Application of saponin in preparation of activated water and device for producing activated water by using saponin

Abstract

The invention belongs to the technical field of cosmetics, and provides application of saponin in preparation of activated water and a device for producing the activated water by using the saponin. Activated water can be obtained by mixing saponin with water, and the saponin is selected from monomeric saponin, total saponin or saponin composition. The half breadth of the activated water is adjusted by adjusting the adding amount of the saponin and the standing time after the saponin and the water are mixed, so that submolecular water or micromolecular water is obtained. On the other hand, saponin and water can be mixed and then added into a device for producing activated water, and small molecule water is prepared through the synergistic effect of saponin and ore. The method for preparing the activated water by utilizing the saponin or the saponin cooperated with the ore is simple, convenient and rapid, is beneficial to cost reduction, and is convenient for mass production and industrial popularization. The prepared submolecular water or micromolecular water has moisturizing and anti-aging effects, is applied to the field of cosmetics, and can improve the product effects of the cosmetics.

Description

Technical Field

[0001] The invention belongs to the technical field of cosmetics, and in particular relates to the application of saponin in the preparation of activated water and a device for producing activated water by using saponin. Background Art

[0002] Ginseng (Panaxginseng C..Meyer) is one of the most important medicinal plants and a traditional beauty and skin care product in my country. Its whitening, anti-wrinkle and anti-aging effects are unanimously recognized. Ginseng plants include ginseng, American ginseng and Panax notoginseng. So far, the chemical components found in ginseng are mainly dammarane-type triterpenoid saponins, polyene ynes, sesquiterpenes, flavonoids, tannin acid, polyamines and polysaccharides. Among them, ginsenoside is the iconic component of ginseng and is also the main active ingredient of American ginseng, Panax notoginseng and other plants of the genus Panax in the Araliaceae family. Studies have found that ginsenosides have multiple effects such as anti-aging, antioxidant, whitening, anti-inflammatory, antibacterial, and hair growth.

[0003] Modern research has found that in nature, water does not exist as a single molecule, but exists in the form of molecular clusters formed by hydrogen bonds. Intermolecular hydrogen bonds are a kind of intermolecular force. In liquid water, they are in a dynamic balance of constant disconnection and combination. Due to the lack of direct means to measure the structure of water molecule clusters, the size of water molecule clusters is generally expressed by the half-width of the water vibration frequency measured by nuclear magnetic resonance. The larger the half-width, the larger the water molecule cluster, and the smaller the half-width, the smaller the water molecule cluster. As the water molecule clusters become smaller, the solubility, permeability, metabolism, diffusivity, and emulsification of water are enhanced. Generally, water with a half-width > 100Hz is classified as large molecule water, and more precisely, water with a half-width > 140Hz is large molecule water. Water with a half-width < 100Hz is classified as small molecule water, and water with a half-width < 80Hz is considered to be small molecule water in the strict sense. Studies have found that natural high-quality drinking water 17 The half-width of O-NMR is generally in the range of 70-90 Hz. Tap water is generally composed of 12-13 water molecules, and its half-width is around 121 Hz; pure water is composed of 30-40 water molecules, and its half-width is around 149 Hz.

[0004] At present, the main technologies and means to improve the activity of water bodies are: mechanical method, heat treatment method, strong field method (strong electric field, micro electric field, magnetic field, ultrasound, etc.), weak field method (biowave catalysis, far infrared energy catalysis, quantum technology, etc.). Due to the particularity of the pharmaceutical and cosmetic fields, the weak field method has become the first choice. Common ones include tourmaline, far infrared ceramics, magnets, and bamboo charcoal activated water technology. For example, the Chinese patent with application number 202410305969.3 discloses a low-hertz water incubator device, including: an incubation bucket, a mineralization plate, and an electrode excitation device. The wall of the incubation bucket is made of a mixture of purple sand mud, kaolin and carbon nanotubes, and the mineralization plate is sintered with kaolin, loess stone, purple sand mud, titanium dioxide and tourmaline. The Chinese patent with application number 202310976395.8 discloses a production process for mineralized small molecule water clusters. High-quality tourmaline, woodfish stone, and medical stone are selected as mineralized raw materials. Tourmaline, woodfish stone, and medical stone are configured in a volume ratio of 30:1:5:5. Pure water is mineralized to produce mineralized small molecule water clusters. Summary of the invention

[0005] In view of the above technical problems, the present invention provides the application of saponin in the preparation of activated water and a device for producing activated water using saponin. Activated water, i.e., submolecular water or small molecule water, is obtained by mixing saponin with water. The half width of the activated water can be adjusted by adjusting the amount of saponin added and the placement time after the saponin and water are mixed, thereby obtaining submolecular water or small molecule water. Alternatively, saponin is mixed with water and then added to a device for producing activated water, and saponin and ore act synergistically to prepare small molecule water. To achieve this purpose, the present invention adopts the following technical scheme: In a first aspect, the present invention provides the use of saponin in preparing activated water, wherein the saponin is mixed with water to obtain activated water.

[0006] Preferably, saponin is extracted from plants of the genus Panax, such as ginseng, American ginseng, Panax notoginseng, etc.

[0007] Preferably, the saponin is any one of a monomeric saponin, a saponin composition or a total saponin.

[0008] Preferably, the monomeric saponin is any one of ginsenoside Rg1, ginsenoside Rb1, ginsenoside Rd, ginsenoside Re, notoginsenoside R1, ginsenoside Rg3, ginsenoside Rh1, ginsenoside Rh2 or ginsenoside CK.

[0009] More preferably, the ratio of monomer saponin to water is 0.025g~0.1g:100ml, and the two are placed for 0.5h~3h after mixing. In the present invention, ginsenoside Rg1 is used as an example for explanation, and other monomer saponins can achieve the same technical effect.

[0010] Preferably, the saponin composition is any one of ginsenodiol saponin, ginsenotriol saponin, notoginsengdiol saponin, notoginsengtriol saponin, quinacridiolus saponin or quinacridoluol saponin.

[0011] More preferably, the ratio of the saponin composition to water is 0.025g~0.1g:100ml, and the two are placed for 0.5h~3h after mixing. In the present invention, the notoginsengtriol type saponin is used as an example for explanation, and other saponin compositions can achieve the same technical effect.

[0012] Preferably, the total saponin is any one of notoginseng total saponin, ginseng total saponin or American ginseng total saponin.

[0013] More preferably, the ratio of total saponins to water is 0.025g~0.1g:100ml, and the two are placed for 0.5h~3h after mixing. In the present invention, American ginseng total saponins are used as an example for illustration, and other total saponins can achieve the same technical effect.

[0014] In a second aspect, the present invention provides a device for producing activated water using saponin, the device comprising a column and ore, saponin and water filled in the column.

[0015] In the present invention, water is deionized water or purified water, and the activated water obtained is submolecular water or small molecule water.

[0016] 17 O-NMR half width of 100Hz is considered to be the boundary between large molecule water and small molecule water. The half width of commonly used tap water is around 121Hz, and the half width of well-known pure water is around 149Hz. The half width of small molecule water in the true sense is <80Hz. The function and effect of water with a half width between 120-80Hz has not been studied. 17 Water with O-NMR half-width in the range of 100±20 Hz was defined as submolecular water for the first time, and water with half-width <80 Hz was defined as small molecule water. The size of water molecules was re-refined, and the preparation process and function of submolecular water were studied, providing a better basis for the high-quality utilization of water.

[0017] Compared with the prior art, the present invention has at least the following beneficial effects: 1. Saponin is mixed with water or added to a device for producing activated water after mixing to obtain activated water. The half width of activated water can be adjusted by adjusting the amount of saponin added and the standing time after mixing, thereby obtaining submolecular water or small molecule water. This method is simple and fast, which is conducive to reducing costs, facilitating mass production and industrial promotion.

[0018] 2. The submolecular water or small molecule water prepared by this method has moisturizing and anti-aging effects, and can be applied in the cosmetics field to enhance the product efficacy of cosmetics.

[0019] 3. The present invention utilizes the characteristics of saponin to activate water molecules and produce submolecular water and small molecule water. Compared with the mechanical method, heat treatment method, strong field method, weak field method and other methods in the prior art, it is more energy-saving and environmentally friendly, has low equipment requirements, and is easy to operate. While activating water, it also has the effects of moisturizing and anti-aging, and is suitable for use in the field of health. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 This is the half-width detection diagram of pure water; Figure 2 This is a half width detection diagram of submolecular water in Example 1; Figure 3 This is a half width detection diagram of submolecular water in Example 2; Figure 4 This is a half width detection diagram of small molecule water in Example 3; Figure 5 This is a half width detection diagram of small molecule water in Example 4; Figure 6 This is a half width detection diagram of submolecular water in Example 5; Figure 7 This is a half width detection diagram of submolecular water in Example 6; Figure 8 This is a half width detection graph of small molecule water in Example 7; Fig. 9 This is a half width detection graph of small molecule water in Example 8; Fig.10 This is a half width detection diagram of submolecular water in Example 9; Fig.11 This is a half width detection diagram of submolecular water in Example 10; Fig.12 This is a half width detection graph of small molecule water in Example 11; Fig.13 This is a half width detection graph of small molecule water in Example 12; Fig.14 This is a half width detection diagram of submolecular water in Example 13; Fig.15 This is a half width detection diagram of submolecular water in Example 14; Fig.16 This is a half width detection graph of small molecule water in Example 15; Fig.17 This is a half width detection graph of small molecule water in Example 16; Fig.18 This is a half width detection graph of small molecule water in Example 17; Fig.19 This is a half width detection diagram of submolecular water in Example 18; Fig. 20 This is a half width detection diagram of submolecular water in Example 19; Fig.21 This is a half width detection graph of small molecule water in Example 20; Fig. 22 This is a half width detection graph of small molecule water in Example 21; Fig.23 This is a schematic diagram of the structure of the device for producing activated water in Example 22; Fig.24 This is a half width detection graph of submolecular water in Example 23; Fig.25 This is a half width detection graph of small molecule water in Example 24; Fig.26 This is a half width detection graph of small molecule water in Example 25; Fig. 27 Submolecular water moisturizing cell efficacy test; Fig.28 Relative expression levels of submolecular water MMP-1, MMP-3, and MMP-9 gene mRNA. DETAILED DESCRIPTION

[0021] The technical solution of the present invention is further described below in conjunction with the accompanying drawings and through specific implementation methods. However, the following examples are only simplified examples of the present invention and do not represent or limit the scope of protection of the present invention. The scope of protection of the present invention shall be subject to the claims.

[0022] Unless otherwise specified, the raw materials and materials used in the examples of the present invention are purchased through general commercial channels.

[0023] The source information of the relevant raw materials and materials involved in the following examples are as follows: Ore, purchased from Shiping Mine in Yunnan; HaCaT cells were purchased from the Cell Bank of the Chinese Academy of Sciences; HFF cells were purchased from the Cell Bank of the Chinese Academy of Sciences; Total RNA extraction kit was purchased from Tiangen, Beijing, China, catalog number DP430; Purified water, commonly available in the market; Ginsenoside Rg1, purchased from Yuanye, catalog number S33043; Total saponins of American ginseng, purchased from Yuanye, catalog number S23035; Notoginseng triol type saponins are prepared in the laboratory, and the preparation method is as follows: the Panax notoginseng medicinal material is crushed and passed through a 24-mesh sieve, and extracted with 85% ethanol water by heating and reflux. The extraction is performed twice, each time for 2 hours, the filtrates are combined, filtered, and concentrated under reduced pressure until the mass of the extract is no more than 100 g. 200 g of D101 macroporous resin is taken, and the column is loaded with pure water. The extract is then loaded and adsorbed overnight, and then 4 BV of water is used for elution, 5 BV of 30% ethanol water is used for elution, and 7 BV of 50% ethanol water is used for elution. The 50% ethanol water elution portion is collected, concentrated under reduced pressure, and freeze-dried to obtain the product.

[0024] Pure water uses NMR-500 to detect the half width of water, such as Figure 1 As shown, the half width of pure water is 146.59 Hz, which belongs to macromolecular water.

[0025] <Examples 1-4>

[0026] According to Table 1 below, add ginsenoside Rg1 to 100 mL of pure water and leave it for 1 hour. Use NMR-500 to detect the half-width of water molecule clusters. See the half-width detection diagram. Figure 2-5 .

[0027] Table 1 Main parameters and results of Examples 1-4 Example Purified water Placement time <![CDATA[Ginsenoside Rg1]]> Half bandwidth (Hz) category Detection diagram 1 100mL 1h 0.025g 117.46 Submolecular water 2 2 100mL 1h 0.05g 110.28 Submolecular water 3 3 100mL 1h 0.075g 71.79 Small molecule water 4 4 100mL 1h 0.1g 67.42 Small molecule water 5 It can be seen from Table 1 that when the storage time remains unchanged, the amount of ginsenoside Rg1 added is inversely proportional to the half-width, that is, the less the amount added, the larger the half-width; the more the amount added, the smaller the half-width (the water molecule clusters are smaller).

[0028] <Example 5-8> According to Table 2 below, add Panax notoginsengtriol saponin to 100 mL of pure water and leave it for 1 hour. Use NMR-500 to detect the half width of water molecule clusters. The half width detection diagram is shown in Figure 6-9 .

[0029] Table 3 Main parameters and results of Examples 5-8 Example Purified water Placement time Notoginsenotriol saponins Half bandwidth (Hz) category Detection diagram 5 100mL 1h 0.025g 118.50 Submolecular water 6 6 100mL 1h 0.05g 110.07 Submolecular water 7 7 100mL 1h 0.075g 79.59 Small molecule water 8 8 100mL 1h 0.1g 54.72 Small molecule water 9 It can be seen from Table 2 that when the storage time remains unchanged, the amount of notoginsengtriol saponin added is inversely proportional to the half-width, that is, the less the amount added, the larger the half-width; the more the amount added, the smaller the half-width (the water molecule clusters are smaller).

[0030] <Examples 9-12> According to Table 3 below, add American ginsenosides to 100 mL of pure water and leave for 1 hour. Use NMR-500 to detect the half width of water clusters. See the half width detection diagram. Figure 10-13 .

[0031] Table 3 Main parameters and results of Examples 9-12 Example Purified water Placement time Total saponins from American ginseng Half bandwidth (Hz) category Detection diagram 9 100mL 1h 0.025g 117.65 Submolecular water 10 10 100mL 1h 0.05g 110.59 Submolecular water 11 11 100mL 1h 0.075g 73.97 Small molecule water 12 12 100mL 1h 0.1g 66.38 Small molecule water 13 It can be seen from Table 3 that when the storage time remains unchanged, the amount of total saponins added in American ginseng is inversely proportional to the half-width, that is, the less the amount added, the larger the half-width; the more the amount added, the smaller the half-width (the water molecule clusters are smaller).

[0032] <Examples 13-15>

[0033] According to Table 4 below, 0.075 g of ginsenoside Rg1 was added to 100 mL of pure water. The half width detection of water molecule clusters was performed using NMR-500. The half width detection diagram is shown in Figure 14-16 For the convenience of comparison, the data of Example 3 are listed in Table 4.

[0034] Table 4 Main parameters and results of Examples 13-15 Example Purified water Ginsenoside Rg1 Placement time Half bandwidth (Hz) category Detection diagram 13 100mL 0.075g 0.5h 108.09 Submolecular water 14 3 100mL 0.075g 1h 71.79 Submolecular water 4 14 100mL 0.075g 2h 68.46 Small molecule water 15 15 100mL 0.075g 3h 63.77 Small molecule water 16 As shown in Table 4, when the amount of ginsenoside added remains unchanged, the placement time is inversely proportional to the half width, that is, the shorter the placement time, the larger the half width; the longer the placement time, the smaller the half width (smaller water molecule clusters). And the change range tends to be stable after 2 hours.

[0035] <Examples 16-18>

[0036] According to Table 5 below, 0.075 g of notoginsengtriol type saponin was added to 100 mL of pure water. The half width of water molecule clusters was detected by NMR-500. For the convenience of comparison, the data of Example 7 are included in Table 5.

[0037] Table 5 Main parameters and results of Examples 16-18 Example Purified water Notoginsenotriol saponins Placement time Half bandwidth (Hz) category Detection diagram 16 100mL 0.075g 0.5h 105.81 Submolecular water 17 7 100mL 0.075g 1h 79.58 Submolecular water 8 17 100mL 0.075g 2h 67.83 Small molecule water 18 18 100mL 0.075g 3h 62.81 Small molecule water 19 As shown in Table 5, when the amount of notoginsengtriol saponin added remains unchanged, the placement time is inversely proportional to the half width, that is, the shorter the placement time, the larger the half width; the longer the placement time, the smaller the half width (smaller water molecule clusters). And the change range tends to be stable after 2 hours.

[0038] <Examples 19-21>

[0039] According to Table 6 below, 0.075 g of total saponins of American ginseng was added to 100 mL of pure water. The half width of water clusters was detected by NMR-500. For the convenience of comparison, the data of Example 11 are included in Table 6.

[0040] Table 6 Main parameters and results of Examples 20-23 Example Purified water Total saponins from American ginseng Placement time Half bandwidth (Hz) category Detection diagram 19 100mL 0.075g 0.5h 106.43 Submolecular water 20 11 100mL 0.075g 1h 73.87 Submolecular water 12 20 100mL 0.075g 2h 63.25 Small molecule water 21 21 100mL 0.075g 3h 58.86 Small molecule water 22 As shown in Table 6, when the amount of total saponins of American ginseng is constant, the placement time is inversely proportional to the half width, that is, the shorter the placement time, the larger the half width; the longer the placement time, the smaller the half width (smaller water molecule clusters). And the change range tends to be stable after 2 hours.

[0041] <Example 22>

[0042] This embodiment introduces in detail the device for producing activated water using saponin and the method for using the same.

[0043] like Fig.23 As shown, the device for producing activated water includes a column 1 and an ore 2 filled in the column 1. The column 1 is made of glass or stainless steel, and is cylindrical in shape with a diameter-to-length ratio of 4:160-400. The upper end of the column 1 is provided with a water inlet, which is connected to a water inlet pipe 3; the lower end is provided with a water outlet, which is connected to an external stainless steel water storage tank through a water outlet pipe 4 ( Fig.23 (not shown). A water control valve 5 is installed on the water outlet pipe 4 to control the flow rate of water. As required, the flow rate of water can be controlled between 0.01 cm / s and 0.2 cm / s. As required, a valve can also be installed on the water inlet pipe 3 to control the water inlet flow rate. In addition, the column made of stainless steel material is selected in this embodiment.

[0044] The ore 2 is an ore containing manganese and strontium. The ore 2 can be selected from particles with a particle size between 0.5 cm and 5 cm, and the filling rate in the column 1 is 40% to 80%, preferably 80%.

[0045] The ore of this example was tested by an X-ray fluorescence spectrometer (ZSX Primus IV), and the element contents were shown in Table 7 below, where w (B) represents the mass fraction of the substance.

[0046] Table 7 Ore element content element unit content Manganese (Mn) <![CDATA[w(B) / (10 -2 )]]> 0.038 Strontium (Sr) <![CDATA[w(B) / (10 -6 )]]> 398 Selenium (Se) <![CDATA[w(B) / (10 -6 )]]> 0.06 Zinc (Zn) <![CDATA[w(B) / (10 -6 )]]> 32.1 Silver (Ag) <![CDATA[w(B) / (10 -2 )]]> <0.05 Chromium (Cr) <![CDATA[w(B) / (10 -6 )]]> 9.95 Copper (Cu) <![CDATA[w(B) / (10 -6 )]]> 5.91 Full Iron (TFe) <![CDATA[w(B) / (10 -2 )]]> 0.74 Phosphorus (P) <![CDATA[w(B) / (10 -2 )]]> 0.007 Cadmium (Cd) <![CDATA[w(B) / (10 -6 )]]> 0.05 Calcium oxide (CaO) <![CDATA[w(B) / (10 -2 )]]> 50.23 Magnesium oxide (MgO) <![CDATA[w(B) / (10 -2 )]]> 1.72 The remaining embodiments described below all use the ore of this embodiment.

[0047] The activated water device can be used to prepare activated water, and the method is as follows: water flows into the top of the column 1 from the water inlet pipe 3 at a certain flow rate, flows through the ore 2, and adjusts the opening of the control valve 5 according to the flow rate of the water, so that the water flows out at a uniform speed. The water flowing out from the bottom is activated water, i.e., small molecule water or submolecular water. The activated water flows into an external stainless steel water storage tank through the water outlet pipe 4 for storage or is connected to a bottled water filling device for filling and collection. When filling, it is filled into a plastic bottle with a specification of 500mL / bottle.

[0048] Water quality analysis is required during the collection process. The analysis frequency can be performed once after each batch of water is produced, or according to a regular testing plan of daily, weekly or monthly to ensure that the water quality is always in the best condition. The test content includes the determination of pH value, conductivity, total dissolved solids and heavy metal content of activated water. At the same time, samples are taken and the half width of water molecules obtained by NMR-500 is detected. The half width within the range of 100±20Hz is a qualified product. Then the qualified products are labeled and put into storage.

[0049] The water may be selected as healthy water such as qualified pure water or deionized water. In this embodiment, the water entering the water inlet pipe 3 is prepared pure water or deionized water, and needs to be tested before entering the water inlet pipe 3.

[0050] More preferably, saponin can be added to water in a ratio of water: saponin = 100 mL: 0.025 g to 0.1 g. The saponin can be a monomer saponin, such as ginsenoside Rg1; it can also be a total saponin, such as American ginseng total saponin; or a saponin composition, such as notoginsengtriol type saponin.

[0051] <Examples 23-25>

[0052] According to Table 8 below, small molecule water was prepared using saponin and the activated water device of Example 22. According to the ratio of pure water: saponin = 100 mL: 0.075 g, water and saponin were mixed and added to the activated water device to convert the water into small molecules. Figure 24-26 .

[0053] Table 8 Main parameters and results of Examples 23-25 Example Length ratio Fill rate Purified Water: Saponins Flow rate Half bandwidth (Hz) category Detection diagram 23 4:400 80% 100mL: 0.075g ginsenoside Rg1 0.01cm / s 50.04 Small molecule water 24 24 4:400 80% 100mL: 0.075g notoginsengtriol saponin 0.01cm / s 48.14 Small molecule water 25 25 4:400 80% 100mL: 0.075g American ginsenosides 0.01cm / s 47.36 Small molecule water 26 It can be seen from Table 8 that the ore in the activated water device is both a filler and a small molecule water activator. Ginsenosides work together with the ore to prepare small molecule water.

[0054] In addition, from the comparison between Examples 23, 24, and 25 and Examples 4, 8, and 12, it can be seen that saponins and minerals act synergistically. When the amount of saponins added is the same, the synergistic effect of ginsenosides and minerals on water activation is better than that of ginsenosides alone (smaller half-width).

[0055] <Test Example 1>

[0056] Submolecular water moisturizing cell efficacy test.

[0057] Blank group: cells without treatment; Positive control group: cells treated with 0.3 mM calcium chloride; Experimental group: cells were treated with submolecular water (Example 16) obtained by adding 0.075 g of notoginsengtriol-type saponin to purified water and leaving it for 0.5 h.

[0058] Experimental principle: Aquaporin, also known as water pore protein, is a protein located on the cell membrane (intrinsic membrane protein). It forms a "pore" on the cell membrane and can control the entry and exit of water in the cell, just like a "cell water pump". When water molecules pass through the aquaporin, they form a single column and enter the curved and narrow channel. The internal dipole force and polarity will help the water molecules rotate and pass through the narrow channel at an appropriate angle. Therefore, the protein conformation of the aquaporin is the reason why only water molecules can pass through.

[0059] This test method sets up a blank group, a positive control group and a test group. The test group is a group containing a certain treatment factor or drug, etc. The moisturizing effect of the drug in the test group can be predicted through the difference in AQP3 gene expression between different groups.

[0060] Experimental steps: 1. Select HaCaT cells in logarithmic growth phase with good morphology and inoculate them into 24-well plates at 1×10 5 The cells / well were incubated in a 37°C incubator for 24 h.

[0061] 2. Set up a blank group, a positive control group and a test group, set up 3 replicates at a concentration of 500 μg / mL, treat the cells with the drug, and continue to incubate in a 37°C, 5% CO2 incubator for 24 h.

[0062] 3. Extract RNA using a total RNA extraction kit.

[0063] 3.1 Collection of adherent monolayer cells: Cells can be lysed directly in the culture vessel (the diameter of the vessel should not exceed 10 cm), or treated with trypsin and then centrifuged to collect the cell pellet.

[0064] 3.2 Lysis treatment: cell number less than 5×10 6 350 μl of lysis buffer was added, 5×10 6 -1×10 7 Add 600 μl of lysis buffer when the amount is reached.

[0065] 3.3 Transfer all the solution to the filter column, centrifuge at 12000rpm for 2min, and collect the filtrate.

[0066] 3.4 Add 1 volume of 70% ethanol (usually 350μl or 600μl) to the filtrate, prepare 70% ethanol and mix with RNase-Free ddH2O, transfer the resulting solution to the adsorption column, centrifuge at 12000rpm for 1min, pour out the waste liquid in the collection tube, and put the adsorption column back into the collection tube.

[0067] 3.5 Add 350 μl of deproteinized solution to the adsorption column, centrifuge at 12,000 rpm for 1 min, pour out the waste liquid in the collection tube, and put the adsorption column back into the collection tube.

[0068] 3.6 Preparation of DNase I working solution: Take 10 μl of DNase I stock solution and put it into a new RNase-Free centrifuge tube, add 70 μl of buffer and mix gently.

[0069] 3.7 Add 80 μl of DNase I working solution to the center of the adsorption column and leave it at room temperature for 15 minutes.

[0070] 3.8 Add 350 μl of deproteinized solution to the adsorption column, centrifuge at 12,000 rpm for 1 min, discard the waste liquid in the collection tube, and put the adsorption column back into the collection tube.

[0071] 3.9 Add 599 μl of rinse solution to the adsorption column, let it stand at room temperature for 2 minutes, centrifuge at 12000 rpm for 1 minute, pour out the waste liquid in the collection tube, and put the adsorption column back into the collection tube.

[0072] 3.10 Repeat 3.9 3.11 Centrifuge at 12000rpm for 2min and discard the waste liquid. Leave the adsorption column at room temperature for several minutes to completely dry the residual rinse liquid in the adsorption material.

[0073] 3.12 Transfer the adsorption column to a new RNase-Free centrifuge tube, add 30-100 μl RNase-Free ddH2O, place at room temperature for 2 minutes, and centrifuge at 12000 rpm for 2 minutes to obtain RNA solution.

[0074] 4. RNA Quantification Use an ultra-micro spectrophotometer, select RNA content determination, select pure water as a blank control, and detect the collected RNA solution.

[0075] 5. Reverse transcription of cDNA 5.1 Preparation of reaction system: Add 5× PrimeScript RT Master Mix, 1 μg RNA, and make up to 20 ul system with pure water.

[0076] 5.2 Reverse transcription reaction at 37℃ for 15min.

[0077] 5.3 Reverse transcriptase inactivation reaction: 85℃, 5sec.

[0078] 5.4 The obtained cDNA was stored in a refrigerator at 4°C.

[0079] 6. Fluorescence quantitative PCR 6.1 Primer synthesis.

[0080] 6.2 Preparation of fluorescent quantitative PCR reaction system: 10 μl of 2×SYBR Green dye, 0.8 μl of upstream primer, 0.8 μl of downstream primer, 1 μl of cDNA, and 7.6 μl of pure water, to make a 20 μl system.

[0081] 6.3 qPCR parameter setting: Set the parameters according to the experimental requirements.

[0082] Experimental results: see Table 9 and Fig. 27 .

[0083] Table 9 Evaluation results of moisturizing effect of test samples

[0084] serial number Sample name Detection concentration AQP3 improvement rate Experimental Group Panax notoginseng 500 μg / mL 111.6%*** Positive control group Calcium chloride 0.3mM 88.43%*** From Table 9 and Fig. 27 It can be concluded that at the detected concentration, notoginsengtriol saponin submolecular water can significantly increase the expression of AQP3 and has a certain moisturizing effect.

[0085] <Test Example 2>

[0086] Testing of the anti-aging cellular efficacy of submolecular water.

[0087] Blank group: cells without treatment; Model group: hydrogen peroxide (H2O2)-induced cell senescence model; Experimental group: cells were treated with submolecular water (Example 16) obtained by adding 0.075 g of notoginsengtriol-type saponin to purified water and leaving it for 0.5 h.

[0088] Experimental principle: One of the main causes of skin aging is the change in the structure of the dermis. There are many reasons for the structural change. For example, external factors activate matrix metalloproteinases (MMPs) in the body, resulting in excessive degradation of collagen and elastin in the dermis that support the skin structure, causing wrinkles, decreased elasticity and other aging symptoms. Hydrogen peroxide, as a small molecule oxidant, can easily cause cell aging through oxidative stress through the biomembrane system. Therefore, a hydrogen peroxide-induced cell aging model can be constructed to detect the expression level of matrix metalloproteinase genes, thereby determining the anti-aging efficacy of the test substance.

[0089] Experimental steps: 1. Select HFF cells in logarithmic growth phase with good morphology and inoculate them into 6-well plates. 2.5×10 5 The cells / well were incubated in a 37°C incubator for 24 h.

[0090] 2. Set up a blank group, a model group and an experimental group, set up 3 replicates at a mass concentration of 0.025%, treat the cells with the drug for 4 h, add 50 μM H2O2 for induction treatment for 2 h, and then re-administer the drug and continue incubating in a 37 ℃, 5% CO2 incubator for 24 h.

[0091] 3. Extract RNA using a total RNA extraction kit.

[0092] 3.1 Collection of adherent monolayer cells: Cells can be lysed directly in the culture vessel (the diameter of the vessel should not exceed 10 cm), or treated with trypsin and then centrifuged to collect the cell pellet.

[0093] 3.2 Lysis treatment: cell number less than 5×10 6 350 μl of lysis buffer was added, 5×10 6 -1×10 7 Add 600 μl of lysis buffer when the amount is appropriate.

[0094] 3.3 Transfer all the solution to the filter column, centrifuge at 12000rpm for 2min, and collect the filtrate.

[0095] 3.4 Add 1 volume of 70% ethanol (usually 350μl or 600μl) to the filtrate, prepare 70% ethanol and mix with RNase-Free ddH2O, transfer the resulting solution to the adsorption column, centrifuge at 12000rpm for 1min, pour out the waste liquid in the collection tube, and put the adsorption column back into the collection tube.

[0096] 3.5 Add 350 μl of deproteinized solution to the adsorption column, centrifuge at 12,000 rpm for 1 min, pour out the waste liquid in the collection tube, and put the adsorption column back into the collection tube.

[0097] 3.6 Preparation of DNase I working solution: Take 10 μl of DNase I stock solution and put it into a new RNase-Free centrifuge tube, add 70 μl of buffer and mix gently.

[0098] 3.7 Add 80 μl of DNase I working solution to the center of the adsorption column and leave it at room temperature for 15 minutes.

[0099] 3.8 Add 350 μl of deproteinized solution to the adsorption column, centrifuge at 12,000 rpm for 1 min, discard the waste liquid in the collection tube, and put the adsorption column back into the collection tube.

[0100] 3.9 Add 599 μl of rinse solution to the adsorption column, let it stand at room temperature for 2 minutes, centrifuge at 12000 rpm for 1 minute, pour out the waste liquid in the collection tube, and put the adsorption column back into the collection tube.

[0101] 3.10 Repeat 3.9 3.11 Centrifuge at 12000rpm for 2min and discard the waste liquid. Leave the adsorption column at room temperature for several minutes to completely dry the residual rinse liquid in the adsorption material.

[0102] 3.12 Transfer the adsorption column to a new RNase-Free centrifuge tube, add 30-100 μl RNase-Free ddH2O, place at room temperature for 2 minutes, and centrifuge at 12000 rpm for 2 minutes to obtain RNA solution.

[0103] 4. RNA Quantification Use an ultra-micro spectrophotometer, select RNA content determination, select pure water as a blank control, and detect the collected RNA solution.

[0104] 5. Reverse transcription of cDNA 5.1 Preparation of reaction system: Add 5× PrimeScript RT Master Mix, 1 μg RNA, and make up to 20 ul system with pure water.

[0105] 5.2 Reverse transcription reaction at 37℃ for 15min.

[0106] 5.3 Reverse transcriptase inactivation reaction: 85℃, 5sec.

[0107] 5.4 The obtained cDNA was stored in a refrigerator at 4°C.

[0108] 6. Fluorescence quantitative PCR 6.1 Primer synthesis 6.2 Preparation of fluorescent quantitative PCR reaction system: 10 μl of 2×SYBR Green dye, 0.8 μl of upstream primer, 0.8 μl of downstream primer, 1 μl of cDNA, and 7.6 μl of pure water, to make a 20 μl system.

[0109] 6.3 qPCR parameter setting: Set the parameters according to the experimental requirements.

[0110] Experimental results: see Table 10 and Fig.28 . Fig.28 In the figure, a, b, and c are the relative RNA expression levels of MMP-1 (matrix metalloproteinase-1), MMP-3, and MMP-9, respectively; in the three figures, H2O2 is the model group and Panax notoginseng is the experimental group.

[0111] Table 10 Evaluation results of anti-aging efficacy of test samples

[0112] serial number Sample name Detection concentration MMP-1 MMP-3 MMP-9 Anti-aging efficacy evaluation results Experimental Group Panax notoginseng 0.025 % <![CDATA[-84.733% ** ]]> <![CDATA[-81.907% *** ]]> <![CDATA[-66.759% ** ]]> Significant effect Model Group <![CDATA[H2O2]]> 50μM 1 1 1 From Table 10 and Fig.28It can be concluded that at the detected concentration, notoginsengtriol saponin submolecular water can significantly reduce the expression of MMP-1, MMP-3, and MMP-9, and has a certain anti-aging effect.

[0113] The applicant declares that the above is only a specific implementation mode of the present invention, but the protection scope of the present invention is not limited thereto. Those skilled in the art should understand that any changes or substitutions that can be easily thought of by those skilled in the art within the technical scope disclosed by the present invention are within the protection scope and disclosure scope of the present invention.

Claims

1. The application of saponin in the preparation of activated water, characterized in that: Mix saponin with water to get activated water.

2. The use of saponin as claimed in claim 1 in preparing activated water, characterized in that, The saponin is extracted from a plant of the genus Panax.

3. The use of saponin as claimed in claim 1 in preparing activated water, characterized in that, The saponin is any one of a monomeric saponin, a saponin composition or a total saponin.

4. The use of saponin as claimed in claim 3 in preparing activated water, characterized in that, The monomer saponin is any one of ginsenoside Rg1, ginsenoside Rb1, ginsenoside Rd, ginsenoside Re, notoginsenoside R1, ginsenoside Rg3, ginsenoside Rh1, ginsenoside Rh2 or ginsenoside CK.

5. The use of saponin as claimed in claim 4 in preparing activated water, characterized in that, The ratio of the monomer saponin to water is 0.025 g to 0.1 g: 100 ml, and the two are mixed and left for 0.5 h to 3 h.

6. The use of saponin as claimed in claim 3 in preparing activated water, characterized in that, The saponin composition is any one of ginsenodiol saponin, ginsenotriol saponin, notoginsengdiol saponin, notoginsengtriol saponin, quinacridiolus saponin or quinacridoluol saponin.

7. The use of saponin as claimed in claim 6 in preparing activated water, characterized in that: The ratio of the saponin composition to water is 0.025 g to 0.1 g: 100 ml, and the two are mixed and left for 0.5 h to 3 h.

8. The use of saponin as claimed in claim 3 in preparing activated water, characterized in that, The total saponin is any one of notoginseng total saponin, ginseng total saponin or American ginseng total saponin.

9. The use of saponin as claimed in claim 8 in preparing activated water, characterized in that, The ratio of total saponin to water is 0.025 g to 0.1 g:100 ml, and the two are mixed and placed for 0.5 h to 3 h.

10. A device for producing activated water using saponin, characterized in that: The device for producing activated water by utilizing saponin comprises a column and ore, saponin and water filled in the column.

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