Preparation method of paris polyphylla extract as well as product and application of paris polyphylla extract

Abstract

The invention relates to a preparation method of a paris polyphylla extract as well as a product and application of the paris polyphylla extract. The preparation method comprises the following steps: crushing paris polyphylla, mixing the crushed paris polyphylla with an ethanol water solution, and carrying out reflux extraction to obtain an extracting solution; concentrating the extracting solution to obtain an extract, dissolving the extract in an ethanol aqueous solution, loading the extract to macroporous adsorption resin, and eluting with the ethanol aqueous solution to obtain an eluent; concentrating the eluent to obtain an extract, dissolving the extract in an ethanol aqueous solution to obtain a decolorized stock solution, mixing the decolorized stock solution with aluminum oxide for decolorization, and filtering to obtain a supernatant to obtain a decolorized solution; and concentrating and drying the decolored solution to obtain the product. According to the method, natural pigments in the plant raw materials are removed while the saponin extraction rate is guaranteed, and the appearance quality of the product is improved.

Description

Technical Field

[0001] The invention belongs to the technical field of material extraction, and relates to a preparation method of a long-column Paris polyphylla extract, a product thereof and an application thereof. Background Art

[0002] Plants have a long history of being used for beauty and skin care. Many ancient books record plants with beauty and skin care effects. In addition, consumers' dual demand for beauty and health has made the market prospects of cosmetics added with plant raw materials broad. Among them, CN117338681A discloses a Paris polyphylla extract, its preparation method and application. Paris polyphylla medicinal materials are extracted 2 to 3 times with 70% ethanol at normal pressure reflux with 15 to 20 times the amount of raw medicine. The single extraction time is 2 to 3 hours. The extract is concentrated under reduced pressure. After the concentrated solution is cooled, it is added to a D101 macroporous adsorption resin that has been pre-treated with 95% ethanol aqueous solution and then balanced with pure water. It is gradient eluted with pure water and 30% to 95% ethanol aqueous solution in turn to obtain an eluent. After the eluent is concentrated and dried, a Paris polyphylla extract is obtained. The Paris polyphylla extract can effectively inhibit melanin synthesis and intracellular tyrosinase activity, and has good antioxidant capacity.

[0003] However, the color of Paris polyphylla extract is dark and is not suitable for direct use as a cosmetic plant raw material. Therefore, it is very important to provide a method for removing pigments from Paris polyphylla extract while ensuring a high content of effective ingredients. Summary of the invention

[0004] In view of the deficiencies in the prior art, the object of the present invention is to provide a method for preparing a Paris polyphylla extract and its product and application.

[0005] In order to achieve the purpose of the invention, the present invention adopts the following technical solutions:

[0006] In a first aspect, the present invention provides a method for preparing a Paris polyphylla extract, the preparation method comprising:

[0007] (1) crushing Paris polyphylla and mixing with ethanol aqueous solution to perform reflux extraction to obtain an extract;

[0008] (2) concentrating the extract to obtain an extract, dissolving it in an ethanol aqueous solution, loading it onto a macroporous adsorption resin, and eluting it with an ethanol aqueous solution to obtain an eluate;

[0009] (3) concentrating the eluate to obtain an extract, dissolving the extract in an ethanol aqueous solution to obtain a decolorized stock solution, mixing the decolorized stock solution with alumina for decolorization, filtering and collecting the supernatant to obtain a decolorized solution;

[0010] (4) Concentrate the decolorizing solution and dry it to obtain the product.

[0011] Preferably, the mass percentage of ethanol in the ethanol aqueous solution in step (1) is 50-70%, for example 50%, 52%, 54%, 56%, 58%, 60%, 62%, 64%, 66%, 68%, 70%, etc. Other specific point values ​​within the above numerical range can be selected and will not be described one by one here.

[0012] Preferably, the mass ratio of the ethanol aqueous solution to the long column Paris polyphylla in step (1) is (8-14):1, wherein the specific point values ​​in (8-14) can be selected from 8, 8.2, 8.5, 8.8, 9, 9.2, 9.5, 9.8, 10, 10.2, 10.5, 10.8, 11, 11.2, 11.5, 11.8, 12, 12.2, 12.5, 12.8, 13, 13.2, 13.5, 13.8, 14, etc. Other specific point values ​​within the above numerical range can be selected, and they will not be repeated here one by one.

[0013] Preferably, the reflux extraction is performed 1-3 times, for example, 1 time, 2 times, or 3 times.

[0014] Preferably, the time for a single reflux extraction is 1-2.5 h, for example, 1 h, 1.2 h, 1.4 h, 1.6 h, 1.8 h, 2 h, 2.2 h, 2.4 h, 2.5 h, etc. Other specific point values ​​within the above numerical range can be selected and will not be described one by one here.

[0015] Preferably, the concentration temperature in step (2) is 35-65°C, for example, 35°C, 37°C, 40°C, 42°C, 45°C, 48°C, 50°C, 52°C, 55°C, 58°C, 60°C, 62°C, 65°C, etc. Other specific points within the above numerical range can be selected and will not be described here one by one.

[0016] Preferably, the macroporous adsorption resin includes any one of D-101 macroporous adsorption resin, D-4020 macroporous adsorption resin, HPD-100A macroporous adsorption resin, HPD-400A macroporous adsorption resin, and AB-8 macroporous adsorption resin, or a combination of at least two thereof, and is further preferably D-101 macroporous adsorption resin, HPD-100A macroporous adsorption resin, HPD-400A macroporous adsorption resin, or AB-8 macroporous adsorption resin.

[0017] Preferably, the mass percentage of ethanol in the ethanol aqueous solution used for dissolving in step (2) is 1-20%, for example 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, etc. Other specific points within the above numerical range can be selected and will not be described one by one here.

[0018] Preferably, the concentration of the extract in the decolorized stock solution is 0.1-1.5kg / L, for example, 0.1kg / L, 0.2kg / L, 0.3kg / L, 0.4kg / L, 0.5kg / L, 0.6kg / L, 0.7kg / L, 0.8kg / L, 0.9kg / L, 1kg / L, 1.1kg / L, 1.2kg / L, 1.3kg / L, 1.4kg / L, 1.5kg / L, etc. Other specific point values ​​within the above numerical range can be selected, which will not be described one by one here.

[0019] Preferably, the ratio of the concentrated solution to the macroporous adsorption resin is (1-3) mL:1 g.

[0020] Among them, the specific point values ​​in (1-3) can be selected as 1, 1.2, 1.4, 1.6, 1.8, 2, 2.2, 2.4, 2.6, 2.8, 3, etc. Other specific point values ​​within the above numerical range can also be selected, and they will not be listed here one by one.

[0021] Preferably, the mass percentage of ethanol in the ethanol aqueous solution used for elution in step (2) is 40-95%, for example 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, etc. Other specific points within the above numerical range can be selected and will not be described one by one here.

[0022] Preferably, the volume of the ethanol aqueous solution in step (2) is 3-10 times the volume of the macroporous adsorption resin, for example, 3 times, 4 times, 5 times, 6 times, 7 times, 8 times, 9 times, 10 times, etc. Other specific point values ​​within the above numerical range can be selected and will not be described one by one here.

[0023] Preferably, the concentration temperature is 35-65°C, for example, 35°C, 40°C, 45°C, 50°C, 55°C, 60°C, 65°C, etc. Other specific point values ​​within the above numerical range can be selected and will not be described here one by one.

[0024] Preferably, the mass percentage of ethanol in the ethanol aqueous solution in step (3) is 40-80%, for example 40%, 42%, 45%, 48%, 50%, 52%, 55%, 58%, 60%, 62%, 65%, 68%, 70%, 72%, 75%, 78%, 80%, etc. Other specific point values ​​within the above numerical range can be selected and will not be described one by one here.

[0025] Preferably, in step (3), the ratio of the extract to the ethanol aqueous solution is (0.01-0.03) g:1 mL.

[0026] The specific point values ​​in (0.01-0.03) can be selected as 0.01, 0.012, 0.014, 0.016, 0.018, 0.02, 0.022, 0.024, 0.026, 0.028, 0.03, etc. Other specific point values ​​within the above numerical range can be selected, and they will not be described one by one here.

[0027] Preferably, the mass ratio of the extract in step (3) to the mass of alumina is 1:(20-80), wherein the specific point values ​​in (20-80) can be selected from 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, etc., and other specific point values ​​within the above numerical range can be selected, which will not be repeated here.

[0028] Preferably, the decolorization temperature is 35-45°C and the time is 15-30 minutes.

[0029] The temperature can be selected as 35°C, 36°C, 37°C, 38°C, 39°C, 40°C, 41°C, 42°C, 43°C, 44°C, 45°C, etc., and the time can be selected as 15min, 16min, 17min, 18min, 19min, 20min, 21min, 22min, 23min, 24min, 25min, 26min, 27min, 28min, 29min, 30min, etc. Other specific point values ​​within the above numerical range can be selected, so they will not be described one by one here.

[0030] Preferably, the pH value of the decolorizing stock solution is 5-8, for example 5, 5.2, 5.4, 5.6, 5.8, 6, 6.2, 6.4, 6.6, 6.8, 7, 7.2, 7.4, 7.6, 7.8, 8, etc. Other specific points within the above numerical range can be selected and will not be described here one by one.

[0031] Preferably, the decolorizing solution is concentrated by vacuum concentration.

[0032] Preferably, the temperature for concentrating the decolorizing solution is 35-65°C, for example, 35°C, 40°C, 45°C, 50°C, 55°C, 60°C, 65°C, etc. Other specific points within the above numerical range can be selected and will not be described here one by one.

[0033] Preferably, the drying method includes any one of vacuum freeze drying, reduced pressure drying or water bath drying, or a combination of at least two thereof.

[0034] In a second aspect, the present invention provides a Paris polyphylla extract prepared according to the preparation method of the Paris polyphylla extract described in the first aspect.

[0035] In a third aspect, the present invention provides a use of the Paris polyphylla extract according to the second aspect in the preparation of cosmetics.

[0036] Preferably, the added amount of the Paris polyphylla extract in the cosmetic is 0.1-0.3%, for example 0.1%, 0.12%, 0.14%, 0.16%, 0.18%, 0.2%, 0.22%, 0.24%, 0.26%, 0.28%, 0.3%, etc. Other specific point values ​​within the above numerical range can be selected, which will not be described here one by one.

[0037] Compared with the prior art, the present invention has the following beneficial effects:

[0038] The invention adopts a reflux alcohol extraction method to extract the anti-photoaging efficacy components (total saponins) in the Paris polyphylla extract to the greatest extent, and can enrich the total saponin components by adopting AB-8 macroporous adsorption resin, thereby increasing the mass fraction of the total saponins in the Paris polyphylla extract, and can decolorize the enriched total saponin components by adopting neutral alumina, thereby removing the natural pigments in the plant raw materials, improving the appearance quality of the product and improving the stability of the product.

[0039] The main chemical components of the long-column Paris extract raw material obtained by the preparation method provided by the present invention are Paris saponin I, Paris saponin II, Paris saponin VI, Paris saponin VII, Paris saponin V, Paris saponin D, Paris saponin H, dioscin and slender dioscin, and the total amount is 25-45%, among which the total amount of Paris saponin I, Paris saponin II and Paris saponin VII in the Paris medicinal material content determination item included in the 2020 edition of the Chinese Pharmacopoeia is 10-25%, which is much higher than the total amount stipulated in the Pharmacopoeia and shall not be less than 0.6%. The long-column Paris extract raw material prepared by the present invention has good anti-photoaging effect, has a protective effect on UVB-induced HaCaT cells, and has good application prospects in anti-skin aging cosmetics; and the preparation method provided by the present invention, that is, the total saponin purity of the long-column Paris obtained by extraction, purification and decolorization is high, the total saponin content can reach 60-90%, the process is stable and controllable, simple and reasonable, and can be promoted for industrialization. BRIEF DESCRIPTION OF THE DRAWINGS

[0040] Figure 1 This is a flow chart for the preparation of long-column Paris polyphylla extract.

[0041] Figure 2 It is the fingerprint of the long-column Paris polyphylla extract obtained in Example 1, wherein chromatographic peaks 1-9 are Paris polyphylla saponin VII chromatographic peak, Paris polyphylla saponin D chromatographic peak, Paris polyphylla saponin H chromatographic peak, Paris polyphylla saponin VI chromatographic peak, Paris polyphylla saponin II chromatographic peak, Dioscorea zingiberensis chromatographic peak, Dioscorea zingiberensis chromatographic peak, Paris polyphylla saponin I chromatographic peak, and Paris polyphylla saponin V chromatographic peak, respectively.

[0042] Figure 3 It is the safe concentration of Paris polyphylla extract on HaCaT cells, compared with the control group, *P<0.05.

[0043] Figure 4 The effect of Paris polyphylla extract on the cell activity of HaCaT cells after UVB irradiation was compared with the control group. # P < 0.05; ## P < 0.01; ### P<0.001, compared with the model group, *P<0.05; **P<0.01; ***P<0.001. DETAILED DESCRIPTION

[0044] The technical solution of the present invention is further described below by specific implementation methods. It should be understood by those skilled in the art that the embodiments are only to help understand the present invention and should not be regarded as specific limitations of the present invention.

[0045] The sources of the functional ingredients contained in the products involved in the following embodiments and comparative examples are as follows (only the functional ingredients are reflected, and the necessary auxiliary ingredients contained in other commercially available raw materials are not repeated):

[0046] AB-8 macroporous adsorption resin is a product purchased from Beijing Solebow Technology Co., Ltd. under the trade name AB-8 macroporous adsorption resin;

[0047] HPD-100A macroporous adsorption resin is a product purchased from Beijing Solebow Technology Co., Ltd. under the trade name of HPD-100A macroporous adsorption resin;

[0048] HPD-400A macroporous adsorption resin is a product purchased from Beijing Solebow Technology Co., Ltd. under the trade name of HPD-400A macroporous adsorption resin.

[0049] Example 1

[0050] This embodiment provides a method for preparing a Paris polyphylla extract, and the preparation flow chart is as follows: Figure 1 As shown, the preparation method comprises:

[0051] (1) 100 g of Paris polyphylla was mixed with 1000 mL of 70% ethanol aqueous solution, and the mixture was refluxed for 1.5 h for 2 extractions, filtered, and the filtrates were combined. The obtained extract was rotary evaporated at 50° C. and −0.09 MPa, and concentrated to 200 mL under reduced pressure, and then dried in a water bath to obtain a dry extract;

[0052] (2) The obtained dry extract was dissolved in a 10% ethanol aqueous solution to a concentration of 0.19 g / mL, and loaded onto an AB-8 macroporous adsorption resin, with the ratio of the mass of the resin to the loading volume being 1 g:1.5 mL, and washed with water to remove impurities (water washing flow rate 4 BV / h, water elution volume 2 BV), and then eluted with a 60% ethanol aqueous solution (elution flow rate 4 BV / h, elution solvent volume 7 BV), and the eluate was collected;

[0053] (3) The obtained eluate was rotary evaporated at 50°C and -0.09MPa, and concentrated to 200mL under reduced pressure, and then dried in a water bath to obtain an extract, which was dissolved in a 60% ethanol aqueous solution to a concentration of 0.01g / mL, mixed with alumina for decolorization, the mass ratio of alumina to the extract being 40:1, the pH of the decolorization stock solution was adjusted to 6, the decolorization time was 15min, the decolorization temperature was 40°C, and filtered to collect the decolorized solution;

[0054] (4) The obtained decolorized solution was rotary evaporated at 50° C. and −0.09 MPa, concentrated to 100 mL under reduced pressure, and then freeze-dried to obtain a long-column Paris polyphylla extract raw material freeze-dried powder.

[0055] Its fingerprint is as follows Figure 2 shown.

[0056] Example 2

[0057] This embodiment provides a method for preparing a Paris polyphylla extract, the preparation method comprising:

[0058] (1) 100 g of Paris polyphylla was mixed with 1400 mL of 60% ethanol aqueous solution, and extracted under reflux for 2.5 h, once in total, filtered, and the filtrate was combined. The obtained extract was rotary evaporated at 35° C. and −0.09 MPa, and concentrated to 200 mL under reduced pressure, and then dried in a water bath to obtain a dry extract;

[0059] (2) The obtained dry extract was dissolved in a 5% ethanol aqueous solution to a concentration of 0.75 g / mL, and loaded onto a HPD-100A macroporous adsorption resin, with the ratio of the mass of the resin to the loading volume being 1 g:1 mL, and washed with water to remove impurities (water washing flow rate 4 BV / h, water elution volume 2 BV), and then eluted with a 40% ethanol aqueous solution (elution flow rate 4 BV / h, elution solvent volume 7 BV), and the eluate was collected;

[0060] (3) The obtained eluate was rotary evaporated at 35°C and -0.09MPa, and concentrated to 200mL under reduced pressure, and then dried in a water bath to obtain an extract, which was dissolved in a 60% ethanol aqueous solution to a concentration of 0.01g / mL, mixed with alumina for decolorization, the mass ratio of alumina to the extract being 40:1, the pH of the decolorization stock solution was adjusted to 6, the decolorization time was 15min, the decolorization temperature was 40°C, filtered, and the decolorized solution was collected;

[0061] (4) The obtained decolorized solution was rotary evaporated at 35° C. and −0.09 MPa, concentrated to 100 mL under reduced pressure, and then freeze-dried to obtain a raw freeze-dried powder of the long-column Paris polyphylla extract.

[0062] Example 3

[0063] This embodiment provides a method for preparing a Paris polyphylla extract, the preparation method comprising:

[0064] (1) 100 g of Paris polyphylla was mixed with 800 mL of 50% ethanol aqueous solution, and the mixture was refluxed for 1 h, and extracted 3 times in total. The filtrates were combined, and the obtained extract was rotary evaporated at 65° C. and −0.09 MPa, and concentrated to 200 mL under reduced pressure, and then dried in a water bath to obtain a dry extract;

[0065] (2) The obtained dry extract was dissolved in a 2% ethanol aqueous solution to a concentration of 1.5 g / mL, and loaded onto a HPD-400A macroporous adsorption resin, with the ratio of the mass of the resin to the loading volume being 1 g:3 mL, and washed with water to remove impurities (water washing flow rate 4 BV / h, water elution volume 2 BV), and then eluted with a 95% ethanol aqueous solution (elution flow rate 4 BV / h, elution solvent volume 7 BV), and the eluate was collected;

[0066] (3) The obtained eluate was rotary evaporated at 65°C and -0.09MPa, and concentrated to 200mL under reduced pressure, and then dried in a water bath to obtain an extract, which was dissolved in a 60% ethanol aqueous solution to a concentration of 0.01g / mL, mixed with alumina for decolorization, the mass ratio of alumina to the extract being 40:1, the pH of the decolorization stock solution was adjusted to 6, the decolorization time was 15min, the decolorization temperature was 40°C, filtered, and the decolorized solution was collected;

[0067] (4) The obtained decolorized solution was rotary evaporated at 65° C. and −0.09 MPa, concentrated to 100 mL under reduced pressure, and then freeze-dried to obtain a long-column Paris polyphylla extract raw material freeze-dried powder.

[0068] Example 4

[0069] This embodiment provides a method for preparing a Paris polyphylla extract, which differs from Embodiment 1 only in that the mass ratio of alumina to the extract in step (3) is 20:1, and other operations remain unchanged.

[0070] Example 5

[0071] This embodiment provides a method for preparing a Paris polyphylla extract, which differs from Embodiment 1 only in that the mass ratio of alumina to the extract in step (3) is 80:1, and other operations remain unchanged.

[0072] Example 6

[0073] This embodiment provides a method for preparing a Paris polyphylla extract, which differs from Embodiment 1 only in that the mass ratio of alumina to the extract in step (3) is 10:1, and other operations remain unchanged.

[0074] Example 7

[0075] This embodiment provides a method for preparing a Paris polyphylla extract, which differs from Embodiment 1 only in that the mass ratio of alumina to the extract in step (3) is 100:1, and other operations remain unchanged.

[0076] Example 8

[0077] This embodiment provides a method for preparing a Paris polyphylla extract, which differs from Embodiment 1 only in that the decolorization time in step (3) is 20 min, and other operations remain unchanged.

[0078] Example 9

[0079] This embodiment provides a method for preparing a Paris polyphylla extract, which differs from Embodiment 1 only in that the decolorization time in step (3) is 25 min, and other operations remain unchanged.

[0080] Example 10

[0081] This embodiment provides a method for preparing a Paris polyphylla extract, which differs from Embodiment 1 only in that the decolorization time in step (3) is 30 min, and other operations remain unchanged.

[0082] Embodiment 11

[0083] This embodiment provides a method for preparing a Paris polyphylla extract, which differs from Embodiment 1 only in that the decolorization temperature in step (3) is 35° C., and other operations remain unchanged.

[0084] Example 12

[0085] This embodiment provides a method for preparing a Paris polyphylla extract, which differs from Embodiment 1 only in that the decolorization temperature in step (3) is 45° C., and other operations remain unchanged.

[0086] Embodiment 13

[0087] This embodiment provides a method for preparing a Paris polyphylla extract, which differs from Embodiment 1 only in that the decolorization temperature in step (3) is 50° C., and other operations remain unchanged.

[0088] Embodiment 14

[0089] This embodiment provides a method for preparing a Paris polyphylla extract, which differs from Embodiment 1 only in that the decolorization temperature in step (3) is 20° C., and other operations remain unchanged.

[0090] Embodiment 15

[0091] This embodiment provides a method for preparing a Paris polyphylla extract, which differs from Embodiment 1 only in that the pH of the decolorizing stock solution in step (3) is 5, and other operations remain unchanged.

[0092] Example 16

[0093] This embodiment provides a method for preparing a Paris polyphylla extract, which differs from Embodiment 1 only in that the pH of the decolorizing stock solution in step (3) is 7, and other operations remain unchanged.

[0094] Embodiment 17

[0095] This embodiment provides a method for preparing a Paris polyphylla extract, which differs from Embodiment 1 only in that the pH of the decolorizing stock solution in step (3) is 8, and other operations remain unchanged.

[0096] Comparative Example 1

[0097] This comparative example provides a method for preparing a Paris polyphylla extract, which differs from Example 1 only in that an equal amount of alumina is replaced by activated carbon, and other operations remain unchanged.

[0098] Test Example 1

[0099] Determination of total saponin content in Paris polyphylla:

[0100] Test method: Refer to the literature "Yang Xiao, Zhang Zhenqiu. Determination of total saponins in Paris polyphylla [J]. Chinese Journal of Traditional Chinese Medicine, 2007, 25(11): 3. DOI: 10.3969 / j.issn.1673-7717.2007.11.089." The total saponins content was determined using the UV-perchloric acid colorimetric method.

[0101] Preparation of reference solution: Take Paris saponin I reference substance and add methanol to prepare a 0.21 mg / mL reference solution.

[0102] Preparation of test solution: Take long-column Paris polyphylla, crush it, pass it through a No. 2 sieve, take out 3.0 g, accurately weigh it, put it in a stoppered conical flask, accurately add 20 mL of methanol, weigh it, ultrasonically treat it for 10 minutes, cool it, weigh it again, make up the lost weight with methanol, shake it well, filter it, and obtain the solution.

[0103] Drawing of standard curve: Using UV-perchloric acid colorimetric method, transfer 0.40, 0.50, 0.60, 0.70, 0.80, 0.90 mL of reference solution respectively, put into 10 mL stoppered colorimetric tube, evaporate to dryness, add 5 mL of perchloric acid, bathe in 60℃ water for 15 min, take out and bathe in ice water for 15 min, adjust to zero with methanol as reference, measure the absorbance A at the maximum absorption wavelength of 407 nm with ultraviolet spectrophotometer, and draw the standard curve with Paris saponin I content and absorbance value as horizontal and vertical coordinates, respectively.

[0104] Determination of test solution: Pipette 0.1 mL of the test solution into a 10 mL stoppered colorimetric tube, determine the absorbance values ​​of the samples at a wavelength of 407 nm in sequence according to the method of drawing the standard curve, and calculate the total saponin content by the standard curve method.

[0105] The test results are shown in Table 1:

[0106] Table 1

[0107] Total saponin content (%) Longzhu Paris 4.24

[0108] Test Example 2

[0109] The decolorized stock solution and decolorized solution obtained in Examples 1-17 and Comparative Example 1 were filtered to obtain the supernatant, and the total saponin mass concentration was determined by the UV-perchloric acid colorimetric method of Test Example 1, and the absorbance at 204 nm was determined by a UV spectrophotometer after diluting it 100 times with 60% ethanol aqueous solution.

[0110] The decolorization rate and total saponin retention rate were calculated according to the following formula. The results are shown in Table 2.

[0111] Decolorization rate = (A1-A2) / A1×100%; Total saponin retention rate = C1 / C0×100%

[0112] Where: A1: absorbance of decolorized original solution, A2: absorbance of decolorized solution;

[0113] C0: total saponin concentration in the decolorized original solution (mg / mL); C1: total saponin concentration in the decolorized solution (mg / mL)

[0114] Table 2

[0115]

[0116]

[0117] As can be seen from the data in Table 2, the extract obtained by the extraction method described in the present application not only has a high total saponin retention rate (about 70%), but also has a high decolorization rate, reduces the pigment content in the product, and makes the product have a wider range of applications. The effect of replacing the decolorizing agent or decolorizing under other conditions prepared by the present application is not as good as that of the present application.

[0118] Test Example 3

[0119] Safety concentration test

[0120] Test method: CCK-8 method to determine cell viability

[0121] Weigh an appropriate amount of the long column Paris polyphylla extract prepared in Example 1 and dissolve it in DMSO to prepare a 10 mg / mL stock solution. HaCaT cells were cultured at 7×10 3 100 μL of the stock solution containing the extract of Paris polyphylla was added to 1 mL of DMEM basal medium, and then diluted into sample solutions of different concentrations, for a total of 6 concentrations. The old culture medium was discarded, and 100 μL of the sample solution was added to each well. The control group was added with 100 μL of DMEM basal medium. The blank group was a DMEM basal medium without cells, with 3 replicate wells in parallel, and cultured in a 37°C, 5% CO2 incubator for 24 hours. The drug-containing culture medium was discarded, and 100 μL of a mixture of DMEM basal medium and CCK-8 reagent was added to each well. The mixture was incubated in the incubator for 1 hour, and the OD value of each well at a wavelength of 450 nm was measured with an enzyme marker. Cell viability was calculated according to the following formula.

[0122] Cell viability (%) = [(A1-A2) / (A0-A2)] × 100%

[0123] Where: A0: absorbance value of control group, A1: absorbance value of sample group, A2: absorbance value of blank group

[0124] The results are as follows Figure 3 As shown by Figure 3 It can be seen that when the concentration of the extract obtained in the present application is below 5 μg / mL, its cell viability has no significant difference from that of the control group and is highly safe.

[0125] Test Example 4

[0126] Extract effect test: Effect of Paris polyphylla extract (Example 1) on cell activity after UVB irradiation of HaCaT cells

[0127] Test method: HaCaT cells were cultured at 7×10 3 100 μL per well was inoculated into a 96-well plate and cultured in a 37°C, 5% CO2 incubator. The next day, after the cells adhered to the wall, the supernatant was discarded and washed twice with PBS. After the liquid in the well was aspirated, 90 mJ / cm 2 The dry wells were irradiated with a UVB irradiation dose. After irradiation, the model group was immediately added with DMEM basal medium. The experimental group was given a dose within the safe concentration range (0.31-5μg / mL) based on the results of the CCK-8 method cell viability assay. Three parallel wells were performed. The blank group contained no cells but only DMEM basal medium. The control group was not irradiated with UVB and directly added with DMEM basal medium. The model group was not given any medicine. After 24 hours of culture, the cell activity was determined by the CCK-8 method. The calculation method is the same as that of Test Example 3.

[0128] The results are as follows Figure 4 shown.

[0129] The applicant declares that the present invention illustrates the preparation method of a long-column Paris polyphylla extract and its product and application through the above-mentioned embodiments, but the present invention is not limited to the above-mentioned embodiments, that is, it does not mean that the present invention must rely on the above-mentioned embodiments to be implemented. Those skilled in the art should understand that any improvement of the present invention, equivalent replacement of the raw materials of the product of the present invention, addition of auxiliary components, selection of specific methods, etc., all fall within the protection scope and disclosure scope of the present invention.

[0130] The preferred embodiments of the present invention are 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, a variety of 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.

[0131] It should also be noted that the various specific technical features described in the above specific embodiments can be combined in any suitable manner without contradiction. In order to avoid unnecessary repetition, the present invention will not further describe various possible combinations.

Claims

1. A method for preparing a long-column Paris polyphylla extract, characterized in that: The preparation method comprises: (1) crushing Paris polyphylla and mixing with ethanol aqueous solution to perform reflux extraction to obtain an extract; (2) concentrating the extract to obtain an extract, dissolving it in an ethanol aqueous solution, loading it onto a macroporous adsorption resin, and eluting it with an ethanol aqueous solution to obtain an eluate; (3) concentrating the eluate to obtain an extract, dissolving the extract in an ethanol aqueous solution to obtain a decolorized stock solution, mixing the decolorized stock solution with alumina for decolorization, filtering and collecting the supernatant to obtain a decolorized solution; (4) Concentrate the decolorizing solution and dry it to obtain the product.

2. The method for preparing the long-column Paris polyphylla extract according to claim 1, characterized in that: The mass percentage of ethanol in the ethanol aqueous solution in step (1) is 50-70%; Preferably, the mass ratio of the ethanol aqueous solution to the long-column Paris polyphylla in step (1) is (8-14):1; Preferably, the number of reflux extraction is 1-3 times; Preferably, the time of a single reflux extraction is 1-2.5 h.

3. The method for preparing the Paris polyphylla extract according to claim 1 or 2, characterized in that: The concentration temperature in step (2) is 35-65°C.

4. The method for preparing the Paris polyphylla extract according to any one of claims 1 to 3, characterized in that: The macroporous adsorption resin includes any one of D-101 macroporous adsorption resin, D-4020 macroporous adsorption resin, HPD-100A macroporous adsorption resin, HPD-400A macroporous adsorption resin, and AB-8 macroporous adsorption resin, or a combination of at least two thereof; Preferably, the macroporous adsorption resin includes D-101 macroporous adsorption resin, HPD-100A macroporous adsorption resin, HPD-400A macroporous adsorption resin or AB-8 macroporous adsorption resin; Preferably, the mass percentage of ethanol in the ethanol aqueous solution used for dissolving in step (2) is 1-20%; Preferably, the concentration of the extract in the decolorized stock solution is 0.1-1.5 kg / L; Preferably, the ratio of the concentrated solution to the macroporous adsorption resin is (1-3) mL:1 g.

5. The method for preparing the Paris polyphylla extract according to any one of claims 1 to 4, characterized in that: The mass percentage of ethanol in the ethanol aqueous solution used for elution in step (2) is 40-95%; Preferably, the volume of the ethanol aqueous solution in step (2) is 3-10 times the volume of the macroporous adsorption resin.

6. The method for preparing the Paris polyphylla extract according to any one of claims 1 to 5, characterized in that: The concentration temperature in step (3) is 35-65° C. Preferably, the mass percentage of ethanol in the ethanol aqueous solution in step (3) is 40-80%; Preferably, in step (3), the ratio of the extract to the ethanol aqueous solution is (0.01-0.03) g:1 mL.

7. The method for preparing the Paris polyphylla extract according to any one of claims 1 to 6, characterized in that: The mass ratio of the extract in step (3) to the mass ratio of alumina is 1:(20-80); Preferably, the decolorization temperature is 35-45°C and the time is 15-30min; Preferably, the pH of the decolorizing stock solution is 5-8.

8. The method for preparing the Paris polyphylla extract according to any one of claims 1 to 7, characterized in that: The method for concentrating the decolorizing solution is vacuum decompression concentration; Preferably, the temperature at which the decolorizing solution is concentrated is 35-65°C; Preferably, the drying method includes any one of vacuum freeze drying, reduced pressure drying or water bath drying, or a combination of at least two thereof.

9. A Paris polyphylla extract prepared by the method for preparing the Paris polyphylla extract according to any one of claims 1 to 8.

10. Use of the Paris polyphylla extract according to claim 9 in preparing cosmetics.

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