Composition capable of carrying rare ginsenoside and preparation method and application thereof

Abstract

The invention relates to a composition capable of carrying rare ginsenoside and a preparation method and application thereof, and belongs to the technical field of daily necessities. The composition capable of carrying the rare ginsenoside comprises the following components in parts by weight: 3-8 parts of peony seed oil, 3-5 parts of solid wax and 1-5 parts of a peony fermentation hydrogen donor, the solid wax comprises at least one of peony unsaponifiable matter and vegetable wax ester; the peony fermentation hydrogen donor comprises peony fermentation glycerol. The peony fermented essential oil is adopted as a hydrogen donor and is compounded with the peony seed oil and the solid wax, grease can be modified, the obtained composition has a space structure carrying non-oil-soluble components, the non-oil-soluble components such as rare ginsenoside can be introduced into a full-oil system, and the composition has the advantages of being excellent in stability, free of toxic and side effects and the like. Therefore, the stability and the utilization rate of non-oil-soluble components are effectively improved.

Description

Technical Field

[0001] The invention belongs to the technical field of daily necessities, and particularly relates to a composition capable of carrying rare ginsenosides, a preparation method and application thereof. Background Art

[0002] The market is hot with various Chinese herbal beverages, such as ginseng, rhodiola rosea, ginseng antlers, etc. These Chinese herbal medicines are metabolized and absorbed in the intestines after oral administration, and then the component structure will be transformed into highly active ingredients. However, this in vivo conversion is subject to individual differences in the human body and multiple conditions, and the conversion efficiency is low. For example, the average absorption rate of ginseng prototype saponins is 2.5%, and the absorption rate of Rb1 is 1.4%. Therefore, the expected functions of Chinese herbal medicines cannot be achieved by non-specifically supplementing the highly active ingredients of Chinese herbal medicines. The market is more focused on the research of ingredients and separation and purification methods for ginseng, and less on the research of its delivery system.

[0003] Rare ginsenosides contain the structure of sugar and aglycone. Aglycones have different degrees of lipophilicity, while sugar chains have strong hydrophilicity, which makes the saponins have certain surface active functions. Rare ginsenosides are certain metabolic derivatives and transformed derivatives obtained after metabolic transformation of prototype ginsenosides (tetracyclic triterpenoid compounds), which means that the molecular weight of rare ginsenosides is smaller than that of prototype ginsenosides, and their pharmacological activity is far superior to that of prototype ginsenosides. Although rare ginsenosides contain polar groups, it is difficult for them to fully form stable hydrogen bonds and other interactions with water molecules, which leads to their poor water solubility; and because they cannot form a stable state with oil molecules in oil, their oil solubility is also poor.

[0004] Therefore, stable oil-soluble rare ginsenosides are relatively scarce, and it is of great significance to develop a product that can stably carry rare ginsenosides. Summary of the invention

[0005] The purpose of the present invention is to overcome the problems existing in the above-mentioned prior art and to provide a composition capable of carrying rare ginsenosides and a preparation method and application thereof.

[0006] The present invention is achieved through the following technical solutions:

[0007] In a first aspect, the present invention provides a composition capable of carrying rare ginsenosides, comprising the following components by weight: 3 to 8 parts of peony seed oil, 3 to 5 parts of solid wax, and 1 to 5 parts of peony fermentation hydrogen donor; the solid wax comprises at least one of peony unsaponifiable matter and plant wax esters; the peony fermentation hydrogen donor comprises peony fermentation glycerol.

[0008] The present invention adopts peony fermented essential oil as a hydrogen donor, and compounded with peony seed oil and solid wax, can modify oil esters, and the obtained composition has a spatial structure carrying non-oil-soluble components, can introduce non-oil-soluble components such as rare ginsenosides into a full oil system, and has excellent stability, thereby effectively improving the stability and utilization rate of the non-oil-soluble components.

[0009] Preferably, the peony fermentation hydrogen donor further comprises a supramolecular peony solution, and the mass ratio of the peony fermentation glycerol to the peony fermentation hydrogen donor is (1-3): (1-3).

[0010] More preferably, the mass ratio of the peony fermented glycerol to the peony fermented hydrogen donor is 3:1.

[0011] Preferably, the mass ratio of the peony seed oil, the solid wax and the peony fermentation hydrogen donor is 2:2:1.

[0012] Preferably, the peony fermented glycerol is prepared by fermenting the whole peony plant with glycerol-producing Candida.

[0013] Preferably, the method for preparing peony fermented glycerol comprises the following steps: treating the whole peony plant with cellulase, sterilizing by high pressure to obtain peony fermentation medium, fermenting with glycerol-producing Candida as the fermentation strain, taking the supernatant, and obtaining the peony fermented glycerol after decolorization and sterilization.

[0014] The DPPH free radical scavenging rate, hydroxyl free radical scavenging rate and tyrosinase inhibition rate of the peony fermented glycerol of the present invention are 76.2±0.5%, 82.9±0.5% and 96.3±0.5% respectively, which is significantly better than non-fermented glycerol and glycerol fermented without plant substrate. The peony fermented glycerol of the present invention as a hydrogen donor can make the composition have better structural stability and antioxidant properties.

[0015] Preferably, the temperature of the cellulase treatment is 48°C-52°C, and the time is 30h-40h.

[0016] Preferably, the weight of the cellulase is 0.08%-0.12% of the weight of the whole peony plant.

[0017] Preferably, the fermentation conditions are: 224.6 g·L -1 -235.4g·L -1 Glucose, speed 340r / min-560r / min, ventilation volume 0.8vvm-1vvm.

[0018] Preferably, the fermentation temperature is 25°C-35°C, and the fermentation time is 75h-85h.

[0019] Preferably, activated carbon is used for decolorization, and the decolorization time is 14 min-16 min.

[0020] In a specific embodiment of the present invention, the specific operation of the fermentation is: pick a ring of glycerol-producing Candida strains from the freshly transferred cultured surface, inoculate it into a 500mL shake flask (the seed culture medium liquid volume is 50mL), shake culture it in a reciprocating shaker at 30°C for 19h (amplitude 10a frequency 110 times min), and then inoculate the shake flask seed liquid into a 5L fermentation tank at an inoculation rate of 6%, the peony fermentation medium liquid volume is 3L, 230.0g·L -1 Glucose, speed 550rmin, temperature 31℃, ventilation 0.9vvm, natural pH.

[0021] Specifically, the method of treating the whole peony plant with cellulase is as follows: remove impurities from the whole peony plant, crush it, add water to boil it, add cellulase to treat it after cooling, filter it, and sterilize it under high pressure to obtain the peony fermentation medium.

[0022] The parameters in the preparation of peony fermented glycerol will affect the yield and antioxidant properties of the fermented glycerol. Under the above preparation parameters and conditions, the peony fermented glycerol can have better performance.

[0023] Preferably, the method for preparing peony seed oil and peony unsaponifiable matter comprises the following steps:

[0024] (1) extracting the pretreated peony seeds with supercritical CO2 fluid to obtain peony seed oil;

[0025] (2) molecularly distilling the peony seed oil obtained in step (1), recovering the light phase fraction residue to obtain the peony seed oil unsaponifiable matter.

[0026] Preferably, in the step (1), the method for pretreating the peony seeds is: drying at a constant temperature of 58°C for 24 hours, crushing and sieving, and the crushing particle size is 20 mesh; the conditions for the supercritical CO2 fluid extraction are: extraction pressure 28mPa-32mPa, extraction temperature 37°C-43°C, carbon dioxide flow rate 30kg / h, extraction time 2h, separation temperature 30°C-40°C, and separation pressure 10.5MPa-11.5MPa.

[0027] Preferably, in step (2), the molecular distillation method is: distillation pressure is 0.018 mm Hg-0.022 mm Hg, distillation temperature is 215° C.-225° C., scraping speed is 250 r / min-290 r / min, and distillation time is 24 h.

[0028] Preferably, the method for preparing the supramolecular peony solution comprises the following steps:

[0029] S1. The peony seed meal is pretreated and fermented with Lactobacillus to obtain an initial fermentation liquid;

[0030] S2. The initial fermentation liquid obtained in step S1 is separated by acid addition, and filtered and sterilized to obtain a supramolecular peony solution.

[0031] Preferably, in step S1, the method for pretreating the peony seed meal is: removing impurities and then crushing.

[0032] Preferably, in step S1, the culture medium for fermentation culture is a supramolecular NaDES solvent, glucose, urea and ultrapure water in a ratio of 2:2:1:15; and the fermentation culture time is 10 days.

[0033] Preferably, the supramolecular NaDES solvent is prepared by heating lactobionic acid, fructose, sorbitol and water in a molar ratio of 1:1:1:5 at 46°C-50°C for 2.5-3.5h and then drying at 58°C-62°C for 71h-73h.

[0034] In a second aspect, the present invention provides a method for preparing the composition capable of carrying rare ginsenosides, comprising the following steps: uniformly mixing peony seed oil, solid wax, and peony fermentation hydrogen donor, adding an alkaline catalyst for catalytic reaction, neutralizing the catalyst after the reaction, and filtering to obtain the composition capable of carrying rare ginsenosides.

[0035] Preferably, the catalyst is an alkaline catalyst, such as an alkaline or alkaline earth hydroxide or alkoxy compound, specifically caustic soda and potassium salts, calcium hydroxide, potassium carbonate or sodium carbonate in alcohol solution, aqueous solution or solid form. The temperature of the catalytic reaction is 200±10°C, and the catalytic reaction is carried out for 8-10 hours under nitrogen inert conditions. At the end of the temperature stabilization period, the catalyst is neutralized with a sufficient amount of acid (such as citric acid, phosphoric acid, etc.), and then filtered to obtain the composition that can carry rare ginsenosides.

[0036] The composition obtained by the ester catalytic reaction of the present invention has a simple preparation method, and the obtained composition has an amphiphilic property and can be used to stabilize non-oil-soluble active ingredients.

[0037] In a third aspect, the present invention provides the use of the composition capable of carrying rare ginsenosides in cosmetics and / or the preparation of health products and / or medicines.

[0038] The composition of the present invention has the effect of carrying non-oil-soluble active ingredients, can be used to stably carry non-oil-soluble active ingredients, and has broad application prospects in health products and medicines.

[0039] In a fourth aspect, the present invention provides a peony gel oil containing rare ginsenosides, comprising the following components in parts by weight: 0.01 to 0.1 parts of rare ginsenosides, 0.1 to 1 parts of the composition capable of carrying rare ginsenosides, and 7 to 9 parts of peony seed oil.

[0040] The peony gel oil of the present invention carries rare ginsenosides in the above-mentioned composition. Since the rare ginsenosides have strong antioxidant capacity, they synergistically stabilize the stability of the peony seed oil and effectively extend the shelf life of the gel oil. The peony gel oil of the present invention has excellent immunity enhancement, anti-inflammatory, anti-allergic and antioxidant effects.

[0041] The present invention does not limit the type of peony seed oil, and it can be peony seed oil obtained by various extraction processes, such as peony seed oil extracted by supercritical fluid, cold-pressed peony seed oil and water-enzyme peony seed oil. Preferably, the peony seed oil is obtained by supercritical fluid extraction.

[0042] In a fifth aspect, the present invention provides the use of the peony gel oil containing rare ginsenosides in the preparation of health products and / or medicines.

[0043] The peony gel oil of the invention contains rare ginsenosides, has excellent immunity enhancement, anti-inflammatory, anti-allergic and anti-oxidation effects, and can be applied to the preparation of health products and medicines.

[0044] Compared with the prior art, the present invention has the following beneficial effects: due to the structural characteristics, the water solubility and oil solubility of rare ginsenosides are poor, and the efficacy of rare ginsenosides cannot be effectively exerted at present. The present invention adopts peony fermented essential oil as a hydrogen donor, compounded with peony seed oil and solid wax, and can modify the oil ester. The obtained composition has a spatial structure carrying rare ginsenosides, can introduce rare ginsenosides into a full oil system, and has excellent stability. The peony gel oil of the present invention contains rare ginsenosides, synergistically stabilizes the stability of peony seed oil, effectively improves the stability of the gel oil, and the peony gel oil of the present invention has excellent immunity enhancement, anti-inflammatory, anti-allergic and antioxidant effects. BRIEF DESCRIPTION OF THE DRAWINGS

[0045] Figure 1 It is the 24h status of the blank group in the stability test;

[0046] Figure 2 The state of comparative example 8 after being placed at 40° C. for 3 months in the stability test;

[0047] Figure 3 This is the anti-allergic test result diagram of the blank group in the anti-allergic test;

[0048] Figure 4 This is the anti-allergic test result diagram of the model group in the anti-allergic test;

[0049] Figure 5 This is a graph showing the anti-allergic test results of the group using the peony gel oil of Example 1 at a dosage of 1.5 μmol / L;

[0050] Figure 6 This is a graph showing the anti-allergic test results of the group using the peony gel oil of Example 1 at a dosage of 0.6 μmol / L;

[0051] Figure 7 This is a representative graph of the test results of the model control group in the neutrophil aggregation inhibition test in the anti-inflammatory test;

[0052] Figure 8 This is a representative graph of the test results of the positive control group in the neutrophil aggregation inhibition test in the anti-inflammatory test;

[0053] Fig. 9 This is a representative graph of the test results of the peony gel oil group of Example 1 in the anti-inflammatory test for inhibiting neutrophil aggregation. DETAILED DESCRIPTION

[0054] To better illustrate the purpose, technical solutions and advantages of the present invention, the present invention will be further described below in conjunction with specific embodiments. It should be understood by those skilled in the art that the specific embodiments described herein are only used to explain the present invention and are not intended to limit the present invention.

[0055] Unless otherwise specified, the experimental methods used in the examples are all conventional methods; the materials, reagents, etc. used, unless otherwise specified, can be obtained from commercial channels.

[0056] Examples 1-6

[0057] The components and weights of the compositions capable of carrying rare ginsenosides of Examples 1-6 are shown in Table 1.

[0058] Table 1 Formula table of Examples 1-6

[0059] Components Example 1 Example 2 Example 3 Example 4 Example 5 Example 6 Peony seed oil(g) 400 500 300 440 400 400 Peony unsaponifiable matter (g) 400 300 300 440 300 400 Peony fermented glycerin (g) 150 100 100 90 225 200 Supramolecular peony solution (g) 50 100 300 30 75 0

[0060] The preparation method of the composition capable of carrying rare ginsenosides of Examples 1-6 comprises the following steps: peony seed oil, peony unsaponifiable matter (solid wax), peony fermented glycerol and supramolecular peony solution (peony fermented hydrogen donor) are mixed according to the weights shown in Table 1, 1000 g of the mixture is introduced into a 2L stirred reactor, 20 g of alkali is added as a catalyst, the reaction is carried out at 200 degrees Celsius for eight hours under nitrogen inert conditions, at the end of the temperature stabilization period, the catalyst is neutralized with sufficient acid, and then filtered, and the obtained wax derivative is the composition capable of carrying rare ginsenosides of Examples 1-6.

[0061] The preparation method of the peony seed oil and peony unsaponifiable matter comprises the following steps:

[0062] (1) After drying the peony seeds at a constant temperature of 58° C. for 24 hours, crushing and sieving (crushing particle size 20 mesh), using supercritical CO2 fluid to extract peony seed oil, wherein the extraction pressure is set to 30.0 mPa, the extraction temperature is set to 40° C., the carbon dioxide flow rate is 30 L / h, the extraction time is set to 2 h, the separation temperature is set to 35° C., and the separation pressure is set to 11.0 MPa, to obtain peony seed oil;

[0063] (2) setting the distillation pressure of the peony seed oil obtained in step (1) to 0.02 mm Hg, the distillation temperature to 220° C., the scraping speed to 270 r / min, and recovering the light phase fraction after 24 hours to obtain the peony seed oil unsaponifiable matter.

[0064] The method for preparing peony fermented glycerol comprises the following steps: removing impurities from the whole peony plant and then crushing it, adding water to steam for 10 minutes, adding 0.1% (w / w) cellulase at 50°C for 36 hours after cooling; cooling and filtering, and sterilizing by high pressure to obtain peony fermentation medium;

[0065] A ring of glycerol-producing Candida strain was picked from the freshly transferred slant and inoculated into a 500 mL shake flask (the volume of seed culture medium was 50 mL), and cultured in a reciprocating shaker at 30°C for 19 h (amplitude 10a frequency 110 times / min), and then the shake flask seed liquid was inoculated into a 5 L fermentation tank (Korea Co. LKF-5L) at a 6% inoculation rate, and the volume of peony fermentation medium was 3 L, and the 230.0 g·L -1 Glucose, a speed of 350r / min, a temperature of 31°C, and a ventilation volume of 0.9vvm were fermented for 80h, and the supernatant was taken, decolorized with activated carbon for 15min and then sterilized to obtain the peony fermented glycerol.

[0066] The method for preparing the supramolecular peony solution comprises the following steps:

[0067] S1. The molar ratio of lactobionic acid, fructose, sorbitol and water is 1:1:1:5, heated at 48°C for 3h and then dried at 60°C for 72h to obtain supramolecular NaDES solvent, and the supramolecular NaDES solvent, glucose, urea and ultrapure water are mixed at a ratio of 2:2:1:15 to obtain fermentation broth. Peony seed meal is crushed after removing impurities, added with lactobacillus and fermented in the aforementioned fermentation broth for 10 days to obtain initial fermentation broth;

[0068] S2. The initial fermentation liquid obtained in step S1 is separated by acid addition, and filtered and sterilized to obtain a supramolecular peony solution.

[0069] Comparative Examples 1-8

[0070] The difference between the compositions carrying rare ginsenosides of Comparative Examples 1-8 and the examples is only in the components and weight parts, as shown in Table 2.

[0071] The difference between the preparation method of peony fermented glycerol and the preparation method of peony fermented glycerol is that the whole peony plant is used to replace the whole peony plant, and the other preparation parameters are the same as those of the preparation method of peony fermented glycerol to obtain peony fermented glycerol.

[0072] Unless otherwise specified, the components and raw materials used in the embodiments and comparative examples of the present invention are all commercially available raw materials, and the components and raw materials used in each parallel experiment are all of the same kind.

[0073] Table 2 Formula table of comparative examples 1-8

[0074] Components Comparative Example 1 Comparative Example 2 Comparative Example 3 Comparative Example 4 Comparative Example 5 Comparative Example 6 Comparative Example 7 Comparative Example 8 Peony seed oil(g) 800 0 500 100 400 400 400 400 Peony unsaponifiable matter (g) 0 800 500 800 400 400 400 400 Peony fermented glycerin (g) 150 150 0 75 0 0 0 0 Supramolecular peony solution (g) 50 50 0 25 0 100 0 200 Polyglycerol-10(g) 0 0 0 0 0 100 0 0 Peony fermented glycerin (g) 0 0 0 0 0 0 200 0 Glycerin(g) 0 0 0 0 200 0 0 0

[0075] The compositions carrying rare ginsenosides obtained in the examples and comparative examples were tested, and the testing method was as follows:

[0076] 1. Recovery amount: refers to the amount of wax derivatives obtained by filtration, that is, the amount of the composition that can carry rare ginsenosides;

[0077] 2. Water absorption multiple: refers to the ratio of the maximum amount of water that the composition carrying rare ginsenosides can absorb to its own weight, indicating that the composition has a certain hydrogen supply capacity;

[0078] 3. DPPH free radical scavenging rate: Set up T, T0, C, and C0 tubes, add 50.0 μL of sample (5% concentration) and 100.0 μL of water to T and T0 tubes, mix well, add 150.0 μL of water to C and C0 tubes, and add 50.0 μL of DPPH solution (0.12 mg mL -1 ), add 50.0L ethanol solution (95%, v / v) to T0 and C0 tubes, mix well, incubate at 30℃ for 5min, and measure absorbance at 517nm;

[0079] The calculation formula of DPPH free radical scavenging rate is: R% = (1-[A i -A j ] / A c )×100%, where Ac is the absorbance value of C tube, Ai is the absorbance value of T tube; Aj is the control, which is the absorbance value of T0 tube;.

[0080] 4. Hydroxyl radical scavenging rate: Set T, T O , C tube, add 30.0μL sample (use water in C tube instead of sample solution), 30.0μL FeSO4 solution (8.0mmo1·L-1 ), 100.0 μL salicylic acid (3.0 mmol·L -1 ) and 100.0 μL H2O2 (8.8 mmol·L -1 ) After mixing, incubate at 37℃ for 10min, and measure absorbance at 510nm. The T and C tubes are the average of three sets of data.

[0081] Among them, hydroxyl radical scavenging rate (%) = [A0-(A x -A x0 ) / A0]·100, where A0 is the absorbance value of the blank control, A x is the absorbance of the added sample, A x0 No color developing agent H2O2 is added.

[0082] The test results of the embodiments and comparative examples are shown in Table 3 and Table 4.

[0083] Table 3 Test results of the embodiment

[0084] Example 1 Example 2 Example 3 Example 4 Example 5 Example 6 Recycling amount (g) 846 798 679 700 668 756 Water absorption multiple 4.5 4.1 4.3 3.9 4.2 4.0 DPPH free radical scavenging rate% 87.7 82.5 86.2 89.6 88.9 81.1 Hydroxyl free radical scavenging rate% 79.1 75.3 77.3 67.3 65.7 74.4

[0085] Table 4 Test results of comparative examples

[0086]

[0087] As can be seen from Table 3 and Table 4, Example 1 has the best performance. In Comparative Examples 1-3, the lack of a certain component will lead to a decrease in the performance of the composition. In Comparative Example 4, the amount of component added is not good, resulting in the overall weakening of the performance of the composition. It can be seen from the results of Comparative Examples 5-6 compared with Example 6 that the additional addition of a non-fermentation-derived hydrogen donor (such as non-fermentation-derived ordinary glycerol, non-fermented polyglycerol-10) will have a relatively large impact on the antioxidant properties of the overall composition. The performance of Comparative Example 8, which only uses a peony supramolecular solution, is also lower than that of Example 6, indicating that the addition of peony fermented glycerol to the composition of the present invention can effectively improve the performance of the composition.

[0088] Comparison between Comparative Example 7 and Example 6 shows that if peony is selected as another substrate for fermentation of glycerol, its antioxidant performance is lower than that of Example 6 using peony as the substrate. In addition, the selection of peony as the substrate for fermentation of glycerol can also optimize the whole chain production of peony, embody the concept of zero waste, and optimize and redevelop the by-products.

[0089] Application Examples 1-6 and Application Comparative Examples 1-8

[0090] A peony gel oil containing rare ginsenosides comprises: 0.1 parts of rare ginsenoside RG3, 0.9 parts of the composition capable of carrying rare ginsenosides of Examples 1-6 and Comparative Examples 1-8, and 9 parts of supercritical peony seed oil.

[0091] The preparation method is as follows: evenly mix the rare ginsenoside RG3 with the composition capable of carrying rare ginsenosides of Examples 1-6 and Comparative Examples 1-8, and then evenly mix with supercritical peony seed oil to obtain the peony gel oil containing rare ginsenosides.

[0092] The supercritical peony seed oil is the peony seed oil obtained by supercritical fluid extraction and used in the composition of the aforementioned embodiment.

[0093] Application Example 7

[0094] A peony gel oil containing rare ginsenosides comprises: 0.1 parts of rare ginsenoside CK, 0.9 parts of the composition capable of carrying rare ginsenosides of embodiment 1, and 9 parts of supercritical peony seed oil.

[0095] The difference between the preparation method and Application Example 1 is that rare ginsenoside CK is used to replace rare ginsenoside RG3, and the rest of the preparation method is the same as Application Example 1.

[0096] Application Example 8

[0097] A peony gel oil containing rare ginsenosides comprises: 0.1 parts of rare ginsenoside RG3, 0.9 parts of the composition capable of carrying rare ginsenosides of embodiment 1, and 9 parts of cold-pressed peony seed oil.

[0098] The preparation method is as follows: evenly mix the rare ginsenoside RG3 and the composition capable of carrying rare ginsenosides in Example 1, and then evenly mix them with cold-pressed peony seed oil to obtain the peony gel oil containing the rare ginsenosides.

[0099] The preparation method of the cold-pressed peony seed oil comprises the following steps: putting peony seeds for oil into a 50°C oven and drying them to less than 10% (refer to GB / T14489.1-2008), storing them in a 4°C refrigerator for standby use, selecting peony seeds with similar particle sizes, putting them into liquid cold pressing for oil production, controlling the pressure to 50Mpa and the temperature to 50°C for pressing for 2 hours, centrifuging the pressed peony seed oil in a 4°C centrifuge at a speed of 5000r / s for 5 minutes, and taking the supernatant to obtain the peony seed oil; and pressing the peony seed oil before use.

[0100] Application Example 9

[0101] A peony gel oil containing rare ginsenosides comprises: 0.1 parts of rare ginsenoside RG3, 0.9 parts of the composition capable of carrying rare ginsenosides of Example 1, and 9 parts of peony seed oil prepared by aqueous enzymatic method.

[0102] The preparation method is as follows: evenly mix the rare ginsenoside RG3 and the composition capable of carrying rare ginsenosides of Example 1, and then evenly mix them with the aqueous enzymatic peony seed oil to obtain the peony gel oil containing the rare ginsenosides.

[0103] The preparation method of the peony seed oil by aqueous enzymatic method comprises the following steps: drying, crushing and sieving peony seeds to obtain peony seed powder, accurately weighing 30 g of the pretreated peony seed powder into a 500 mL beaker, adding ultrapure water at a material-liquid ratio of 1:7.5 (w / v), ultrasonicating in an ultrasonic cleaner for 5 minutes, adjusting the pH value of the system to 9.0 with HCl or NaOH, adding 2% alkaline protease (Alcalase 2.4L) at 57° C., performing enzymolysis for 150 minutes in a water bath constant temperature oscillator, heating to 90° C. to inactivate the enzyme for 15 minutes after the enzymolysis, then centrifuging at 4500 r / min for 15 minutes, allowing the oil and water layers to stand in a separatory funnel for 2 minutes to separate the peony seed oil, centrifuging again at 5000 r / min for 10 minutes, collecting the upper clear oil and drying it, and obtaining the peony seed oil by aqueous enzymatic method.

[0104] Application Comparative Example 9

[0105] A peony gel oil comprises: 0.9 parts of the composition capable of carrying rare ginsenosides of embodiment 1 and 9 parts of supercritical peony seed oil.

[0106] The preparation method is as follows: the composition capable of carrying rare ginsenosides in Example 1 is uniformly mixed with supercritical peony seed oil to obtain the peony gel oil containing rare ginsenosides.

[0107] The supercritical peony seed oil is the peony seed oil obtained by supercritical fluid extraction and used in the composition of the aforementioned embodiment.

[0108] The stability of the peony gel oil of application examples 1-7 and application comparative examples 1-8 was tested, and the blank group was: 0.1 parts of rare ginsenoside RG3 and 9.9 parts of supercritical peony seed oil; that is, the blank group did not contain a composition that can carry rare ginsenosides. The test results are shown in Tables 5 and Figure 1 , Figure 2 As shown, Figure 1 For the blank group 24h status, Figure 2 This is the state of Comparative Example 8 after being placed at 40°C for 3 months.

[0109] Table 5 Stability test results of peony gel oil of application examples 1-7 and application comparative examples 1-8

[0110]

[0111]

[0112] From the results of using Examples 1-6 and Comparative Examples 1-5 in Table 5, the peony gel oil obtained by the non-optimal ratio of the composition that can carry rare ginsenosides can dissolve the rare ginsenosides in the peony seed oil at the beginning, but after a short period of heat stability test, RG3 precipitation occurs and it cannot be stabilized. This may be due to the low water absorption performance of the comparative composition, the weak hydrogen supply capacity, and the inability to form a stable hydrogen bond structure between the hydroxyl groups of the rare ginsenosides. Therefore, for Comparative Example 8, which has poor water absorption performance itself, the 3-month heat resistance test phenomenon further confirms this conclusion ( Figure 2 ). From the blank results, it can be seen that without the addition of the composition of the present invention, RG3 is indeed unable to dissolve in peony seed oil ( Figure 1 ), this is because the composition of the present invention is a wax ester derivative obtained after esterification, and the esters therein are amphiphilic. Due to the presence of such amphiphilic molecules, it can be miscible with the rare ginsenoside RG3 and has the ability to disperse it into oil. It can be seen from Application Example 7 that the composition of the present invention also has excellent dispersion stability for rare ginsenosides of other configurations.

[0113] The performance test of the peony gel oil of application examples 1, 8-9 and application comparative examples 6 and 8-9 was carried out, and the test method was as follows:

[0114] 1. Test POV value: store peony gel oil, supercritical peony seed oil and cold-pressed peony seed oil in a 60°C oven for 18 days, and test the POV value of the sample every 3 days to examine the changes in POV values ​​of peony seed oil and peony gel oil under accelerated conditions. The peroxide value determination method refers to GB 5009.227-2016 Determination of Peroxide Value in Food.

[0115] 2. Test TBA value: Place peony gel oil and peony seed oil crude oil in a 60°C oven for 18 days, measure the TBA value of the sample every 3 days, and examine the changes in TBA values ​​of peony seed oil and peony gel oil under accelerated oxidation conditions.

[0116] The TBA value was determined by a direct method: 50-100 mg (accurate to 0.1 mg) of sample was weighed into a 25 mL volumetric flask, diluted with n-butanol, 5 mL of the diluted sample was mixed with TBA solution (TBA solution: 200 mg TBA was placed in a 100 mL volumetric flask, diluted with n-butanol and stored at room temperature for 12 h before filtration, the filtrate was made up to 100 mL with n-butanol and stored at 4 °C for later use) in a stoppered test tube and heated in a 95 °C water bath for 2 h, cooled to room temperature for 10 min, and the absorbance was measured at 530 nm.

[0117] The test results are shown in Table 6.

[0118] Table 6 Performance test results of peony gel oil of application examples 1, 8-9 and application comparative examples 6, 8-9

[0119]

[0120] It can be seen from Table 6 that under room temperature conditions, the shelf life of supercritical peony seed oil is 80 days, while the shelf life of peony seed oil after adding peony gel oil containing rare ginsenosides is 272 days, indicating that after adding peony gel oil containing rare ginsenosides, the stability and oxidation resistance of the oil are improved, and the shelf life of the product is extended.

[0121] The peony gel oil of Application Example 1 was tested for anti-allergy and anti-inflammatory properties, and the testing method was as follows:

[0122] 1. Anti-allergic test - inhibiting extracellular Ca2+ in mast cells 2+ internal flow

[0123] In the early stage of allergic reaction, the receptors on mast cells bind to a large amount of IgE antibodies in the body, bridging the molecules on the cell surface, causing changes in the cell membrane structure and extracellular Ca 2+ Influx, intracellular Ca 2+ accumulation, activating the tyrosine phosphorylation pathway, leading to mast cell degranulation. 2+ Influx of intracellular Ca 2+ Increased concentration is an important sign of mast cell degranulation, inhibiting intracellular Ca 2+ The increase in concentration is one aspect of screening anti-allergic drugs. The fluorescent probe Fluo 3-AM used in the experiment is a lipophilic compound that can penetrate the cell membrane into the cell and be hydrolyzed into free Fluo3 molecules. Fluo3 molecules can bind to free intracellular Ca 2+ , produces fluorescence at 490-500nm, and the fluorescence intensity of the cells is related to the intracellular Ca 2+ The concentration was positively correlated.

[0124] Test method:

[0125] 1) Cell seeding: Keratinocytes were seeded into 6-well plates at a seeding density of 18×10 cells / well and incubated overnight in an incubator (37°C, 5% CO).

[0126] 2) Solution preparation: Prepare the working solutions of the test substances according to the test groups in Table 7 below.

[0127] Table 7

[0128]

[0129] 3) Probe loading: Aspirate the keratinocyte culture medium, wash 3 times with PBS, add 0.5 mL of probe loading working solution (calcium ion fluorescent Flu03-AM probe: Japan Tongren / F026, 2uM) to each well, and incubate at 37C for 45 minutes. After the incubation, aspirate the probe working solution and wash 3 times with PBS.

[0130] 4) Stimulation and administration: 0.5 mL of PBS solution was added to each well of the blank group, 0.05 μM capsaicin CAP was added to each well of the model group, and 0.05 uM capsaicin CAP and PBS solution corresponding to the concentration of Example 1 were added to each well of the sample group. After administration, the cells were incubated in an incubator for 1 h.

[0131] 5) Calcium flow detection: After incubation, observe and take pictures using a fluorescence microscope (excitation wavelength: 460nm-550nm, emission wavelength: 590nm), and collect cell morphology under a 200x field of view. The photography time is controlled within 30 minutes.

[0132] The test results are shown in Figure 3-Figure 6 Among them Figure 3 For the blank group, Figure 4 For the model group, Figure 5 The peony gel oil of Example 1 was used in an amount of 1.5 μmol / L. Figure 6 The peony gel oil used in Example 1 is used in an amount of 0.6 μmol / L.

[0133] from Figure 3-Figure 6 It can be seen that compared with the control group, the fluorescence intensity of the cells in the model group was significantly enhanced, and the fluorescence intensity of the sample group gradually weakened with the increase of concentration, indicating that the peony gel oil containing rare ginsenosides can inhibit the intracellular Ca 2+ The concentration of β-actin increases, thereby inhibiting mast cell degranulation.

[0134] 2. Anti-inflammatory test

[0135] 2.1 Anti-inflammatory test - inhibiting the expression of macrophage inflammatory factors

[0136] Inflammation is one of the more common diseases. It is the basic pathological process of many diseases and also a defensive response of the body for self-protection. Inflammation is closely related to the occurrence and development of many diseases, such as nephritis, hepatitis, and cancer. The main manifestations of inflammation are edema, fever, redness of wounds, and even chills, coma, etc. LPS (lipopolysaccharide) is an endotoxin that can show a variety of biological activities by acting on human or animal cells. Therefore, the in vitro inflammation model of mouse mononuclear macrophage RAW2647 cells induced by LPS is often used to evaluate the anti-inflammatory activity of drugs.

[0137] Experimental protocol (in-vitro)

[0138] Cells: Mouse macrophage cell line Raw 264.7

[0139] Sample: Peony Gel Oil of Application Example 1

[0140] Irritant: Lipopolysaccharide (LPS), one of the main components of the cell wall of Gram-negative bacteria.

[0141] Treatment method: Determine the safe concentration range of the peony gel oil in Application Example 1 on Raw264.7 cells, ② 1 μg / mL LPS and 0.3% peony gel oil in Application Example 1 co-treat Raw264.7 cells for 24 hours, and use neutral red NR to detect cell viability. Collect the supernatant, and detect the expression levels of IL-6, IL-8, PGE-2, TNF-α, and COX-2 by ELISA.

[0142] The test results are shown in Table 8.

[0143] Table 8 Anti-inflammatory test results

[0144] Inflammatory factor inhibition rate (%) IL-6 IL-8 PGE-2 TNF-α COX-2 0.3% Example 3-1 69.8 99.5 96.0 88.21 83.68

[0145] As can be seen from the above table, the strong inhibition of anti-inflammatory activity proves that the present invention has a strong anti-inflammatory activity, suggesting that it has certain medicinal and health care value.

[0146] 2.2 Anti-inflammatory test - inhibition of neutrophil aggregation

[0147] Test method: copper sulfate was used to induce damage to the neuromast cells in the lateral line region of zebrafish embryos to cause neutrophil aggregation model for testing. 24 fish embryos were exposed to 0.16 mg / L copper sulfate pentahydrate and 5 g / L peony gel oil solution of application example 1, and 3 fish embryos were exposed to 0.16 mg / L copper sulfate pentahydrate solution as a model control group, and 3 fish embryos were exposed to 0.16 mg / L copper sulfate pentahydrate and 0.0036 mg / L indomethacin solution as a positive control group. After exposure for 40 minutes, the fish embryos were fixed and stained with Sudan black, and the number of neutrophils in the lateral line region was counted and statistically analyzed.

[0148] The experimental results show that the soothing effect of the model control group is not significant (0% neutrophil aggregation inhibition rate), the soothing effect of the positive control group is better (30% neutrophil aggregation inhibition rate), and the peony gel oil of Application Example 1 of the present invention has a neutrophil aggregation inhibition rate of 92%, and the soothing effect is more significant.

[0149] The representative picture of the test results is as follows Figure 7-9 As shown, Figure 7 is a representative figure of the model control group. Figure 8The positive control group: a representative image of 0.0036 mg / L indomethacin. Fig. 9 It is a representative diagram of the peony gel oil of Application Example 1.

[0150] The results of the inflammation test show that the peony gel oil of the present invention can significantly inhibit the aggregation of neutrophils in zebrafish embryos, has an anti-inflammatory effect, and can inhibit the inflammatory response after injury by inhibiting the aggregation of neutrophils.

[0151] Finally, it should be noted that the above embodiments are only used to illustrate the technical solution of the present invention rather than to limit the scope of protection of the present invention. Although the present invention has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that the technical solution of the present invention can be modified or replaced by equivalents without departing from the essence and scope of the technical solution of the present invention.

Claims

1. A composition capable of carrying rare ginsenosides, characterized in that: The invention comprises the following components by weight: 3-8 parts of peony seed oil, 3-5 parts of solid wax and 1-5 parts of peony fermentation hydrogen donor; the solid wax comprises at least one of peony unsaponifiable matter and plant wax ester; the peony fermentation hydrogen donor comprises peony fermentation glycerol.

2. The composition capable of carrying rare ginsenosides according to claim 1, characterized in that: The peony fermentation hydrogen donor also includes a supramolecular peony solution, and the mass ratio of the peony fermentation glycerol to the peony fermentation hydrogen donor is (1-3): (1-3).

3. The composition capable of carrying rare ginsenosides according to claim 1, characterized in that: The peony fermented glycerol is prepared by fermenting the whole peony plant with glycerol-producing Candida.

4. The composition capable of carrying rare ginsenosides according to claim 1, characterized in that: The method for preparing peony fermented glycerol comprises the following steps: treating the whole peony plant with cellulase, sterilizing by high pressure to obtain peony fermentation medium, fermenting with glycerol-producing Candida as fermentation strain, taking supernatant, decolorizing and sterilizing to obtain the peony fermented glycerol.

5. The composition capable of carrying rare ginsenosides according to claim 4, characterized in that: The temperature of the cellulase treatment is 48° C.-52° C., and the time is 30 h-40 h; and / or the weight of the cellulase is 0.08%-0.12% of the weight of the whole peony plant.

6. The composition capable of carrying rare ginsenosides according to claim 4, characterized in that: The fermentation temperature is 25°C-35°C, and the fermentation time is 75h-85h.

7. A method for preparing a composition carrying rare ginsenosides according to any one of claims 1 to 6, characterized in that: The following steps are involved: The peony seed oil, solid wax and peony fermentation hydrogen donor are uniformly mixed, and an alkaline catalyst is added to carry out a catalytic reaction. After the reaction is completed, the catalyst is neutralized and filtered to obtain the composition capable of carrying rare ginsenosides.

8. Use of the composition carrying rare ginsenosides according to any one of claims 1 to 6 in cosmetics and / or preparation of health products and / or preparation of medicines.

9. A peony gel oil containing rare ginsenosides, characterized in that: The invention comprises the following components in parts by weight: 0.01 to 0.1 parts of rare ginsenosides, 0.1 to 1 parts of the composition capable of carrying rare ginsenosides according to any one of claims 1 to 6, and 7 to 9 parts of peony seed oil.

10. Use of the peony gel oil containing rare ginsenosides according to claim 9 in the preparation of health products and / or medicines.

Images