Extract of Rhus chinensis bark with whitening and soothing effects, preparation method and application thereof

Abstract

The invention discloses a rhus chinensis bark extract with whitening and soothing effects, a preparation method and an application thereof, comprising the following steps: crushing the rhus chinensis bark, heating and refluxing with ethanol for extraction, reducing the pressure on the obtained extract to recover the solvent, and concentrating to obtain an extract; mixing the obtained extract with MCI small-pore resin, loading it into an MCI column, and washing it with ethanol to obtain an active segment; the invention provides a rhus chinensis bark extract with whitening and soothing effects, a preparation method and an application thereof, optimizes the process parameters in the extraction process, optimizes the extraction part of the rhus chinensis, and obtains the active segment HDYFM-1 after separation and purification of the ethanol extract of the rhus chinensis bark by using MCI small-pore resin, which has whitening, antioxidant and soothing effects; the invention uses the rhus chinensis bark for extraction, which is better than the conventional extraction using the rhus chinensis leaf on the market, and has excellent effects in whitening, antioxidant and soothing effects.

Description

Technical Field

[0001] The present invention relates to the daily chemical field, in particular to a sumac bark extract with whitening and soothing effects, a preparation method thereof, and an application thereof. Background Art

[0002] With the development of society, people pay more attention to beauty, and beauty care and skin care have gradually become an important part of people's daily lives. External factors such as environmental pollution, dust, impurities in the air, and ultraviolet radiation in modern life will all have a negative impact on the skin. Long-term exposure to such an environment, if proper care is not taken, the skin is prone to various problems, such as enlarged pores, dull skin tone, dryness, fine lines, and skin spots.

[0003] Sumac extract has gradually attracted attention in the cosmetics field in recent years due to its antioxidant, anti-inflammatory, and moisturizing properties. It is rich in various active ingredients, such as flavonoids and polyphenols, which can effectively protect the skin from free radical damage and promote skin repair. In addition, its natural source also meets the preferences of modern consumers for natural skin care products.

[0004] Common extraction methods of sumac extract include solvent extraction, ultrasonic extraction, and water extraction, etc. Solvent extraction has high efficiency, but may leave residual solvents; ultrasonic extraction can improve the extraction rate, but the equipment cost is relatively high; water extraction has good safety, but the extraction efficiency is low; at the same time, there is a lack of research on the effective parts in the existing sumac extract, and there is a need for sumac extract with clear skin care effects on the market. Summary of the Invention

[0005] In order to overcome the above-mentioned drawbacks of the prior art, the purpose of the present invention is to provide a sumac bark extract with whitening and soothing effects, a preparation method thereof, and an application thereof.

[0006] The technical solution adopted by the present invention to solve its technical problems is: a sumac bark extract, which is obtained by chromatographic separation with an organic solvent using sumac bark as a raw material.

[0007] As a further improvement of the present invention: the organic solvent is methanol.

[0008] As a further improvement of the present invention: the organic solvent is ethanol.

[0009] The present invention also includes a preparation method of sumac bark extract, which includes the following steps:

[0010] Take sumac bark, crush it, and extract it by heating under reflux with ethanol. The solvent of the obtained extract is recovered under reduced pressure, and after concentration, an extract paste is obtained; the obtained extract paste is mixed with MCI macroporous resin, MCI is loaded into a column, and rinsed with ethanol; the active segment HDYFM-1 is obtained.

[0011] As a further improvement of the present invention: 90% ethanol is used in the ethanol heating reflux extraction after the sumac bark is crushed.

[0012] As a further improvement of the present invention: in the sample mixing of the obtained extract with MCI macroporous resin, the sample is mixed with 2 times the amount of MCI macroporous resin, and 5 times the amount of MCI is packed into a column.

[0013] As a further improvement of the present invention: in the ethanol rinsing, 10% ethanol, 20% ethanol, 30% ethanol, 50% ethanol, and 100% ethanol are used to rinse for 5 column volumes.

[0014] As a further improvement of the present invention: 100 kg of sumac bark is taken, crushed, and extracted by heating under reflux with 2 times the amount of 90% ethanol, repeated three times, and the extraction liquids are combined. The solvent of the obtained extraction liquid is recovered under reduced pressure, concentrated to obtain an extract. The obtained extract is mixed with 2 times the amount of MCI macroporous resin, packed into a column with 5 times the amount of MCI, and rinsed with 10% ethanol, 20% ethanol, 30% ethanol, 50% ethanol, and 100% ethanol for 5 column volumes respectively to obtain active segments 1 - 5, wherein active segment 4 is HDYFM - 1.

[0015] As a further improvement of the present invention: rinse with 10% ethanol, 20% ethanol, 30% ethanol, 50% ethanol, and 100% ethanol for 5 column volumes. Active segment 1 is obtained with 10% rinsing ethanol, active segment 2 is obtained with 20% rinsing ethanol, active segment 3 is obtained with 30% rinsing ethanol, active segment 4 is obtained with 50% rinsing ethanol, and active segment 5 is obtained with 100% rinsing ethanol.

[0016] The present invention also includes the application of sumac bark extract in cosmetics.

[0017] Compared with the prior art, the beneficial effects of the present invention are as follows: the present invention provides a sumac bark extract with whitening and soothing effects, its preparation method and application. The process parameters in the extraction process are optimized, and the extraction part of sumac is optimized. The ethanol extract of sumac bark is separated and purified by MCI macroporous resin to obtain the active segment HDYFM - 1, which has whitening, antioxidant and soothing effects; the present invention uses sumac bark for extraction, which is superior to the conventional extraction using sumac leaves on the market and has excellent effects in terms of whitening, antioxidant and soothing effects. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] In order to more clearly illustrate the technical solution, the drawings required for use in the embodiments will be briefly introduced below. Obviously, the drawings in the following description are only some embodiments of the present invention. For those of ordinary skill in the art, other drawings can be obtained based on these drawings without creative efforts.

[0019] Figure 1 For the tyrosinase IC in the embodiments of the present invention 50 The IC value and activity diagram corresponding to the HDYFM-Y concentration in the determination 50 In the figure, the X-axis is the HDYFM-Y concentration and the Y-axis is the activity.

[0020] Figure 2 For the tyrosinase IC in the embodiments of the present invention 50 The IC value and activity diagram corresponding to the HDYFM-P concentration in the determination 50 In the figure, the X-axis is the HDYFM-P concentration and the Y-axis is the activity.

[0021] Figure 3 For the ABST activity and IC in the embodiments of the present invention 50 The IC value and activity diagram corresponding to the HDYFM-Y concentration in the determination 50 In the figure, the X-axis is the HDYFM-Y concentration and the Y-axis is the activity.

[0022] Figure 4 For the ABST activity and IC in the embodiments of the present invention 50 The IC value and activity diagram corresponding to the HDYFM-P concentration in the determination 50 In the figure, the X-axis is the HDYFM-P concentration and the Y-axis is the activity.

[0023] Figure 5 For the DPPH activity and IC in the embodiments of the present invention 50 The IC value and activity diagram corresponding to the HDYFM-Y concentration in the determination 50 In the figure, the X-axis is the HDYFM-Y concentration and the Y-axis is the activity.

[0024] Figure 6 For the DPPH activity and IC in the embodiments of the present invention 50 The IC value and activity diagram corresponding to the HDYFM-P concentration in the determination 50 In the figure, the X-axis is the HDYFM-P concentration and the Y-axis is the activity.

[0025] Figure 7 Test result diagram of HDYFM-1 in the zebrafish soothing efficacy test in the embodiments of the present invention.

[0026] Figure 8 Column chart of the relative expression level of the il-6 gene of HDYFM-1 in the zebrafish soothing efficacy test in the embodiments of the present invention, where *p < 0.05 and ***p < 0.001.

[0027] Figure 9 Test result diagram of HDYFM-1 in the zebrafish antioxidant efficacy test in the embodiments of the present invention.

[0028] Figure 10This is a bar graph of the PEG2 content of HDYFM-1 in the cell experiment for testing the soothing effect of the embodiments of the present invention, where **p < 0.01 in the figure.

[0029] Figure 11 This is a bar graph of the relative expression level of the TNF-α gene of HDYFM-1 in the cell experiment for testing the soothing effect of the embodiments of the present invention, **p < 0.01.

[0030] Figure 12 This is a bar graph of the SOD activity of HDYFM-1 in the cell experiment for testing the antioxidant effect of the embodiments of the present invention.

[0031] Figure 13 This is a typical graph of the ROS fluorescence intensity of the ROS scavenging rate test of HDYFM-1 in the cell experiment for testing the antioxidant effect of the embodiments of the present invention. Detailed implementation manners

[0032] To make the objectives, technical solutions and advantages of the present invention clearer, the technical solutions of the present invention will be clearly and completely described below in conjunction with the specific embodiments of the present invention and the corresponding drawings. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without making creative efforts shall fall within the protection scope of the present invention.

[0033] It should be noted that the terms "first", "second", etc. in the specification and claims of the present invention and the above-mentioned drawings are used to distinguish similar objects, and do not have to be used to describe a specific order or sequence. It should be understood that such data can be interchanged under appropriate circumstances so that the embodiments of the present invention described herein can be implemented in an order other than those illustrated or described herein. In addition, the terms "comprising" and "having" and any variations thereof are intended to cover non-exclusive inclusion. For example, a process, method, system, product or device comprising a series of steps or units does not necessarily have to be limited to those steps or units clearly listed, but may include other steps or units not clearly listed or inherent to these processes, methods, products or devices.

[0034] Sumac (Rhus chinensis Mill.) is a deciduous small tree plant belonging to the genus Rhus of the Anacardiaceae family. It can reach up to 10 meters in height. Its small branches are brownish. The leaves are polymorphic, with a round base, dark green on the leaf surface, powdery green on the leaf back, and no petiole. The panicle is broad and much branched, and the female inflorescence is shorter, densely pubescent; the bracts are white, the petals are oblong, and curl outwards when blooming; the drupe is spherical and red when mature. It blooms from August to September and bears fruit in October.

[0035] Rhus chinensis is widely distributed in the arid and semi-arid regions of Asia, Europe, and Africa. Rhus chinensis can grow in harsh environments, such as saline-alkali soil, desert margins, and riverbank areas, showing its strong adaptability and vitality.

[0036] Rhus chinensis has a certain application history in traditional medicine and is used to treat various diseases, including skin diseases. Its extracts contain various bioactive substances, such as phenolic compounds, flavonoids, and alkaloids. These components are considered to have the potential for antioxidant, anti-inflammatory, and whitening effects. In particular, its antioxidant components can help resist free radical damage caused by ultraviolet rays and reduce pigmentation.

[0037] Common extraction methods of Rhus chinensis extracts include solvent extraction, ultrasonic extraction, and water extraction, etc. Solvent extraction has high efficiency, but may have residual solvents; ultrasonic extraction can improve the extraction rate, but the equipment cost is relatively high; water extraction has good safety, but the extraction efficiency is relatively low. At the same time, there is a research gap in the effective parts of the existing Rhus chinensis extracts. The present invention provides a preparation method and application of the bark extract of Rhus chinensis with whitening and soothing effects, optimizes the process parameters in the extraction process, optimizes the extraction parts of Rhus chinensis, and has whitening, antioxidant, and soothing effects.

[0038] Example 1:

[0039] Preparation of ethanol extract of Rhus chinensis leaves:

[0040] Take 100 g of Rhus chinensis leaves, crush them, and extract them with twice the amount of 90% ethanol by heating under reflux for three times. Combine the extracts, recover the solvent under reduced pressure for the obtained extract, and concentrate to obtain the ethanol extract HDYFM-Y of Rhus chinensis leaves.

[0041] Example 2:

[0042] Preparation of ethanol extract of Rhus chinensis bark:

[0043] Take 100 g of Rhus chinensis bark, crush them, and extract them with twice the amount of 90% ethanol by heating under reflux for three times. Combine the extracts, recover the solvent under reduced pressure for the obtained extract, and concentrate to obtain the ethanol extract HDYFM-P of Rhus chinensis bark.

[0044] Example 3:

[0045] Preparation of active segments HDYFM-Y-1~4 of Rhus chinensis leaves:

[0046] Mix the ethanol extract HDYFM-Y of Rhus chinensis leaves obtained in Example 1 with twice the amount of MCI macroporous resin, load the column with 5 times the amount of MCI, and rinse with 30% ethanol, 50% ethanol, 70% ethanol, and 100% ethanol for 5 column volumes respectively. Obtain the active segments HDYFM-Y-1~4.

[0047] Example 4:

[0048] Preparation of the active segments HDYFM-P-1 to 4 of Rhus chinensis Mill. bark:

[0049] The ethanol extract HDYFM-P of Rhus chinensis Mill. bark obtained in Example 2 was mixed with 2 times the amount of MCI small-pore resin, and 5 times the amount of MCI was used to pack the column. The column was rinsed with 30% ethanol, 50% ethanol, 70% ethanol, and 100% ethanol for 5 column volumes respectively. The active segments HDYFM-P-1 to 4 were obtained.

[0050] Example 5:

[0051] Preparation of the active segment HDYFM-1:

[0052] 100 kg of Rhus chinensis Mill. bark was crushed and extracted by heating under reflux with 2 times the amount of 90% ethanol for three times. The extraction solutions were combined, and the solvent was recovered under reduced pressure. The obtained extract was concentrated to obtain an extract. The obtained extract was mixed with 2 times the amount of MCI small-pore resin, and 5 times the amount of MCI was used to pack the column. The column was rinsed with 10% ethanol, 20% ethanol, 30% ethanol, 50% ethanol, and 100% ethanol for 5 column volumes respectively. The active segments 1 to 5 were obtained, and the active segment 4 was HDYFM-1.

[0053] Effect experiments were carried out on the above-prepared active segments:

[0054] Determination of tyrosinase inhibition rate:

[0055] The test drug was mixed with L-Dopa, and tyrosinase (final concentration 25 U / mL) was added to start the reaction. Three replicate wells were set, and at the same time, a blank control without the drug and a 377 positive control were set. At room temperature, for 5 min, the OD value was measured with an enzyme-labeled instrument, and the detection wavelength was 490 nm. The tyrosinase activity inhibition rate was calculated.

[0056] Tyrosinase activity inhibition rate (%) = (1 - OD of sample well 490 nm / OD of experimental control well 490 nm ) × 100%.

[0057] Table 1 Results table of tyrosinase activity inhibition rate corresponding to each active segment

[0058] Number Concentration Tyrosinase inhibition rate HDYFM-Y 100 mg / ml -15.21±0.81 HDYFM-Y-1 100 mg / ml 27.89±1.66 HDYFM-Y-2 100 mg / ml 17.9±0.68 HDYFM-Y-3 100 mg / ml 4.21±0.23 HDYFM-Y-4 100 mg / ml 11.15±0.25 HDYFM-P 100 mg / ml 43.06±0.99 HDYFM-P-1 100 mg / ml 31.74±0.73 HDYFM-P-2 100 mg / ml 38.02±1.57 HDYFM-P-3 100 mg / ml 5.19±0.98 HDYFM-P-4 100 mg / ml 9.17±0.12 HDYFM-1 100 mg / ml 53.76±0.13

[0059] Determination of tyrosinase IC 50 :

[0060] Add 20 ml of the test drug solution to a 96-well culture plate, divided into five concentration experimental groups (gradient: 1000 mg / ml, 250 mg / ml, 62.5 mg / ml, 15.625 mg / ml, 3.90625 mg / ml), set 3 replicate wells, and at the same time set a background group without tyrosinase (add the test drug at the same concentration). Add 105 ml of PBS buffer to the experimental groups and 130 ml of PBS buffer to the background group, mix well. Add 25 ml of tyrosinase solution (200 U / mL) to the experimental groups, mix well, then add 50 ml of the substrate L-DOPA, react at 25 ºC for 5 min, measure the absorbance (OD value) with an enzyme-linked immunosorbent assay (ELISA) reader, and the detection wavelength is 490 nm. Use GraphPad Prism 10 to calculate the IC 50 value.

[0061] Table 2 Statistical table of IC 50 measurement values of the active segments HDYFM-1 and HDYFM-P tyrosinase

[0062] Number <![CDATA[IC 50 value]]> HDYFM-1 79.16 mg / ml HDYFM-P 83.64 mg / ml

[0063] ABST activity and IC 50 measurement:

[0064] Add 20 ml of the test sample solution to a 96-well plate (divided into six concentrations (gradient: 1000 mg / ml, 200 mg / ml, 40 mg / ml, 8 mg / ml, 1.6 mg / ml, 0.32 mg / ml), with 3 replicates in each group), as well as two blank groups (ultrapure water with the same extract added with DMSO at the same concentration) and a positive control water-soluble vitamin E solution. Add ABTS to one blank group and the sample groups, and do not add it to the other. The blank group without ABTS is called the blank background reaction group, and the sample group without ABTS is called the sample background reaction group. The remaining experimental groups all add 180 ml of the diluted ABTS working solution, and place it in the dark at room temperature for 5 minutes. Measure the absorbance at 734 nm and calculate the ABTS radical scavenging activity. Use GraphPad Prism 10 to calculate the IC 50 value.

[0065] Table 3 Statistical table of IC 50 measurement values of the active segments HDYFM-1 and HDYFM-P ABST

[0066] Number <![CDATA[IC 50 value]]> HDYFM-1 122.2 mg / ml HDYFM-P 213.7 mg / ml

[0067] DPPH activity and IC 50 measurement:

[0068] Add 20 μL of the sample solution to be tested into a 96-well plate, divided into six concentrations (gradients are 1000 mg / ml, 200 mg / ml, 40 mg / ml, 8 mg / ml, 1.6 mg / ml, 0.32 mg / ml), set 3 replicate wells, and a blank control (anhydrous ethanol with the same concentration of DMSO as the extract). Continue to add 180 μL of DPPH solution to the 96-well plate, and then place it in an incubator at 37 °C for 1 h. Measure the absorbance at 517 nm and calculate the DPPH radical scavenging activity. Use GraphPad Prism 10 to calculate the IC 50 value.

[0069] Table 4 Statistical table of measured values of IC 50 of active segments HDYFM-1 and HDYFM-P

[0070] Number <![CDATA[IC 50 value]]> HDYFM-1 154.9 mg / ml HDYFM-P 221.4 mg / ml

[0071] According to the tyrosinase inhibition results, the tyrosinase inhibition rate of the crude extract of HDYFM-P (43.059 ± 0.989%) is better than that of the crude extract of HDYFM-Y (-15.211 ± 0.805). Therefore, the part of Rhus chinensis Mill. bark was selected for further development and efficacy testing. The tyrosinase inhibition rate of the active segment HDYFM-1 (53.76 ± 0.126%), tyrosinase inhibition rate IC 50 (IC 50 : 79.16 mg / ml), ABST scavenging rate IC 50 (IC 50 : 122.2 mg / ml), DPPH scavenging rate IC 50 (IC 50 : 154.9 mg / ml) are all better than the tyrosinase inhibition rate of the crude extract of HDYFM-P (43.06 ± 0.99%), tyrosinase inhibition rate IC 50 (IC 50 : 83.64 mg / ml), ABST scavenging rate IC 50 (IC 50 : 213.7 mg / ml), DPPH scavenging rate IC 50 (IC 50 : 221.4 mg / ml). Therefore, the active segment HDYFM-1 was selected for further efficacy testing.

[0072] Zebrafish soothing efficacy test:

[0073] 1) Randomly select zebrafish into a 6-well plate, 15 tails per well.

[0074] 2) Establish a zebrafish skin inflammation model by administering SLS in aqueous solution.

[0075] 3) Administer the sample in aqueous solution, and set up a normal control group and a model control group simultaneously, with a volume of 3 mL per well.

[0076] 4) Incubate in the dark at 28 °C for 18 h.

[0077] 5) Randomly select 10 zebrafish from each experimental group and place them under a fluorescence microscope for photographing. Analyze and collect data using advanced image processing software, analyze the number of neutrophils (N) in the zebrafish skin, and calculate and determine whether the sample has a soothing effect according to the formula.

[0078] Soothing effect (%) = (N (model control group) - N (sample group)) * 100% / (N (model control group) - N´ (normal control group)).

[0079] Table 5 Table of corresponding efficacy and test results of detection concentrations

[0080] Number Detection concentration % Efficacy % P value Detection result HDYFM-1 0.008 61 < 0.001 Significant

[0081] 1) Randomly select zebrafish and place them in a 6-well plate, with 30 zebrafish per well.

[0082] 2) Establish a zebrafish skin inflammation model by administering SLS in aqueous solution.

[0083] 3) Administer the sample in aqueous solution, and set up a normal control group and a model control group simultaneously, with a volume of 3 ml per well. Three biological replicates.

[0084] 4) Incubate in the dark at 28 °C for 18 h.

[0085] 5) Extract the total RNA of zebrafish from each experimental group, synthesize cDNA, and detect the gene expression of β-actin and the target gene using q-PCR.

[0086] 6) Use β-actin as an internal reference for gene expression and calculate the relative RNA expression level of the target gene.

[0087]

[0088] Table 6 Table of corresponding relative expression levels of il-6 gene and test results of detection concentrations

[0089] Number Detection concentration % Relative expression level of il-6 gene P value Detection result HDYFM-1 0.005 0.567 < 0.001 Significant

[0090] Zebrafish antioxidant efficacy test:

[0091] 1) Randomly select zebrafish and place them in a 6-well plate, with 15 zebrafish per well.

[0092] 2) Establish a zebrafish oxidative stress model by aqueous administration of menadione.

[0093] 3) Administer the sample aqueous solution, and set up a normal control group and a model control group simultaneously, with a volume of 3 mL per well.

[0094] 4) Incubate in the dark at 28 °C for 22 h.

[0095] 5) Stain the zebrafish with a specific ROS fluorescent reagent. After staining, randomly select 10 zebrafish from each experimental group and place them under a fluorescence microscope for photography. Analyze and collect data using advanced image processing software, analyze the fluorescence intensity (S) of the yolk sac of the zebrafish, calculate the antioxidant effect of the sample according to the formula, and determine whether it has antioxidant activity.

[0096]

[0097] Table 7 Table of corresponding efficacy and test results for detection concentrations

[0098] Number Detection concentration % Efficacy % P value Detection result HDYFM-1 0.008 24 < 0.001 Significant

[0099] Cell experiment for soothing efficacy test:

[0100] I. PGE2 content

[0101] 1) Inoculate RAW264.7 cells into a 6-well plate (3×105 cells / well) and incubate at 37 °C, 5% CO 2 for 24 h.

[0102] 2) After incubation, remove the culture medium, wash 1-2 times with D-Hanks. Replace the medium in the normal control group with fresh medium, replace the medium in the model control group with fresh medium containing the modeling agent (0.5 μg / mL LPS), and replace the medium in the sample group with fresh medium containing the sample and the modeling agent (0.5 μg / mL LPS). Continue to incubate at 37 °C, 5% CO 2 for 24 h.

[0103] 3) Collect the cell culture supernatant into a 1.5 mL sterile centrifuge tube and detect it according to the operation manual of the ELISA detection kit.

[0104]

[0105] II. TNF-α gene expression

[0106] 1) Inoculate HaCaT cells into a 6-well plate (6×105 cells / well) and incubate at 37 °C, 5% CO 2 for 24 h.

[0107] 2) After incubation, remove the culture medium, wash with D-Hanks 1-2 times, and then expose the sample group and the model control group to ultraviolet light (80 mJ / cm 2 ).

[0108] 3) After model establishment, replace the normal control group and the model control group with fresh culture medium, add fresh culture medium containing the sample to the sample group, and continue to incubate at 37 °C and 5% CO 2 for 24 h.

[0109] 4) Extract the total RNA of each experimental group, synthesize cDNA, and detect the gene expression of β -actin and the target gene.

[0110] 5) Use β -actin as the internal reference for gene expression, and calculate the relative RNA expression of the target gene.

[0111]

[0112] Table 8 Corresponding efficacy and test results of PGE2 content detection concentration

[0113] Number Detection index Detection concentration % Efficacy % P value Detection result HDYFM-1 PGE2 content 0.003125 58 < 0.01 Significant

[0114] Table 9 Detection results of TNF-α gene expression detection concentration

[0115] Number Detection index Detection concentration % Relative gene expression level P value Detection result HDYFM-1 TNF-α gene expression 0.0015625 0.324 < 0.01 Significant

[0116] Antioxidant efficacy test cell experiment:

[0117] 1. SOD activity test

[0118] 1) Inoculate cells in a 6-well plate (6×105 cells / well) and incubate at 37 °C and 5% CO 2 for 24 h.

[0119] 2) Model establishment: After incubation, remove the culture medium, gently wash the cells with D-Hanks once or twice, and then perform model establishment treatment. The model control group and the sample group are irradiated with UVB at 120 mJ / cm 2 , and at the same time place the normal control group in the dark.

[0120] 3) Sample administration: Replace the normal control group and the model control group with fresh culture medium, and replace the sample group with fresh culture medium containing the sample, and continue to incubate at 37 °C and 5% CO 2 for 24 h.

[0121] 4) After the incubation, remove the culture medium, wash the cells 3 times with D-Hanks, collect the cells, and perform the detection according to the instructions of the SOD assay kit. Calculate the antioxidant effect of the sample according to the following formula.

[0122]

[0123] Table 10 Table of corresponding efficacy and test results of detection concentrations

[0124] Number Detection concentration % Effect % P value Detection result HDYFM-1 0.0015625 33 < 0.01 Significant

[0125] 2. ROS scavenging rate test

[0126] 1) Seed the cells in a 12-well plate (1×105 cells / well) and incubate at 37 °C and 5% CO 2 for 24 h.

[0127] 2) Sample addition: Remove the culture medium and gently wash the cells once or twice with D-Hanks. Replace the normal control group and the model control group with fresh culture medium, and replace the sample group with fresh culture medium containing the sample, and continue to incubate at 37 °C and 5% CO 2 for 24 h.

[0128] 3) Model establishment: After the incubation, remove the culture medium, gently wash the cells once or twice with D-Hanks, and then perform the model establishment treatment. The model control group and the sample group are irradiated with UVA (9 J / cm 2 ), and at the same time place the normal control group in the dark.

[0129] 4) After the model establishment, stain the cells with the reactive oxygen species detection kit, take pictures with a fluorescence microscope, analyze the average fluorescence intensity (S) of each group of cells with Image J, and calculate the ROS scavenging rate of the sample group according to the formula.

[0130] Table 11 Table of corresponding efficacy and test results of detection concentrations

[0131] Number Detection concentration % ROS scavenging rate % P value Detection result HDYFM-1 0.0015625 73 < 0.001 Significant

[0132] Perform the cosmetic freckle removal and whitening efficacy test on the sumac bark extract emulsion prepared by applying the sumac bark extract of the present application. The test method is as follows:

[0133] 1.1 Test product: The original product of the cosmetic final product prepared with the sumac bark extract HDYFM-1 prepared by the present application as the only whitening efficacy raw material in the formula.

[0134] 1.2 Negative control: Blank control in the blackened area.

[0135] 1.3 Positive control: 7% ascorbic acid (vitamin C) product.

[0136] Subjects: A total of 20 people, including 3 males and 17 females, aged 23 to 57 years, with an average age of 45.70 ± 9.34 years, meeting the voluntary inclusion criteria for subjects.

[0137] Test method: The test was carried out in accordance with the specific requirements of the "Technical Specifications for Cosmetics Safety" (2015 edition). The non-exposed areas of the thighs or backs of the subjects were selected as the test sites. The minimum erythema dose (MED value) of the subjects' skin to ultraviolet irradiation was predicted 24 hours before the test. The xenon arc lamp of the solar simulator was used to irradiate once a day at the same irradiation point at a dose of 0.75 times the MED for 4 consecutive days. The 4 days after the irradiation were the skin darkening period, without any treatment. On the 5th day after the irradiation, the skin color of each test area was visually evaluated and detected by a skin color instrument, and the test areas with poor consistency (the areas where the ITA° value differed from the average value of all test areas by more than 5) were excluded. On the same day, the corresponding test substances were applied to each darkening test area according to the random table, and the test substances were continuously applied for 4 weeks. The skin color was visually evaluated and instrumentally detected at 1 week, 2 weeks, 3 weeks, and 4 weeks after the application, and the results were recorded.

[0138] Statistical method: Statistical analysis of the data was performed using statistical analysis software. Measurement data were expressed as: mean ± standard deviation, and a normal distribution test was conducted. If it met the requirements of normal distribution, paired t-tests were used for comparison before and after; otherwise, the Wilcoxon signed-rank test for two related samples was used; for the comparison of ranked data before and after, the Wilcoxon signed-rank test for two related samples was used; for the comparison between the test product and the control group, independent sample t-tests or the Wilcoxon rank-sum test were used.

[0139] At the same time, the regression coefficients of each parameter changing with time were calculated, and the significance level was P < 0.05.

[0140] The test results of the ITA° value and MI value of the test product and the control are as follows:

[0141] Test product:

[0142]

[0143] Negative control:

[0144]

[0145] Positive control:

[0146]

[0147] The test results of the self-evaluation of the visual skin color grade of the volunteers for the test product and the control:

[0148] Test product:

[0149]

[0150] Negative control:

[0151]

[0152] Positive control:

[0153]

[0154] Analysis of the time point difference before and after applying the test product and the negative control:

[0155]

[0156] Note: "NS" indicates no significant difference with P≥0.05; "S" indicates significant difference with P<0.05.

[0157] Regression coefficient analysis:

[0158]

[0159] Note: "NS" indicates no significant difference with P≥0.05; "S" indicates significant difference with P<0.05.

[0160] Test conclusion:

[0161] The test results of the freckle-removing and whitening effects of the cosmetics show that after applying the test product for 1 week, 2 weeks, 3 weeks, and 4 weeks, the difference in the ITA° value of the skin before and after application is significantly different from that of the negative control (P<0.05); after applying the test product for 1 week, 2 weeks, 3 weeks, and 4 weeks, the difference in MI before and after application is significantly different from that of the negative control (P<0.05); after applying the test product for 1 week, 2 weeks, 3 weeks, and 4 weeks, the difference in the visual skin color grade before and after application is significantly different from that of the negative control (P<0.05); compared with the negative control, the regression coefficients of ITA°, MI, and the visual skin color grade of the test product are all significantly different (P<0.05), and the test product has freckle-removing and whitening effects.

[0162] In summary, after reading the present invention document, those of ordinary skill in the art can make various other corresponding transformation schemes without creative mental labor according to the technical solutions and technical concepts of the present invention, and all of them fall within the scope protected by the present invention.

Claims

1. An extract of Rhus chinensis bark having whitening and soothing effects, characterized in that: The Rhus chinensis bark extract is obtained by using Rhus chinensis bark as a raw material and separating by organic solvent chromatography; The organic solvent is ethanol; The following steps are involved: The bark of Rhus chinensis is crushed and then extracted with ethanol under heating and reflux, and the obtained extract is decompressed to recover the solvent and concentrated to obtain an extract; the obtained extract is mixed with MCI small pore resin, and loaded with MCI column, Wash 5 column volumes with 10% ethanol, 20% ethanol, 30% ethanol, 50% ethanol and 100% ethanol; obtain active segments 1 to 5, among which active segment 4 is HDYFM-1.

2. The method for preparing the Rhus chinensis bark extract according to claim 1, wherein After the bark of Rhus chinensis is crushed, the bark is subjected to ethanol heating reflux extraction, and the ethanol used is 60% to 90%.

3. The method for preparing the Rhus chinensis bark extract according to claim 1, wherein The obtained extract is mixed with MCI small pore resin in an amount of 1 to 3 times the amount of MCI small pore resin, and loaded into the column in an amount of 2 to 8 times the amount of MCI.

4. The method for preparing the Rhus chinensis bark extract according to claim 3, wherein: The obtained extract was mixed with MCI small pore resin with 2 times the amount of MCI small pore resin and loaded into the column with 5 times the amount of MCI.

5. The method for preparing the Rhus chinensis bark extract according to claim 1, wherein The following steps are involved: Take 100 kg of Rhus chinensis bark, grind it and extract it with 1 to 3 times the amount of 60% to 90% ethanol by heating and reflux, repeat three times, combine the extracts, reduce the pressure to recover the solvent of the obtained extracts, and concentrate to obtain an extract. The obtained extract is mixed with 2 times the amount of MCI small-pore resin, loaded with 5 times the amount of MCI column, and rinsed with ethanol.

6. Use of the Rhus chinensis bark extract according to claim 1 in cosmetics.

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