A ginsenoside Rb1 preparation with high content and high stability, and its preparation method and application

Abstract

The present invention relates to a ginsenoside Rb1 preparation with high content and high stability, and its preparation method and application. The ginsenoside Rb1 preparation includes ginsenoside Rb1, oil, emulsifier and co-emulsifier. This preparation solves the problems of poor solubility, low bioavailability and limited uses of ginsenoside Rb1. Moreover, the ginsenoside Rb1 preparation has a skin-soothing effect in an in vitro 3D skin desensitization model and can be widely applied to product forms such as functional cosmetic emulsions, aqueous solutions, sprays, etc.

Description

Technical Field

[0001] The present invention belongs to the technical field of pharmaceutical preparations, and relates to a panaxadiol preparation with high content and high stability, and a preparation method and application thereof. Background Art

[0002] Ginsenosides are the main active ingredients of ginseng, notoginseng, etc. They have attracted wide attention because of their anti-inflammatory, antioxidant, vasodilating, anti-allergic, anti-tumor and other effects. However, ginsenosides contain a large number of sugar groups, and it is difficult for the human body to directly absorb and utilize them after taking. After metabolism in the human body, the sugar groups of these saponins need to be hydrolyzed step by step to generate secondary saponins (protopanaxadiol, protopanaxatriol) or aglycones (panaxadiol, panaxatriol) with smaller molecular weights and more convenient for human absorption. Panaxadiol (PD) is a saponin aglycone containing two OH groups obtained after strong acid hydrolysis of diol-type saponins or protopanaxadiol. The purpose of hydrolysis is to improve the drug activity of saponins in the body (anti-inflammatory and soothing, anti-tumor, neuroprotection, regulation of cardiovascular and cerebrovascular). However, panaxadiol has very poor water solubility and low bioavailability, so the application of panaxadiol in various fields is relatively limited.

[0003] The 3D skin model is to obtain a small amount of living skin tissue from the body, separate the cells from the tissue, culture and amplify them in vitro, and then mix the amplified cells and biomaterials in a certain proportion and adhere the cells. The 3D skin model is formed by growing on the biomaterials. The principle of the 3D skin soothing model is to simulate and reproduce the biological reactions in the real skin during the soothing process by imitating the cells and tissue structures in the skin, so as to reflect that the active ingredients can reduce or reverse the inflammatory reaction in the model to achieve soothing (collect the skin culture solution for detecting inflammatory factors IL-1α and IL-8, and represent whether there is a soothing effect according to the relative expression level).

[0004] Chinese Patent Application CN103505462 B discloses the use of "20(S)-protopanaxadiol" in protopanaxadiol injection for the treatment of myocardial ischemia. Chinese Patent Application CN1526407A discloses a "protopanaxadiol freeze-dried emulsion and its preparation method" for intravenous targeted delivery to the lymphatic system. Chinese Patent Application CN 103800286A discloses a "20(S)-protopanaxadiol self-microemulsion composition and its preparation method and application". The self-microemulsion is composed of protopanaxadiol, non-ionic emulsifier, saturated fatty acid, etc. However, some components of the emulsifier and co-emulsifier have high cytotoxicity and are not easy to be added to cosmetics or pharmaceutical excipients. Moreover, the content of protopanaxadiol in this preparation is at most 20%, although the bioavailability is improved, there are also certain limitations in product development. Summary of the Invention

[0005] Aiming at the deficiencies of the prior art, the purpose of the present invention is to provide a ginsenoside diol preparation with high content and high stability, and its preparation method and application. The content of ginsenoside diol in the ginsenoside diol preparation of the present invention is as high as 30%, which solves the problems of low water solubility and low bioavailability of ginsenoside diol. The emulsifier used meets the requirements of the Cosmetics Technology Specification and can simultaneously meet the applications in the fields of cosmetics and pharmaceutical excipients.

[0006] To achieve the purpose of this invention, the following technical solutions are adopted:

[0007] In the first aspect, the present invention provides a ginsenoside diol preparation with high content and high stability. The raw materials for preparing the ginsenoside diol preparation include ginsenoside diol, oil, emulsifier and co-emulsifier.

[0008] The present invention mainly uses ginsenoside diol as the raw material to prepare a ginsenoside diol water-soluble preparation with high content and high stability, so as to improve the skin topical absorption and increase its bioavailability, and broaden the application of ginsenoside diol in cosmetics and pharmaceutical excipients. The ginsenoside diol preparation of the present invention has a high content of ginsenoside diol. The mass percentage content of ginsenoside diol in the preparation is above 30%. And the ginsenoside diol preparation can be dissolved in water in any proportion, clear and transparent without precipitation. After standing for 6 months, the content of ginsenoside diol hardly changes significantly, and the appearance remains clear and transparent, with good solubility and high stability. This preparation solves the problems of poor solubility, low bioavailability and limited uses of ginsenoside diol. And this ginsenoside diol preparation has a skin soothing effect in an in vitro 3D skin desensitization model and can be widely applied to product dosage forms such as functional cosmetic emulsions, aqueous solutions, sprays, etc.

[0009] Preferably, the ginsenoside diol preparation includes 10 - 40% of ginsenoside diol, 10 - 20% of oil, 30 - 40% of emulsifier and 10 - 30% of co-emulsifier by mass percentage.

[0010] The mass percentage content of the ginsenoside diol can be selected as 10%, 12%, 14%, 16%, 18%, 20%, 22%, 24%, 26%, 28%, 30%, 32%, 34%, 36%, 38%, 40%, etc. The mass percentage content of the oil can be selected as 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, etc. The mass percentage content of the emulsifier can be selected as 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, etc. The mass percentage content of the co-emulsifier can be selected as 10%, 12%, 14%, 16%, 18%, 20%, 22%, 24%, 26%, 28%, 30%, etc. Other specific point values within the above numerical ranges can be selected and will not be elaborated one by one here.

[0011] Preferably, the oil and fat includes any one or a combination of at least two of pricklyash fruit oil, camellia seed oil, olive oil or grape seed oil, and is further preferably pricklyash fruit oil.

[0012] The present invention creatively discovers that the effect of pricklyash fruit oil is superior to other oils and fats, and it can cooperate with panaxadiol to enhance its effect on soothing efficacy.

[0013] Preferably, the emulsifier includes PEG-100 stearate and / or polysorbate 60; and is further preferably PEG-100 stearate and polysorbate 60.

[0014] The present invention discovers that the addition of PEG-100 stearate and polysorbate 60 can further improve the stability of the preparation, and the emulsifier used is a combined emulsifier of non-ionic emulsifiers with an HLB value greater than 12.

[0015] Preferably, the mass ratio of PEG-100 stearate to polysorbate 60 is (2-3):1, and specific point values in (2-3) can be selected as 2, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3, etc. Other specific point values within the above numerical range can be selected and will not be elaborated here one by one.

[0016] The present invention discovers that adding the emulsifier in the above ratio can better exert the emulsifying effects of the two emulsifiers, is more conducive to transdermal absorption, enables panaxadiol to be evenly dispersed in the preparation system without precipitation, and its aqueous solution can also be clear and transparent, being more conducive to transdermal absorption.

[0017] Preferably, the co-emulsifier includes any one or a combination of at least two of glycerol, butylene glycol, pentylene glycol, polyethylene glycol or 1,2-propanediol.

[0018] In addition to increasing the emulsifying effect, the co-emulsifier can also increase the solubility of panaxadiol. Among them, glycerol has a promoting effect on reducing the particle size of the whole preparation, reducing the surface tension, and better forming microemulsion droplets.

[0019] In a second aspect, the present invention provides a preparation method of a panaxadiol preparation with high content and high stability according to the first aspect, and the preparation method includes: mixing panaxadiol with oil and fat, and then mixing with an emulsifier and a co-emulsifier to obtain the product.

[0020] The preparation method of the ginsenoside diol preparation of the present invention creatively uses a base oil to first dissolve ginsenoside diol, so that ginsenoside diol is fully wrapped in the oil phase, and then an emulsifier and a co-emulsifier are added. On the one hand, ginsenoside diol is dispersed and dissolved in the oil phase, making it easier to be emulsified by the emulsifier and improving the emulsification effect. On the other hand, using this preparation method can improve the stability of ginsenoside diol.

[0021] Preferably, the temperature of the mixing is independently 50 - 70 °C, such as 50 °C, 52 °C, 54 °C, 56 °C, 58 °C, 60 °C, 62 °C, 64 °C, 66 °C, 68 °C, 70 °C, etc. Other specific point values within the above numerical range can be selected and will not be elaborated one by one here.

[0022] In the third aspect, the present invention provides an application of the ginsenoside diol preparation according to the first aspect in the preparation of a cosmetic having a soothing effect.

[0023] Preferably, the addition amount of the ginsenoside diol preparation in the cosmetic is 0.05% - 1%, such as 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1%, etc. Other specific point values within the above numerical range can be selected and will not be elaborated one by one here.

[0024] Compared with the prior art, the present invention has the following beneficial effects:

[0025] The ginsenoside diol preparation of the present invention has a high ginsenoside diol content. The mass percentage content of ginsenoside diol in the preparation is above 30%. And the ginsenoside diol preparation can be dissolved in water in any proportion, being clear and transparent without precipitation. After being placed for 6 months, the content of ginsenoside diol hardly changes significantly, and the appearance remains clear and transparent, with good solubility and high stability. This preparation solves the problems of poor solubility, low bioavailability, and limited uses of ginsenoside diol. And this ginsenoside diol preparation has a skin soothing effect in an in vitro 3D skin desensitization model and can be widely applied to product forms such as functional cosmetic emulsions, aqueous solutions, sprays, etc. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 is the liquid chromatogram of the ginsenoside diol preparation and the ginsenoside diol standard product.

[0027] Figure 2 is the result chart of the relative expression level of IL-1α. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0028] The technical solution of the present invention will be further described below through specific embodiments. Those skilled in the art should understand that the embodiments are only for helping to understand the present invention and should not be regarded as specific limitations on the present invention.

[0029] The sources of the active ingredients contained in the products involved in the following examples and comparative examples are as follows (only the active ingredients are shown, and the necessary auxiliary ingredients contained in other commercially available raw materials are not described in detail):

[0030] Pricklyash fruit oil, olive oil, grape seed oil, camellia seed oil, PEG-100 stearate, and polysorbate-6 were all purchased from Shanghai Yuanye Bio-Technology Co., Ltd.

[0031] Example 1

[0032] This example provides a ginsenoside diol preparation. The ginsenoside diol, calculated by mass fraction, includes 1 part of ginsenoside diol with a content of 95%, 0.5 part of pricklyash fruit oil, 1 part of emulsifier, and 0.5 part of glycerol. The emulsifier is PEG-100 stearate and polysorbate-60, and their mass ratio is 2.5:1.

[0033] Its preparation method is as follows:

[0034] Mix ginsenoside diol and pricklyash fruit oil at 60 °C, then mix with the emulsifier and glycerol, and homogenize for 10 min to obtain the product.

[0035] Example 2

[0036] This example provides a ginsenoside diol preparation. The ginsenoside diol, calculated by mass fraction, includes 0.9 part of ginsenoside diol with a content of 95%, 0.4 part of pricklyash fruit oil, 1.3 parts of emulsifier, and 0.8 part of glycerol. The emulsifier is PEG-100 stearate and polysorbate-60, and their mass ratio is 3:1.

[0037] The preparation method refers to Example 1.

[0038] Example 3

[0039] This example provides a ginsenoside diol preparation. The ginsenoside diol, calculated by mass fraction, includes 1.2 parts of ginsenoside diol with a content of 95%, 0.6 part of pricklyash fruit oil, 0.9 part of emulsifier, and 0.3 part of glycerol. The emulsifier is PEG-100 stearate and polysorbate-60, and their mass ratio is 2:1.

[0040] The preparation method refers to Example 1.

[0041] Example 4

[0042] This example provides a ginsenoside diol preparation. The difference from Example 1 is only that the pricklyash fruit oil is replaced equally with camellia seed oil, and other components and their contents remain unchanged.

[0043] The preparation method refers to Example 1.

[0044] Example 5

[0045] This example provides a ginsenoside diol preparation, which is only different from Example 1 in that the pricklyash seed oil is replaced with olive oil in equal amounts, and other components and contents remain unchanged.

[0046] The preparation method refers to Example 1.

[0047] Example 6

[0048] This example provides a ginsenoside diol preparation, which is only different from Example 1 in that the pricklyash seed oil is replaced with grape seed oil in equal amounts, and other components and contents remain unchanged.

[0049] The preparation method refers to Example 1.

[0050] Example 7

[0051] This example provides a ginsenoside diol preparation, which is only different from Example 1 in that 0.5 parts of pricklyash seed oil is replaced with 0.2 parts of pricklyash seed oil, and other components and contents remain unchanged.

[0052] The preparation method refers to Example 1.

[0053] Example 8

[0054] This example provides a ginsenoside diol preparation, which is only different from Example 1 in that 0.5 parts of pricklyash seed oil is replaced with 1 part of pricklyash seed oil, and other components and contents remain unchanged.

[0055] The preparation method refers to Example 1.

[0056] Example 9

[0057] This example provides a ginsenoside diol preparation, which is only different from Example 1 in that 1 part of emulsifier is replaced with 1.5 parts of emulsifier, and other components and contents remain unchanged.

[0058] The preparation method refers to Example 1.

[0059] Example 10

[0060] This example provides a ginsenoside diol preparation, which is only different from Example 1 in that 1 part of emulsifier is replaced with 0.6 parts of emulsifier, and other components and contents remain unchanged.

[0061] Other components and contents remain unchanged.

[0062] The preparation method refers to Example 1.

[0063] Example 11

[0064] This example provides a ginsenoside diol preparation. The ginsenoside diol includes 1 part of ginsenoside diol with a content of 95% by mass, 0.5 part of pricklyash fruit oil, 1 part of emulsifier, and 0.5 part of glycerol. The emulsifier is PEG - 100 stearate and polysorbate - 60, and their mass ratio is 1:1.

[0065] The preparation method refers to Example 1.

[0066] Example 12

[0067] This example provides a ginsenoside diol preparation. The ginsenoside diol includes 1 part of ginsenoside diol with a content of 95% by mass, 0.5 part of pricklyash fruit oil, 1 part of emulsifier, and 0.5 part of glycerol. The emulsifier is PEG - 100 stearate and polysorbate - 60, and their mass ratio is 4:1.

[0068] The preparation method refers to Example 1.

[0069] Example 13

[0070] This example provides a ginsenoside diol preparation. The ginsenoside diol includes 1 part of ginsenoside diol with a content of 95% by mass, 0.5 part of pricklyash fruit oil, 1 part of emulsifier, and 0.5 part of glycerol. The emulsifier is PEG - 100 stearate.

[0071] The preparation method refers to Example 1.

[0072] Example 14

[0073] This example provides a ginsenoside diol preparation. The ginsenoside diol includes 1 part of ginsenoside diol with a content of 95% by mass, 0.5 part of pricklyash fruit oil, 1 part of emulsifier, and 0.5 part of glycerol. The emulsifier is polysorbate - 60.

[0074] The preparation method refers to Example 1.

[0075] Example 15

[0076] This example provides a ginsenoside diol preparation, and the only difference from Example 1 is the preparation method.

[0077] Its preparation method is as follows:

[0078] Mix ginsenoside diol, oil, emulsifier, and glycerol together at 60°C and homogenize for 10 min to obtain the product.

[0079] Comparative Example 1

[0080] This comparative example provides a ginsenoside diol preparation, which is only different from Example 1 in that it does not contain pricklyash seed oil, and the reduced mass is proportionally distributed to the masses of ginsenoside diol, emulsifier and glycerol.

[0081] The preparation method refers to Example 1.

[0082] Test Example 1

[0083] Stability investigation and determination of ginsenoside diol content

[0084] Test samples: Examples 1 - 15, Comparative Example 1

[0085] Test method: Place the test samples for 6 months, and detect the content of ginsenoside diol at the time points of 3 months and 6 months. Mix 100 μL of the test sample with 900 μL of deionized water, and determine the content of ginsenoside diol by ELSD - HPLC.

[0086] Chromatographic column: ZORBAX SB - C18 column (4.6×250 mm; 5 - Micron); Mobile phase: water (A) - acetonitrile (B) (0 - 5 min; 50% B - 60% B; 5 - 10 min; 60% B - 70% B; 10 - 15 min; 70% B - 75% B; 15 - 20 min; 75% B; 20 - 35 min 75% B - 80% B) Evaporation temperature: 75 °C; Atomization temperature: 30 °C; Gas flow rate: 1.8 SLM; Flow rate: 1 mL / min; Column temperature: 40 °C; Injection volume: 10 μL.

[0087] The liquid chromatograms of the ginsenoside diol preparation and the ginsenoside diol reference standard are as Figure 1 shown.

[0088] Table 1

[0089]

[0090] It can be seen from the data in Table 1 that the ginsenoside diol preparation described in the present invention has a relatively high content of ginsenoside diol. The mass percentage content of ginsenoside diol in this preparation is above 30%. After being placed for 6 months, the content of ginsenoside diol hardly changes significantly, the appearance remains clear and transparent, the solubility is good, and it has relatively high stability.

[0091] Test Example 2

[0092] Efficacy detection of in vitro reconstructed 3D epidermal model

[0093] Test samples: Example 1, Example 4 and ginsenoside diol monomer

[0094] Test method:

[0095] A blank control group, an SLS-induced stimulation group, a positive control group (dexamethasone, dex), and a sample group (the concentration of the sample group was 600 μg / mL) were set up and incubated in an incubator at 37 °C with 5% CO 2 for 2 h. Except for the blank control group, SLS was added to each well and incubated in the incubator for 24 h. The cell supernatant was collected, and the inhibitory effect on IL-1α was detected by ELISA. The data were analyzed by one-way ANOVA, and the evaluation results are shown in Table 2. Among them, Example 1 had a significant inhibitory effect on IL-1α (P < 0.001, ****), and the inhibitory effect of Example 4 on IL-1α was slightly weaker, as Figure 2 shown.

[0096] Table 2

[0097] Sample Name Efficacy Results of 3D Skin Soothing Model Example 1 Significantly inhibit the expression of IL-1α Example 4 Inhibit the expression of IL-1α Ginsenoside Rb1 Inhibit the expression of IL-1α

[0098] As can be seen from Table 2 and Figure 2 it can be known that the ginsenoside diol preparation of the present invention has a significantly excellent effect of inhibiting IL-1α. According to the relative expression levels of inflammatory factors IL-1α and IL-8, the soothing effect of the ginsenoside diol preparation can be judged, and if the expression of one of the factors is inhibited, it can be determined that the product has a soothing effect. Moreover, the ginsenoside diol preparation described in the present invention has a better soothing effect than ginsenoside diol monomer, and after the oil is replaced with other oils, the effect is equivalent to that of ginsenoside diol monomer, indicating that Prunus africana oil can cooperate with ginsenoside diol to improve the soothing effect of ginsenoside diol.

[0099] The applicant declares that the present invention uses the above embodiments to illustrate a ginsenoside diol preparation with high content and high stability, its preparation method and application, but the present invention is not limited to the above embodiments, that is, it does not mean that the present invention must rely on the above embodiments to be implemented. Those skilled in the art should understand that any improvement of the present invention, the equivalent replacement of each raw material of the product of the present invention, the addition of auxiliary components, the selection of specific methods, etc. all fall within the protection scope and disclosure scope of the present invention.

[0100] The preferred embodiments of the present invention have been described in detail above. However, the present invention is not limited to the specific details in the above embodiments. Within the technical concept of the present invention, various simple modifications can be made to the technical solution of the present invention, and these simple modifications all belong to the protection scope of the present invention.

[0101] In addition, it should be noted that in the above specific embodiments, the various specific technical features described can be combined in any appropriate manner without contradiction. To avoid unnecessary repetition, the present invention will not separately describe various possible combination methods.

Claims

1. A ginsenoside diol preparation with high content and high stability, characterized in that, the ginsenoside diol preparation comprises 10 - 40% of ginsenoside diol, 10 - 20% of oil, 30 - 40% of emulsifier and 10 - 30% of co-emulsifier by mass percentage; the oil is any one or a combination of at least two of pricklyash fruit oil, olive oil or grape seed oil; the emulsifier is PEG-100 stearate and polysorbate 60 with a mass ratio of (2 - 3):1; the ginsenoside diol preparation is obtained by the following preparation method: mixing ginsenoside diol with oil, and then mixing with emulsifier and co-emulsifier.

2. The ginsenoside diol preparation with high content and high stability according to claim 1, characterized in that, the co-emulsifier comprises any one or a combination of at least two of glycerol, butanediol, pentanediol, polyethylene glycol or 1,2-propanediol.

3. The preparation method of the ginsenoside diol preparation with high content and high stability according to any one of claims 1 - 2, characterized in that, the preparation method is: mixing ginsenoside diol with oil, and then mixing with emulsifier and co-emulsifier.

4. The preparation method of the ginsenoside diol preparation with high content and high stability according to claim 3, characterized in that, the temperature of the mixing is independently 50 - 70°C.

5. The application of the ginsenoside diol preparation according to any one of claims 1 - 2 in the preparation of a cosmetic with a soothing effect.

6. The application according to claim 5, characterized in that, the addition amount of the ginsenoside diol preparation in the cosmetic is 0.05 - 1%.

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