pine pollen compound fermentation composition, which is then stored at -18°C for later use.

[0026] Example 2 A preparation method of a pine pollen compound fermentation composition includes the following steps: (1) Water extraction: according to weight parts, 15 parts of Cistanche, 10 parts of Astragalus, and 15 parts of Taraxacum are crushed and sieved, then mixed with 30 parts of pine pollen, 12 times 100°C hot water is added, stirred uniformly, extracted at 100°C for 2h, and a water extract is obtained; (2) Enzymolysis: according to an enzyme amount of 1.5% (w / v), hemicellulase and pectinase are added to the water extract, stirred at 45°C for 1h, and an enzyme hydrolysate is obtained; the volume ratio of hemicellulase and pectinase is 1:1.5, both of which are purchased from Novozymes (China) Investment Co., Ltd. with an enzyme activity of 100U / mg; (3) Sterilization: the enzyme hydrolysate is heated in a boiling water bath for 15 min, cooled to room temperature, then heated in a boiling water bath for 10 min, cooled to 40°C, and a sterilized solution is obtained; (4) Fermentation: inoculate Lactobacillus rhamnosus and Lactobacillus plantarum into the sterilized solution at an inoculation amount of 0.05% (w / v) and ferment at 42°C for 96h to obtain a fermentation liquor; the weight ratio of Lactobacillus rhamnosus and Lactobacillus plantarum is 1:1, both of which are purchased from Qingdao Nosan Biotechnology Co., Ltd. with the item number 20241208001; (5) Sterilization: sterilize the fermentation liquor in a boiling water bath for 20 min, cool to room temperature, and obtain a fermentation sterilized liquor; (6) Drying: freeze-dry the fermentation sterilized liquor, crush it, and obtain a pine pollen compound fermentation composition, which is then stored at -18°C for later use.

[0027] Example 3 A preparation method of a pine pollen compound fermentation composition includes the following steps: (1) Water extraction: according to weight parts, 15 parts of Cistanche, 10 parts of Astragalus, and 15 parts of Taraxacum are crushed and sieved, then mixed with 30 parts of pine pollen, 12 times 100°C hot water is added, stirred uniformly, extracted at 100°C for 2h, and a water extract is obtained; (2) Enzymolysis: 1.5% (w / v) enzyme was added to the water extract, and hemicellulase and pectinase were added to the water extract, and the mixture was stirred at 45°C for 1 h to obtain an enzyme hydrolysate; the volume ratio of hemicellulase to pectinase was 2:1, both purchased from Novozymes (China) Investment Co., Ltd., and the enzyme activity was 100 U / mg; (3) Sterilization: the enzyme hydrolysate was boiled in a water bath for 15 min to inactivate the enzyme, cooled to room temperature, then boiled in a water bath for 15 min to sterilize, cooled to 40°C, and a sterilized solution was obtained; (4) Fermentation: 0.04% (w / v) of the inoculum was inoculated into the sterilized solution, and Lactobacillus rhamnosus and Lactobacillus plantarum were inoculated at 40°C for 96 h to obtain a fermentation broth; the weight ratio of Lactobacillus rhamnosus to Lactobacillus plantarum was 1:1, both purchased from Qingdao Nuosen Biotechnology Co., Ltd., and the product code was 20241208001; (5) Sterilization: the fermentation broth was sterilized in a boiling water bath for 20 min, and then cooled to room temperature to obtain a fermentation sterilized solution; (6) Drying: the fermentation sterilized solution was freeze-dried and crushed to obtain a pine pollen compound fermentation composition, which was then packaged and stored at -18°C for later use.

[0028] Comparative Example 1 A preparation method of a pine pollen composition, comprising the following steps: According to the weight parts, 15 parts of Cistanche, 10 parts of Astragalus, and 15 parts of Taraxacum were crushed and sieved, then mixed with 40 parts of pine pollen, 10 times 100°C hot water was added, stirred uniformly, extracted at 100°C for 2 h, cooled to room temperature, freeze-dried, crushed, and packaged.

[0029] Comparative Example 2 A preparation method of a pine pollen water extract, comprising the following steps: According to the weight parts, 80 parts of pine pollen were added to 10 times 100°C hot water, stirred uniformly, extracted at 100°C for 2 h, cooled to room temperature, freeze-dried, crushed, and packaged.

[0030] Comparative Example 3 In step (1), according to the weight parts, the amount of pine pollen was 80 parts, and the others were the same as in Example 1, which will not be repeated here.

[0031] Comparative Example 4 In step (1), according to the weight parts, Cistanche 30 parts, Astragalus 20 parts, and Taraxacum 30 parts were used, and the others were the same as in Example 1, which will not be repeated here.

[0032] Comparative Example 5 In step (1), according to the weight parts, Cistanche 40 parts were used, and the others were the same as in Example 1, which will not be repeated here.

[0033] Evaluation experiment of enrichment effect of flavonoids and polysaccharides in different processing process samples In order to further clarify the advantages of the pine pollen compound fermentation composition in the enrichment of active ingredients, the flavonoid and polysaccharide contents in the pine pollen compound fermentation composition (Examples 1-3), the non-fermentation composition (Comparative Examples 1 and 2), and other fermentation compositions (Comparative Examples 3-5) were determined according to the following method.

[0034] (1) Extraction and determination of flavonoids The determination was carried out according to the method of "SN / T 4592-2016 Determination of Total Flavonoids in Export Food". The flavonoids in the sample were complexed with aluminum salt, and a yellow complex was formed under alkaline conditions. The absorbance was determined at a wavelength of 420 nm, and the absorbance was proportional to the content of flavonoid compounds within a certain concentration range. Compared with the rutin standard, the quantitative determination of total flavonoids in the test substance was carried out.

[0035] (2) Extraction and determination of polysaccharides The polysaccharides in the pine pollen fermentation product were extracted by water extraction and alcohol precipitation to obtain a polysaccharide sample solution. Different concentrations of glucose standard solution were prepared, 1.0 mL of standard solution and 1.0 mL of polysaccharide extraction sample were taken and placed in 10 mL centrifuge tubes, respectively, and water was added to each to 2.0 mL, and then 1.0 mL of phenol solution was added, 5.0 mL of concentrated sulfuric acid was added dropwise, shaken well, and placed for 5 min, then placed in a boiling water bath for 15 min, taken out and cooled to room temperature, and the absorbance was determined at 490 nm, and the polysaccharide content was calculated.

[0036] The detection results are shown in Table 1: Table 1 Detection results

[0037] Compared with the non-fermented pine pollen sample of Comparative Example 1 and the other fermentation composition of Comparative Example 3, the flavonoids of the pine pollen compound fermentation composition of Example 1 increased by 258.04% and 104.45%, respectively, and the polysaccharide content increased by 149.86% and 87.65%, respectively, indicating that the compound fermentation technology of the present application has the effect of promoting the rapid dissolution of key active ingredients and realizing the enrichment of components, and pine pollen and Cistanche have the effect of synergistically improving the content of flavonoids and polysaccharides, which lays a solid foundation for improving the effect of the compound fermentation product on improving benign prostatic hyperplasia.

[0038] Sensory evaluation experiment of compound fermented pine pollen product before and after fermentation Evaluation method: sensory evaluation was performed on the samples by human taste. 50 people were selected as the test population (25 men and 25 women, aged 30-60 years old), and the samples were tasted and comprehensively evaluated by the testers without knowing the sample, and the higher the score, the better the taste and quality. The test score is 100 points, including five aspects: overall taste acceptance (20 points), bitter taste acceptance (15 points), astringent taste acceptance (15 points), sour taste acceptance (15 points), salty taste acceptance (15 points), odor (10 points), and dissolution (10 points). The test results are expressed in points per person.

[0039] Experimental results and conclusions: The sensory evaluation results of the pine pollen compound fermentation composition (Example 1) and the pine pollen non-fermentation composition (Comparative Example 1) are shown in Table 2.

[0040] Table 2 Sensory evaluation results

[0041] As can be seen from the evaluation results in Table 2, after fermentation, the sensory quality of the pine pollen compound is obviously improved. Compared with the pine pollen non-fermentation composition (Comparative Example 1), the pine pollen compound fermentation (Example 1) is obviously improved in overall taste acceptance, bitter taste acceptance, and astringent taste acceptance, respectively increased by 41.46%, 66.67%, and 65.06%, respectively, indicating that the fermentation process effectively reduces the inherent bitter and astringent taste of the pine pollen compound, improves the palatability of the product, and improves the consumer's enjoyment.

[0042] Animal experiment for evaluating the effect of pine pollen compound fermentation composition on improving benign prostatic hyperplasia Experimental method: In this experiment, a rat model of prostatic hyperplasia was used to evaluate the intervention effect of each test sample on benign prostatic hyperplasia by measuring the prostate index and the oxidative stress level of the prostate tissue.

[0043] (1) Experimental animals: 6-week-old SPF healthy male SD rats were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd.

[0044] (2) Experimental drugs and test samples Experimental drugs: testosterone propionate was purchased from China National Pharmaceutical Group Corporation, and superoxide dismutase (SOD), malondialdehyde (MDA), and glutathione (GSH) test kits were purchased from Lanjiekeli Technology Co., Ltd.

[0045] Test samples: pine pollen compound fermentation composition (Example 1), pine pollen compound non-fermentation (Comparative Example 1 and Comparative Example 2), and other fermentation compositions (Comparative Example 3-Comparative Example 5).

[0046] (3) Experimental animal grouping: animal experiment grouping, gavage dose and number of experimental animals are shown in Table 3: Table 3 Animal experiment grouping, gavage dose and number of experimental animals

[0047] (4) Experimental specific implementation process Forty-eight 6-week-old SPF healthy male SD rats were raised in a barrier environment (specific pathogen-free, temperature 20-26℃, humidity 55-70%, 12-hour day-night alternation), and the rats were allowed to eat and drink freely during the feeding period. After one week of adaptive feeding, they were divided into 8 groups, 6 rats in each group. Except for the blank control group, intraperitoneal injection of testosterone propionate 5 mg / (kg·d) for 28 days was used to establish a benign prostatic hyperplasia model. During the modeling period, the rats' bedding was checked daily for dryness and moisture, and their mental state was observed. The bedding was quickly moistened, the rats were irritable, their fur was yellow and withered, some of their skin was swollen (due to subcutaneous injection of testosterone propionate), and their urine output was significantly reduced at the 4th week of modeling. These were considered as successful modeling, and the successfully modeled rats were used for subsequent experiments. When conducting pharmacodynamic studies based on this model, the treatment period of the drug should be as close as possible to the modeling period to reduce the self-recovery ability of the body after modeling and the false positive effects on the experimental results. Therefore, the intervention was performed simultaneously with the modeling, and the experimental period was 5 weeks.

[0048] The drugs in each experimental group were prepared according to the formula in Table 1 and dissolved in normal saline. The volume of gavage was 1 mL / 100g body weight. The concentration of the test drug was 50 mg / mL. Gavage was used for drug administration, and the blank and model groups were given normal saline of the same volume. The drug administration frequency was once a day, and the administration lasted for 4 weeks. The rats were allowed to eat and drink freely during the experiment. Intraperitoneal injection was performed 30 minutes after the end of gavage. The concentration of testosterone propionate was 5 mg / mL, and the injection dose was 5 mg / (kg·d). The blank group was injected with the same volume of normal saline. The fasting body weight was measured every week. On the 5th week, the rats were anesthetized with isoflurane inhalation, and blood samples were collected from the abdominal aorta after anesthesia. The blood samples were collected in disposable coagulation blood collection tubes and centrifuged at 3000 rpm for 20 minutes at 4℃. The supernatant (serum) was stored in a -80℃ freezer. Then the rat's prostate was removed and weighed, half of it was quickly stored in liquid nitrogen, and the other half was placed in a 4% paraformaldehyde solution for storage.

[0049] (5) Test of prostatic hyperplasia phenotype The removed prostate was washed with normal saline and dried with filter paper. It was then weighed on an analytical balance and the prostate index was calculated using the following formula: Prostate index (mg / g) = Prostate wet weight (mg) / Body weight (g) x 100 (6) Determination of oxidative stress level in rat prostate tissue The collected prostate tissue was homogenized and then the oxidative stress level was determined according to the kit.

[0050] (7) Statistical methods All experiments were repeated at least three times, and the experimental data are presented as mean ± SD. One-way ANOVA was performed using SPSS 20.0 software, where p < 0.05 indicated a significant difference.

[0051] Experimental results and conclusions (1) Intervention of pine pollen compound fermentation composition on prostate hyperplasia index in rats The prostate index of rats after oral administration of each composition was as follows: Figure 1 As shown, after testosterone propionate modeling, the prostate index of the model group rats increased significantly compared with the blank control group, indicating that the model group rats had prostate hyperplasia. After oral administration of the test sample, the prostate index decreased to varying degrees. Compared with the model group, the prostate index of the rats in the low-dose experimental group of Example 1, the experimental group of Comparative Example 3, the experimental group of Comparative Example 4 and the experimental group of Comparative Example 5 were significantly reduced (p < 0.05) ( Figure 1 ), while the prostate index of rats in the high-dose group of Example 1 was significantly decreased (p < 0.01) ( Figure 1 ), and even recovered to the level of the normal group, which shows that the pine pollen compound fermentation composition (Example 1) has a better effect in improving benign prostatic hyperplasia.

[0052] (2) Intervention of pine pollen compound fermentation composition on prostate oxidative stress in rats with benign prostatic hyperplasia Oxidative stress is one of the main causes of cell damage, inflammatory response and tissue fibrosis. Prostate tissues of patients with benign prostatic hyperplasia often show higher MDA levels and lower SOD and GSH levels. Figure 2 、 Figure 3 and Figure 4 As shown in the figure, after testosterone propionate modeling, the SOD and GSH levels of prostate tissue were significantly reduced, and the MDA level was significantly increased, indicating that testosterone propionate caused a severe oxidative stress reaction in the prostate tissue. The SOD and GSH levels of rats gavaged with different compositions increased to varying degrees, while the MDA level was reduced. Compared with the model group, the SOD level and GSH level of the low-dose experimental group and high-dose experimental group in Example 1 were significantly different (p < 0.01) ( Figure 2 、 Figure 3 ), while the MDA level in the high-dose experimental group of Example 1 was significantly reduced (p < 0.05) ( Figure 4), which indicates that the compound fermented composition of pine pollen (Example 1) has a better effect of improving the oxidative stress of the prostate tissue of the rats with prostate hyperplasia.

[0053] The above merely describes the preferred embodiments of the present application and is not intended to limit the present application. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present application shall be included in the protection scope of the present application.

Claims

1. A method for preparing a pine pollen compound fermentation composition, characterized in that: The steps include: (1) Water extraction: by weight, 10-15 parts of Cistanche deserticola, 8-12 parts of Astragalus membranaceus, and 12-18 parts of Taraxacum officinale were crushed and sieved, and then mixed with 30-40 parts of pine pollen, and extracted with water to obtain a water extract; (2) Enzymatic hydrolysis: adding hemicellulase and pectinase to the aqueous extract for enzymatic hydrolysis to obtain an enzymatic hydrolyzate; (3) Sterilization: sterilize the enzymatic hydrolyzate to obtain a sterilized solution; (4) Fermentation: inoculating Lactobacillus rhamnosus and Lactobacillus plantarum into the sterilized solution and fermenting to obtain a fermentation liquid; (5) Sterilization: Sterilize and cool the fermentation liquid to obtain a sterilized fermentation liquid; (6) Drying: freeze-dry the fermentation sterilized liquid to obtain the pine pollen compound fermentation composition.

2. The method for preparing the pine pollen compound fermentation composition according to claim 1, characterized in that: In step (1), the water extraction temperature is 95-100° C., the extraction time is 1-3 h, and the solid-liquid ratio is 1:8-12.

3. The method for preparing the pine pollen compound fermentation composition according to claim 1, characterized in that: In step (2), the total amount of hemicellulase and pectinase added is 1.0~1.5% (w / v), the volume ratio of hemicellulase to pectinase is (1~2):(1~2), the enzymatic hydrolysis temperature is 40~45°C, and the enzymatic hydrolysis time is 1~2h.

4. The method for preparing the pine pollen compound fermentation composition according to claim 1, characterized in that: In step (3), the sterilization conditions are as follows: sterilize in a boiling water bath for 15 to 20 minutes, cool to room temperature, and then sterilize in a boiling water bath for 5 to 20 minutes.

5. The method for preparing the pine pollen compound fermentation composition according to claim 1, characterized in that: In step (4), the total inoculation amount of Lactobacillus rhamnosus and Lactobacillus plantarum is 0.03-0.05% (w / v), the inoculation ratio of Lactobacillus rhamnosus and Lactobacillus plantarum is 1:1, the fermentation temperature is 37-45°C, and the fermentation time is 72-120h.

6. The method for preparing the pine pollen compound fermentation composition according to claim 1, characterized in that: In step (5), the sterilization conditions are: boiling water bath sterilization for 15 to 20 minutes.

7. A pine pollen compound fermentation composition, characterized in that: The composition is prepared by the preparation method of the pine pollen compound fermentation composition according to any one of claims 1 to 6.

8. Use of the pine pollen compound fermentation composition according to claim 7 in preparing a drug for improving benign prostatic hyperplasia.

9. The use according to claim 8, characterized in that The drug includes pharmaceutical excipients.

10. The use according to claim 8, characterized in that The dosage form of the medicine is tablet, granule, oral solution, capsule or wine.

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