A compound propolis composition, its quality detection method and application

Abstract

The present invention belongs to the technical field of traditional Chinese medicines, and particularly relates to a compound propolis composition, a quality detection method thereof and an application. The compound propolis composition is prepared from the following raw materials: propolis, ginkgo leaves, and bletilla striata. The compound propolis composition of the present invention has a remarkable effect on preventing and treating skin chaps; the preparation method of the compound propolis composition is simple and controllable; the quality detection method uses high performance liquid chromatography (HPLC) to simultaneously determine the contents of two active ingredients, quercetin and kaempferol, in the product, and the detection method is simple, accurate and has good reproducibility.

Description

Technical Field

[0001] The present invention relates to the technical field of traditional Chinese medicines, and particularly relates to a compound propolis composition, a quality detection method thereof and an application.

Background Art

[0002] Skin chapping refers to the appearance of crisscross cracks or fissures on the skin surface. This usually occurs on dry skin, especially in cold and dry winters. Skin chapping usually appears on relatively thin-skinned areas such as the hands, feet, lips and face.

[0003] The main cause of skin chapping is the loss of moisture in the skin, resulting in a dry, hardened and fragile stratum corneum. Common factors leading to skin chapping include dry climate, frequent handwashing, excessive use of chemicals and detergents, prolonged immersion in water or contact with irritating substances, etc.

[0004] Skin chapping can cause pain, discomfort and itching. Severe skin chapping may lead to bleeding, infection and scar formation.

[0005] Methods for preventing skin chapping include keeping the skin moderately moist, avoiding the use of irritating substances, avoiding frequent handwashing, using mild skin care products, and maintaining sufficient water intake.

[0006] In recent years, some natural plant extracts have been found to have the effect of repairing and protecting the skin and are suitable for the treatment of skin chapping. For example: aloe vera extract, green tea extract, lavender extract, witch hazel extract, etc. Most of these natural plant extracts are used in skin care products in the form of ingredients or essential oils. Among the above extracts, some are extracted with water, some are extracted with alcohol, and the concentrations of the alcohols selected are also different. As is well known, traditional Chinese herbs contain very complex components, including various active components, inactive components, and toxic components. When different solvents and different methods are used for extraction, the obtained active components are different, which will affect the quality stability and effectiveness of the products.

[0007] The inventors found in preliminary experiments that propolis, ginkgo leaves and Bletilla striata have a preliminary anti-skin chapping effect. If the processes and quality control methods of propolis, ginkgo leaves and Bletilla striata can be determined in combination with pharmacodynamic experiments, the quality stability and uniformity of the products can be ensured, and their clinical efficacy can be guaranteed.

Summary of the Invention

[0008] To solve the above technical problems, the present invention provides a compound propolis composition, its quality detection method and application. The compound propolis composition has a significant effect on preventing and treating skin chapping; the preparation method of the compound propolis composition is simple and controllable; the quality detection method uses high performance liquid chromatography (HPLC) to simultaneously determine the contents of two active ingredients, quercetin and kaempferol, in the product. The detection method is simple, accurate and has good reproducibility, ensuring the quality stability and uniformity of the product and the clinical efficacy.

[0009] To achieve the above object, the technical scheme adopted by the present invention is as follows:

[0010] A compound propolis composition is prepared from the following raw materials: propolis, ginkgo biloba leaves, and bletilla striata.

[0011] Further, it is prepared from the following raw materials in parts by weight: 1 part of propolis, 0.1 - 0.3 part of ginkgo biloba leaves, and 0.1 - 0.3 part of bletilla striata.

[0012] Further, it is prepared from the following raw materials in parts by weight: 1 part of propolis, 0.2 part of ginkgo biloba leaves, and 0.2 part of bletilla striata.

[0013] A method for preparing the compound propolis composition of the present invention includes the following steps:

[0014] Step 1), after freeze - drying the propolis, it is pulverized into fine powder of 60 - 80 meshes, put into an ethanol solution with a volume fraction of 90 - 95% and a weight 6 - 8 times that of the total weight of the propolis, and ultrasonically extracted at 50 - 60 °C for 1 - 3 h to obtain a propolis ethanol solution. The ethanol is recovered by reduced pressure concentration until there is no alcohol smell, and a concentrated solution A is obtained for standby.

[0015] Step 2), put the ginkgo biloba leaves and bletilla striata into an extraction container, and reflux extract twice. Each time, an ethanol solution with a volume fraction of 50 - 70% and a weight 6 - 8 times that of the total weight of the ginkgo biloba leaves and bletilla striata is added. The first extraction is for 2 - 3 hours, and the second extraction is for 1.5 - 2 hours. The filtrates of the two extractions are concentrated by reduced pressure to recover ethanol until there is no alcohol smell, and a concentrated solution B is obtained for standby.

[0016] Step 3), mix the concentrated solution A prepared in the above step 1) with the concentrated solution B of step 2), or further add excipients to make a spray preparation, and that's it.

[0017] The quality detection method of the compound propolis composition of the present invention is to simultaneously determine the liquid chromatographic contents of two chemical components, quercetin and kaempferol, contained in the compound propolis composition under the same chromatographic conditions.

[0018] Further, the quality detection method is carried out according to the following steps:

[0019] (1) Chromatographic conditions: The chromatographic column is a C18 column, the mobile phase is methanol - 0.4% phosphoric acid; the ultraviolet detection wavelength is 360 nm; the column temperature is 30 °C; the injection volume is 10 μl;

[0020] (2) Preparation of reference solution: Weigh appropriate amounts of quercetin and kaempferol reference substances accurately, place them in the same volumetric flask, dissolve with methanol solution with a volume fraction of 80%, and obtain.

[0021] (3) Preparation of test solution: Take the compound propolis composition, add methanol, ultrasonically treat, cool, filter, take the continuous filtrate, add hydrochloric acid, hydrolyze in a water bath, cool, make up the volume with methanol, filter, and take the continuous filtrate to obtain.

[0022] (4) Determination method: Determine according to the chromatographic conditions described in step (1) to obtain.

[0023] Furthermore, in step (1), in the mobile phase, the volume ratio of methanol to 0.4% phosphoric acid is 55:45.

[0024] Furthermore, step (3) includes taking 5 ml of the compound propolis composition, weighing accurately, placing it in a 50 ml volumetric flask, adding an appropriate amount of methanol, ultrasonically treating for 60 min, ultrasonic power 250 W, ultrasonic frequency 40 kHz, cooling, making up the volume with methanol to the scale, shaking well, filtering, taking 25 ml of the continuous filtrate, placing it in a 50 ml volumetric flask, adding 5 ml of hydrochloric acid with a mass fraction of 10%, hydrolyzing in a water bath at 90 °C for 1 h, taking out, cooling, making up the volume with methanol, filtering, and taking the continuous filtrate to obtain.

[0025] The present invention also provides the application of the compound propolis composition in the preparation of products for preventing and treating skin chapping.

[0026] The raw material sources of the present invention are as follows:

[0027] Propolis, this product is a viscous solid gum formed by the mixture of plant resins collected by worker bees of the Italian bee Apis mellifera L. of the bee family and their secretions from the mandibular gland, wax gland, etc.

[0028] Ginkgo leaf, this product is the dried leaf of the Ginkgo biloba L. plant of the Ginkgoaceae family. It is harvested when the leaves are still green in autumn and dried in time.

[0029] Bletilla striata, this product is the dried tuber of the Bletilla striata (Thunb.) Reichb.f. plant of the Orchidaceae family. It is dug in summer and autumn, the fibrous roots are removed, washed, boiled or steamed in boiling water until there is no white core, dried to half dry, the outer skin is removed, and then dried in the sun.

[0030] In summary, due to the adoption of the above technical solutions, the beneficial effects of the present invention are:

[0031] (1) The compound propolis composition of the present invention has a significant effect on preventing and treating skin chapping and has good skin repair function.

[0032] (2) The determination method of the present invention realizes one-time injection and simultaneously determines the contents of two active ingredients, quercetin and kaempferol, in the compound propolis composition. The chromatographic peaks have good shapes and high resolution, shorten the detection time, and improve the work efficiency. This method is simple, accurate, has good precision, reproducibility and stability, and can provide guarantee for the quality stability and effectiveness of the product.

Description of Drawings

[0033] Figure 1 : High performance liquid chromatography (HPLC) chromatogram (360 nm) of the mixed solution of quercetin and kaempferol reference substances.

[0034] Figure 2 : Chromatogram (360 nm) for the determination of the compound propolis composition prepared in Example 1 (batch number 20230901).

[0035] Figure 1 and Figure 2 In, the chromatographic peaks correspond to: A is quercetin; B is kaempferol.

[0036] Figure 3 : Expression of inflammatory factor IL-6 in sera of each group.

[0037] Figure 4 : Expression of inflammatory factor IL-10 in sera of each group.

Specific Embodiments

[0038] Example 1: Preparation of the compound propolis composition of the present invention

[0039] Prescription: 100 g of propolis, 20 g of Ginkgo biloba leaves, 20 g of Bletilla striata, and made into 1000 ml.

[0040] Preparation method:

[0041] Step 1), after freeze-drying propolis, it is pulverized into fine powder of 60 meshes, put into an ethanol solution with a volume fraction of 95% and a weight 7 times that of the total weight of the propolis, and ultrasonically extracted at 55 °C for 2 h to obtain a propolis ethanol solution. The ethanol is recovered by decompression concentration until there is no alcohol smell, and the concentrated solution A is reserved for later use;

[0042] Step 2), put Ginkgo biloba leaves and Bletilla striata into an extraction container and reflux extract twice. Each time, add an ethanol solution with a volume fraction of 60% and a weight 7 times that of the total weight of Ginkgo biloba leaves and Bletilla striata. The first extraction is for 2 h, and the second extraction is for 1.5 h. The filtrates of the two extractions are concentrated by decompression to recover ethanol until there is no alcohol smell, and the concentrated solution B is reserved for later use;

[0043] Step 3): Mix the concentrated liquid A prepared in the above step 1) with the concentrated liquid B of step 2), and add water to 1000 ml to obtain the product.

[0044] Example 2: Preparation of the compound propolis composition of the present invention

[0045] Prescription: 100 g of propolis, 10 g of Ginkgo biloba leaves, 30 g of Bletilla striata, to make 1000 ml.

[0046] Preparation method:

[0047] Step 1): After freeze-drying propolis, it is crushed into fine powder of 80 meshes, put into an ethanol solution with a volume fraction of 92% and a six-fold amount of the total weight of the propolis, and ultrasonically extracted at 50 °C for 1 h to obtain a propolis ethanol solution. The ethanol is recovered by reduced pressure concentration until there is no alcohol smell, and the concentrated liquid A is reserved for later use.

[0048] Step 2): Put Ginkgo biloba leaves and Bletilla striata into the extraction container, and extract by reflux twice. Each time, add an ethanol solution with a volume fraction of 50% and a six-fold amount of the total weight of Ginkgo biloba leaves and Bletilla striata. Extract for 2 h for the first time and 1.5 h for the second time. The filtrates of the two extractions are concentrated by reduced pressure to recover ethanol until there is no alcohol smell, and the concentrated liquid B is reserved for later use.

[0049] Step 3): Mix the concentrated liquid A prepared in the above step 1) with the concentrated liquid B of step 2), and add water to 1000 ml to obtain the product.

[0050] Example 3: Preparation of the compound propolis composition of the present invention

[0051] Prescription: 100 g of propolis, 30 g of Ginkgo biloba leaves, 10 g of Bletilla striata, to make 1000 ml.

[0052] Preparation method:

[0053] Step 1): After freeze-drying propolis, it is crushed into fine powder of 60 meshes, put into an ethanol solution with a volume fraction of 90% and an eight-fold amount of the total weight of the propolis, and ultrasonically extracted at 60 °C for 3 h to obtain a propolis ethanol solution. The ethanol is recovered by reduced pressure concentration until there is no alcohol smell, and the concentrated liquid A is reserved for later use.

[0054] Step 2): Put Ginkgo biloba leaves and Bletilla striata into the extraction container, and extract by reflux twice. Each time, add an ethanol solution with a volume fraction of 70% and an eight-fold amount of the total weight of Ginkgo biloba leaves and Bletilla striata. Extract for 3 h for the first time and 2 h for the second time. The filtrates of the two extractions are concentrated by reduced pressure to recover ethanol until there is no alcohol smell, and the concentrated liquid B is reserved for later use.

[0055] Step 3): Mix the concentrated liquid A prepared in the above step 1) with the concentrated liquid B of step 2), and add water to 1000 ml to obtain the product.

[0056] Comparative Example 1: Preparation of the compound propolis composition

[0057] The difference between Comparative Example 1 and Example 1 is that the prescription of Comparative Example 1 does not contain ginkgo leaves, and the original share of ginkgo leaves is replaced by bletilla striata, as follows:

[0058] Prescription: 100 g of propolis and 40 g of bletilla striata, made into 1000 ml.

[0059] Preparation method:

[0060] Step 1), after freeze-drying propolis, it is pulverized into fine powder of 60 meshes, put into an ethanol solution with a volume fraction of 95% which is 7 times the total weight of the propolis, and ultrasonically extracted at 55 °C for 2 h to obtain a propolis ethanol solution. Ethanol is recovered by reduced pressure concentration until there is no alcohol smell, and the concentrated solution A is obtained for standby;

[0061] Step 2), put bletilla striata into the extraction container, and reflux extract twice. Each time, add an ethanol solution with a volume fraction of 60% which is 7 times the total weight of the bletilla striata. The first extraction is for 2 h, and the second extraction is for 1.5 h. The filtrates of the two extractions are concentrated by reduced pressure to recover ethanol until there is no alcohol smell, and the concentrated solution B is obtained for standby;

[0062] Step 3), mix the concentrated solution A prepared in Step 1) above with the concentrated solution B in Step 2), and add water to 1000 ml to obtain it.

[0063] Comparative Example 2: Preparation of Compound Propolis Composition

[0064] The difference between Comparative Example 2 and Example 1 is that the prescription of Comparative Example 2 does not contain bletilla striata, and the original share of bletilla striata is replaced by ginkgo leaves, as follows:

[0065] Prescription: 100 g of propolis and 40 g of ginkgo leaves, made into 1000 ml.

[0066] Preparation method:

[0067] Step 1), after freeze-drying propolis, it is pulverized into fine powder of 60 meshes, put into an ethanol solution with a volume fraction of 95% which is 7 times the total weight of the propolis, and ultrasonically extracted at 55 °C for 2 h to obtain a propolis ethanol solution. Ethanol is recovered by reduced pressure concentration until there is no alcohol smell, and the concentrated solution A is obtained for standby;

[0068] Step 2), put ginkgo leaves into the extraction container, and reflux extract twice. Each time, add an ethanol solution with a volume fraction of 60% which is 7 times the total weight of the ginkgo leaves. The first extraction is for 2 h, and the second extraction is for 1.5 h. The filtrates of the two extractions are concentrated by reduced pressure to recover ethanol until there is no alcohol smell, and the concentrated solution B is obtained for standby;

[0069] Step 3), mix the concentrated solution A prepared in Step 1) above with the concentrated solution B in Step 2), and add water to 1000 ml to obtain it.

[0070] Comparative Example 3: Preparation of Compound Propolis Composition

[0071] The difference between Comparative Example 3 and Example 1 is that in Comparative Example 3, propolis, Ginkgo biloba leaves, and Bletilla striata are all extracted by water extraction method, specifically as follows:

[0072] Prescription: 100 g of propolis, 20 g of Ginkgo biloba leaves, and 20 g of Bletilla striata, made into 1000 ml.

[0073] Preparation method:

[0074] Step 1), after freeze-drying propolis, it is crushed into fine powder of 60 meshes, put into water with a weight 7 times that of the total weight of the propolis, and ultrasonically extracted at 55 °C for 2 h to obtain a propolis solution, which is concentrated under reduced pressure to obtain concentrated solution A for standby;

[0075] Step 2), put Ginkgo biloba leaves and Bletilla striata into the extraction container, and reflux extract twice. Each time, add water with a weight 7 times that of the total weight of Ginkgo biloba leaves and Bletilla striata. The first extraction is for 2 h, and the second extraction is for 1.5 h. The filtrates of the two extractions are concentrated by the method of concentration under reduced pressure to obtain concentrated solution B for standby;

[0076] Step 3), mix the concentrated solution A prepared in Step 1) above with the concentrated solution B in Step 2), and add water to 1000 ml to obtain it.

[0077] The following verifies the technical effects of the present invention through experimental research:

[0078] Experiment 1: Application effect of products for preventing and treating skin chapping

[0079] Design an experimental plan with reference to the following literature:

[0080] Wang Lu, Zhao Chunli, Li Xing, etc. Influence of Bletilla striata gum on interleukin expression in serum of rats with skin chapping [J]. Science and Technology Innovation, 2019(23): 2. DOI: 10.3969 / j.issn.1673 - 1328.2019.23.007..

[0081] Specifically as follows:

[0082] 1. Test drugs

[0083] Compound propolis compositions obtained from Example 1 - Example 3 and Comparative Example 1 - Comparative Example 3.

[0084] 2. Experimental animals

[0085] 72 healthy adult SD rats, half male and half female, with a body weight of about 230 g, provided by the Experimental Center of Guangxi University of Chinese Medicine.

[0086] 3. Experimental conditions

[0087] The laboratory temperature is 22℃ - 24℃, and the humidity is 60 - 70%.

[0088] 4. Experimental methods

[0089] 72 rats were randomly divided into nine groups for the experiment: blank control group, model group, positive control group, Example 1 group - Example 3 groups, Comparative Example 1 group - Comparative Example 3 groups, with 8 rats in each group. Adaptive feeding was carried out for 5 days before the experiment.

[0090] 24 hours before the experiment, hair removal was performed on an area of 3.0 cm × 3.0 cm on the back of the rats. After hair removal, the blank control group was smeared with physiological saline, and the remaining groups were continuously smeared with 5 mol / L sodium hydroxide solution for 7 days to establish a skin crack wound model. After successfully establishing the model, the model group was not administered medicine and was allowed to recover naturally. The positive control group was smeared with urea ointment, and the administered groups (Example 1 group - Example 3 groups and Comparative Example 1 group - Comparative Example 3 groups) were respectively sprayed with the corresponding compound propolis composition, once in the morning and once in the evening, for 7 consecutive days.

[0091] After 7 days, the upper serum of the abdominal aortic blood of the rats was collected for standby. The ELISA method was used to measure the inflammatory factors interleukin IL-6 and IL-10 in the serum. The operation was carried out according to the kit instructions, and the absorbance was detected using an enzyme-labeled analyzer. According to the measured absorbance data, a standard curve of the inflammatory factors was drawn, and their average dilution concentration was calculated. A column chart was drawn to show the results of each inflammatory factor.

[0092] The experimental result data was statistically analyzed and expressed in the form of (x ± SD). ANOVA test was used to analyze the differences between groups. A P value less than 0.05 indicates statistical significance, and a P value less than 0.01 indicates a significant difference.

[0093] 5. Experimental results

[0094] As Figure 3 and Figure 4 shown. Figure 3 and Figure 4 In, compared with the blank control group, *: p < 0.05; **: p < 0.01; compared with the model group, #: p < 0.05; ##: p < 0.01; compared with Example 1 group, ▲: p < 0.05; ▲▲: p < 0.01 for Comparative Example 1 group; compared with Example 1 group, ●: p < 0.05; ●●: p < 0.01 for Comparative Example 2 group; compared with Example 1 group, &: p < 0.05; &&: p < 0.01 for Comparative Example 3 group.

[0095] The results showed that after the model was established, compared with the blank control group, the contents of inflammatory factors IL-6 and IL-10 in the model group increased significantly, with a significant difference (P<0.01). After 7 days of administration, compared with the model group, the administration group showed a significant difference (P<0.05 - 0.01), indicating that the compound propolis composition of the present invention has a significant effect on preventing and treating skin chapping. Among them, the groups of Examples 1 - 3 had the most significant decrease. In addition, compared with Example 1, the groups of Comparative Examples 1 - 3 had significantly higher contents of inflammatory factors IL-6 and IL-10, indicating that there is a synergistic effect among propolis, ginkgo leaf and bletilla striata, and the combination of the three herbs can exert the maximum medicinal effect, and the product obtained by the ethanol extraction method has a stronger medicinal effect than the water extraction method. The above results show that the component selection of the present invention is reasonable and the preparation method is appropriate, and the maximum medicinal effect is fully exerted.

[0096] Experiment 2: Determination of the contents of two chemical components, quercetin and kaempferol, contained in the compound propolis composition by liquid chromatography.

[0097] 1. Instruments and test drugs

[0098] 1.1 Instruments Shimadzu LC-20AD XR high performance liquid chromatography system (Shimadzu Corporation, Japan); analytical balance (Sartorius Scientific Instruments Beijing Co., Ltd.); ultrasonic cleaner (Shenzhen Jiemeng Cleaning Equipment Co., Ltd.).

[0099] 1.2 Reagents Quercetin and kaempferol reference standards (purity mass fraction > 98%, provided by Shanghai Anpu Chuisheng Standard Technology Service Co., Ltd.). Methanol is of chromatographic purity; pure water for experiments, and other reagents are all of analytical purity.

[0100] 1.3 Medicinal materials Propolis, ginkgo leaf and bletilla striata were all purchased from Nanning Renyuan Chinese Medicinal Materials Professional Cooperative.

[0101] 2. Chromatographic conditions and system suitability test

[0102] Chromatographic column: ZORBAX Eclipse C18 column (250mm × 4.6mm, 5μm);

[0103] Mobile phase: methanol - 0.4% phosphoric acid (55:45);

[0104] Flow rate: 0.8ml / min; ultraviolet detection wavelength is 360nm;

[0105] Column temperature 30°C; injection volume 10μl.

[0106] Under the above conditions, quercetin and kaempferol were well separated, the resolution between adjacent peaks was greater than 1.5, and the calculated theoretical plate number was not less than 5000. The results are as Figure 1 shown.

[0107] 3. Preparation of Test Solution

[0108] 3.1 Preparation of Reference Solution

[0109] Weigh an appropriate amount of quercetin reference substance and kaempferol reference substance accurately, dissolve them in 80% methanol, make up the volume to a definite value, shake well, and prepare a mixed solution containing 50 μg of quercetin and 30 μg of kaempferol per 1 ml, that is, obtain it.

[0110] 3.2 Preparation of Test Sample Solution

[0111] Take 5 ml of the compound propolis composition, weigh it accurately, place it in a 50-ml volumetric flask, add an appropriate amount of methanol, perform ultrasonic treatment for 60 min, with an ultrasonic power of 250 W and an ultrasonic frequency of 40 kHz. Let it cool, make up the volume to the scale with methanol, shake well, filter, take 25 ml of the subsequent filtrate, place it in a 50-ml volumetric flask, add 5 ml of hydrochloric acid with a mass fraction of 10%, hydrolyze it in a 90°C water bath for 1 h, take it out, cool it, make up the volume with methanol, filter, and take the subsequent filtrate, that is, obtain it.

[0112] 4. Investigation of Linear Relationship

[0113] Take quercetin reference substance and kaempferol reference substance, weigh them accurately, dissolve them in 80% methanol, make up the volume to a definite value, shake well, and prepare a mother liquor of mixed reference substances containing 1026 μg of quercetin and 612.8 μg of kaempferol per 1 ml. Gradually dilute the mother liquor with 80% methanol to obtain a mixed reference solution with concentrations of quercetin 0.06412 mg / ml, 0.1282 mg / ml, 0.2565 mg / ml, 0.5130 mg / ml, 1.026 mg / ml, and kaempferol 0.0380 mg / ml, 0.0766 mg / ml, 0.1532 mg / ml, 0.3064 mg / ml, 0.6128 mg / ml. Determine the peak areas of quercetin and kaempferol according to the above chromatographic conditions. Take the reference substance concentration (X) (mg / ml) as the abscissa and the peak area integral value (Y) as the ordinate to draw a standard curve. The results are shown in Table 1. The results show that quercetin and kaempferol show a good linear relationship within their respective concentration ranges.

[0114] Table 1 Investigation of Linear Relationship of Two Active Ingredients in the Spray Preparation of Compound Propolis Composition

[0115]

[0116] 5. Precision Test

[0117] Precisely pipette 10 μl of the reference solution in "3.1", and under the chromatographic conditions in "2", inject samples continuously for 6 times, calculate the RSD of the peak areas of each peak. The results show that the RSDs of quercetin and kaempferol are 1.35% and 1.16% respectively, both less than 2, indicating good instrument precision.

[0118] 6. Repeatability Test

[0119] Accurately weigh the same sample in six parallel portions, prepare the solution according to the preparation method under "3.2", and determine it under the chromatographic conditions in item "2". Calculate the RSD of the peak areas of each peak. The results show that the RSDs of quercetin and kaempferol are 0.89% and 0.64% respectively, both less than 2, indicating that the established method in this study has good repeatability.

[0120] 7. Stability Test

[0121] Precisely pipette 10 μl of the test solution of the same sample consistent with the repeatability test, inject it at 0, 1, 3, 8, 10, and 12 h respectively to investigate the stability of the two components. The results show that the RSDs of the peak areas of quercetin and kaempferol are 1.08% and 1.26% respectively, both less than 2, indicating that the test solution has good stability within 12 h.

[0122] 8. Recovery Test

[0123] Accurately weigh six portions of the compound propolis composition of the present invention with known content of the same batch number. Adopt the recovery test, accurately add appropriate amounts of quercetin and kaempferol reference substances respectively, and accurately weigh. Prepare the test solution according to the method under "3.2", determine it under the conditions in item "2" in six parallel portions, and calculate the recovery rate. The results are shown in Tables 2 - 3.

[0124] Table 2 Recovery Test of Quercetin (n = 6)

[0125]

[0126] Table 3 Recovery Test of Kaempferol (n = 6)

[0127]

[0128] 9. Determination of Sample Content

[0129] Accurately weigh 5 ml of the compound propolis compositions of three batches (Batch 20230901, Batch 20230902, Batch 20230903) prepared in Example 1 in three parallel portions, prepare each test solution according to "3.2", respectively pipette 10 μl of each sample, inject it into the liquid chromatograph according to the law for determination. The results are shown in Table 4, and the chromatogram is as Figure 2 shown.

[0130] Table 4 Determination Results of the Content in the Sample (n = 3)

[0131]

[0132] 10. Summary: The method has good impurity separation effect, high precision and accuracy, and good reproducibility and stability, which can ensure the quality stability and effectiveness of the compound propolis composition product.

[0133] The above description is a detailed description of the preferred feasible embodiment of the present invention, but the embodiment is not intended to limit the scope of the patent application of the present invention. Any equivalent changes or modifications made under the technical spirit disclosed by the present invention shall fall within the scope of the patent covered by the present invention.

Claims

1. A compound propolis composition for preventing and treating skin chaps, characterized in that, It is prepared from the following raw materials: 1 part of propolis, 0.1 - 0.3 part of ginkgo leaves, and 0.1 - 0.3 part of bletilla striata; the preparation method of the compound propolis composition comprises the following steps: Step 1), freeze-dry the propolis and then pulverize it into fine powder of 60 - 80 meshes, put it into an ethanol solution with a volume fraction of 90 - 95% and a weight 6 - 8 times that of the total weight of the propolis, carry out ultrasonic extraction at 50 - 60 °C for 1 - 3 h to obtain a propolis ethanol solution, concentrate under reduced pressure to recover ethanol until there is no alcohol smell, and obtain concentrated solution A for standby; Step 2), put the ginkgo leaves and bletilla striata into an extraction container, carry out reflux extraction twice, each time adding an ethanol solution with a volume fraction of 50 - 70% and a weight 6 - 8 times that of the total weight of the ginkgo leaves and bletilla striata, extract for 2 - 3 hours for the first time and 1.5 - 2 hours for the second time, concentrate the filtrates of the two times under reduced pressure to recover ethanol until there is no alcohol smell, and obtain concentrated solution B for standby; Step 3), mix the concentrated solution A prepared in Step 1) above with the concentrated solution B in Step 2), or further add excipients to make a spray preparation, and thus obtain it.

2. The compound propolis composition according to claim 1, characterized in that, It is prepared from the following raw materials in parts by weight: 1 part of propolis, 0.2 part of ginkgo leaves, and 0.2 part of bletilla striata.

3. Use of the compound propolis composition according to claim 1 in the preparation of a medicament for preventing and treating skin chapping.

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