Method for extracting high-purity salmon sperm-nodule-derived pdrn and application thereof in improving skin condition

Abstract

This invention discloses a method for extracting high-purity PDRN from salmon testes and its application in improving skin condition, belonging to the field of bioactive ingredient extraction technology. Frozen salmon testis tissue is cleaned of blood vessels and fatty impurities, then cryogenically pulverized at -20~-25℃. The pulverized tissue is added to a pre-treatment dispersion buffer and homogenized under high pressure to obtain a DNA fragmentation solution. The precipitate is then separated by centrifugation. The precipitate is dispersed in a dedicated enzyme digestion buffer, and enzymatic digestion with protease and RNase A is performed to inactivate the enzyme. DNase I is added to the digestion solution for enzymatic cleavage. After the reaction, a stop solution is added to terminate the reaction, and the supernatant is separated by centrifugation. Ethanol is added to the supernatant for precipitation, and centrifugation yields PDRN from salmon testes. This invention, through specific buffer systems and enzymatic digestion conditions, increases the PDRN content by 40~60 kDa, allowing for better absorption and utilization by the skin, improving facial hydration and elasticity, effectively reducing facial wrinkles, and improving facial skin condition. It can be used to prepare functional cosmetics.

Description

Technical Field

[0001] This invention belongs to the field of bioactive ingredient extraction and application technology, specifically relating to a method for extracting high-purity PDRN from salmon testes and its application in improving skin condition. Background Technology

[0002] Polydeoxyribonucleotides (PDRN) are naturally derived, low-molecular-weight DNA derivatives with a molecular weight ranging from 50 to 1500 kDa. PDRN is a linear polymer of deoxyribonucleotides with phosphodiester bonds, containing 50% double-stranded deoxyribonucleotides linked by hydrogen bonds between base pairs. PDRN is primarily extracted or purified from salmon or rainbow trout sperm cells (testicular tissue). Its base composition is up to 98% similar to human DNA, demonstrating significant effects in tissue repair, wound healing, anti-ischemia, and anti-inflammation, with extremely low side effects.

[0003] Different molecular weight PDRNs have different applications. PDRNs above 700 kDa are used for skin regeneration, articular cartilage regeneration, and promoting bone formation; those between 350 and 500 kDa are used in pharmaceuticals for tissue regeneration, corneal regeneration, and treatment of musculoskeletal diseases; and those between 40 and 60 kDa are used in cosmetics and food for barrier repair, wrinkle improvement, and skin rejuvenation. Therefore, it is necessary to prepare PDRNs of different molecular weights according to their intended use. However, PDRNs prepared using existing methods often have a wide molecular weight distribution, which cannot meet the requirements of cosmetics for a narrow molecular weight distribution. Summary of the Invention To address the problem of a wide molecular weight distribution of PDRN in existing technologies, this invention provides a method for extracting high-purity PDRN from salmon testes and its application in improving skin condition. By using specific buffer systems and enzymatic hydrolysis conditions, the content of low molecular weight PDRN (40-60 kDa) is increased, allowing for better absorption and utilization by the skin. This improves facial hydration and elasticity, effectively reduces facial (eye) wrinkles, and has a good effect on improving facial skin. It can be used to prepare functional cosmetics.

[0004] This invention is achieved through the following technical solution: In a first aspect, this invention provides a method for extracting high-purity PDRN from salmon testes, comprising the following steps: (1) Remove blood vessels and fat impurities from frozen salmon testis tissue, perform cryogenic pulverization at -20 to -25°C, add to pretreatment dispersion buffer, and perform high-pressure homogenization at 80 to 120 MPa to obtain DNA lysate, and centrifuge to obtain precipitate; under 80 to 120 MPa conditions, untangle the ultra-long DNA strands without destroying the medium nucleic acids of 40 to 120 kDa, as a pre-disruption aid; (2) Disperse the precipitate from step (1) in a special buffer for enzymatic hydrolysis, add protease and RNase A for enzymatic hydrolysis to obtain the hydrolysate, and then perform enzyme inactivation treatment; (3) After the enzyme inactivation in step (2), DNase I was added to the enzyme hydrolysate for enzyme digestion. After the enzyme digestion reaction was completed, the stop solution was added to terminate the enzyme digestion reaction. The supernatant was obtained by centrifugation and protein and RNA impurities were removed. (4) Add 2 to 3 times the volume of ethanol to the supernatant of step (3) for precipitation, centrifuge to collect the precipitate, and obtain PDRN from salmon testes.

[0005] Further, the pretreatment dispersion buffer described in step (1) includes the following concentrations of substances: 0.05~0.15 mol / L Tris, 0.1~0.2 mol / L NaCl, 0.001~0.1 mol / L EDTA, and 15~20 mmol / L sodium citrate, with the pH adjusted to 7.5~8.0 using hydrochloric acid; the pretreatment dispersion buffer contains low concentrations of sodium citrate and EDTA, which can chelate endogenous DNase in salmon testicular tissue and prevent spontaneous DNA degradation during homogenization.

[0006] Further, the enzyme-specific buffer solution described in step (2) comprises the following concentrations of substances: 0.04~0.06 mol / L Tris, 0.01~0.05 mol / L NaCl, 3~4 mmol / L MgCl2, 0.4~0.7 mmol / L CaCl2, 0.05~0.1 mol / L EDTA, and 20~30 mmol / L sodium citrate, with the pH adjusted to 7.3~7.5 using hydrochloric acid.

[0007] Furthermore, the protease in step (2) is one or more of trypsin, papain and proteinase K, the concentration of the protease after addition is 2~5 g / L, the concentration of RNase A after addition is 0.05~0.1 g / L, the enzymatic hydrolysis temperature is 35~45℃, and the enzymatic hydrolysis time is 3~5 h.

[0008] Furthermore, in step (3), the concentration of DNase I after addition is 0.05~0.1g / L, and the enzyme digestion reaction conditions are to react at 50~55℃ for 3~4h, and then cool down to 40~45℃ for 0.5~2h; DNase I treatment is a key step to achieve narrow distribution of PDRN.

[0009] Further, in step (1), the mass ratio of frozen salmon testis tissue to pretreatment dispersion buffer is 1:6~10, and it is subjected to high-pressure homogenization at 0~4℃ 1~2 times, each time for 5~15min.

[0010] Further, in step (2), the mass ratio of the precipitate and the enzyme digestion buffer is 1:5~8; in step (1), the centrifugation conditions are 0~4℃, 6000~8000r / min, 10~15min; in step (3), the centrifugation conditions are 0~4℃, 10000~12000r / min, 10~15min; in step (4), the centrifugation conditions are 8000~10000r / min, 8~10min.

[0011] Furthermore, in step (2), the enzyme inactivation treatment conditions are 60~70℃ for 8~10min; in step (3), EDTA-2Na is added to terminate the enzyme digestion reaction, and the concentration of EDTA-2Na after addition is 15~20mmol / L.

[0012] In a second aspect, the present invention provides high-purity salmon testis-derived PDRN prepared by the aforementioned preparation method.

[0013] Furthermore, the high-purity salmon testis-derived PDRN with a molecular weight distribution of 40–60 kDa accounts for ≥85%, and the OD... 260 / OD 280 The OD value is 1.85~1.9. 260 and OD 280 The absorbance of high-purity salmon testes-derived PDRN at wavelengths of 260 and 280 nm are respectively.

[0014] In a third aspect, the present invention provides the application of the aforementioned high-purity salmon testis-derived PDRN in improving skin condition.

[0015] Furthermore, the high-purity salmon testis-derived PDRN is used to prepare pharmaceuticals or cosmetics, particularly cosmetics with effects such as moisturizing, increasing facial elasticity, firming the skin, and anti-wrinkle properties.

[0016] Compared with the prior art, the beneficial effects achieved by the present invention are as follows: (1) The pretreatment dispersion buffer of the present invention contains EDTA and sodium citrate, which can chelate endogenous nucleases in salmon testes, avoid enzymatic strand breaks during homogenization to produce small molecule impurities, and sodium citrate and nucleic acid form a weak hydration protective layer, reducing the probability of medium fragments of 200kDa being sheared by fluid, limiting the high pressure homogenization pressure to a mild range of 80~120MPa. Combined with the chelation protection buffer containing sodium citrate and EDTA, it selectively breaks giant DNA wrapped in strands of >200 kDa, and hardly produces excessive ultra-short nucleic acid impurities.

[0017] (2) DNase I is a DNA-specific endonuclease that is activated by magnesium and calcium ions. It is used to controllably cleave long DNA chains to prepare narrowly distributed PDRNs of 40-60 kDa. The amount used and the reaction conditions directly determine the molecular weight distribution of the product. This invention uses a temperature-controlled two-stage digestion method. High-temperature (50-55℃) digestion breaks the long DNA backbone, generating most of the 80-120 kDa intermediates. Low-temperature (40-45℃) gentle cleavage of the intermediates reduces them to the 40-60 kDa range. A special digestion buffer is used, with a low concentration of divalent metals to moderately activate the digestion. Sodium citrate provides dynamic buffering ion strength and inhibits exonucleases, thus inhibiting fragment degradation to below 40 kDa.

[0018] (3) This invention prepares low molecular weight PDRN of 40~60 kDa by combining specific buffer and enzymatic hydrolysis conditions. It can penetrate the stratum corneum gap and reach the microvessels and fibroblasts in the dermis, and be better absorbed and utilized by the skin. It can improve the moisture content and elasticity of facial skin and effectively reduce facial (eye) wrinkles. It has a good effect on improving facial skin and can be used to prepare functional cosmetics. Detailed Implementation

[0019] The present invention is further illustrated below with reference to specific embodiments. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Experimental methods not specifically described in the following examples are generally performed under conventional conditions or as recommended by the manufacturer.

[0020] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of skill in the art. All reagents and materials used in this invention are readily available through conventional means, and unless otherwise specified, they shall be used in accordance with conventional methods in the art or as per the product instructions.

[0021] The proteases, RNase A, and DNase I used in the following examples and comparative examples are all commercially available products.

[0022] Example 1 Pretreatment dispersion buffer: 0.10 mol / L Tris, 0.15 mol / L NaCl, 0.01 mol / L EDTA, 20 mmol / L sodium citrate, pH adjusted to 7.8 with hydrochloric acid; Enzymatic hydrolysis buffer: 0.05 mol / L Tris, 0.02 mol / L NaCl, 3.5 mmol / L MgCl2, 0.6 mmol / L CaCl2, 0.06 mol / L EDTA, 25 mmol / L sodium citrate, pH adjusted to 7.4 with hydrochloric acid; The extraction method for high-purity salmon testes PDRN is as follows: (1) Remove blood vessels and fat impurities from frozen salmon testis tissue, and perform ultra-low temperature pulverization at -20℃ (pulverize to a paste, with no visible hard tissue blocks). Add 8 times the mass of frozen salmon testis tissue to pretreatment dispersion buffer, and perform high-pressure homogenization twice at 0~4℃ and 100MPa for 10 min each time to obtain DNA lysate. Centrifuge at 0~4℃ and 6000r / min for 12 min to separate the precipitate. (2) Disperse the precipitate in step (1) in 6 times the mass of the enzyme hydrolysis buffer, add trypsin and RNase A for enzymatic hydrolysis. The concentration of trypsin after addition is 4 g / L, the concentration of RNase A after addition is 0.08 g / L, the hydrolysis temperature is 37℃, the hydrolysis time is 4 h, and the enzyme hydrolysate is obtained. The enzyme is then inactivated at 65℃ for 8 min. (3) After step (2), DNase I was added to the enzyme hydrolysate. The concentration of DNase I after addition was 0.06 g / L. The reaction was carried out at 52℃ for 3.5 h, and then cooled to 42℃ for 1.5 h. After the reaction was completed, EDTA-2Na was added to terminate the enzyme digestion reaction. The concentration of EDTA-2Na after addition was 18 mmol / L. The supernatant was obtained by centrifugation at 0~4℃ and 12000 r / min for 12 min. (4) Add 2 times the volume of ethanol to the supernatant of step (3) for precipitation, centrifuge at 10000r / min for 10min, collect the precipitate, freeze dry to obtain PDRN from salmon testes.

[0023] Example 2 The pretreatment dispersion buffer and the enzyme digestion buffer are the same as in Example 1; The extraction method for high-purity salmon testes PDRN is as follows: (1) Remove blood vessels and fat impurities from frozen salmon testis tissue, and perform ultra-low temperature pulverization at -20℃ (pulverize to a paste, with no visible hard tissue blocks). Add 10 times the mass of frozen salmon testis tissue to pretreatment dispersion buffer, and perform high-pressure homogenization twice at 0~4℃ and 120MPa for 5 min each time to obtain DNA lysate. Centrifuge at 0~4℃ and 6000r / min for 12 min to separate and obtain the precipitate. (2) Disperse the precipitate in step (1) in 6 times the mass of the enzyme hydrolysis buffer, add trypsin and RNase A for enzymatic hydrolysis. The concentration of trypsin after addition is 3 g / L, the concentration of RNase A after addition is 0.1 g / L, the enzymatic hydrolysis temperature is 37℃, the enzymatic hydrolysis time is 4.5 h, and the enzyme hydrolysate is obtained. The enzyme is then inactivated at 65℃ for 8 min. (3) After step (2), DNase I was added to the enzyme hydrolysate after enzyme inactivation. The concentration of DNase I after addition was 0.08 g / L. The reaction was carried out at 50℃ for 4 h, and then cooled to 40℃ for 2 h. After the reaction was completed, EDTA-2Na was added to terminate the enzyme digestion reaction. The concentration of EDTA-2Na after addition was 16 mmol / L. The supernatant was obtained by centrifugation at 0~4℃ and 12000 r / min for 12 min. (4) Add 2 times the volume of ethanol to the supernatant of step (3) for precipitation, centrifuge at 10000r / min for 10min, collect the precipitate, freeze dry to obtain PDRN from salmon testes.

[0024] Example 3 The pretreatment dispersion buffer and the enzyme digestion buffer are the same as in Example 1; The extraction method for high-purity salmon testes PDRN is as follows: (1) Remove blood vessels and fat impurities from frozen salmon testis tissue, and perform ultra-low temperature pulverization at -20℃ (pulverize to a paste, with no visible hard tissue blocks). Add 8 times the mass of frozen salmon testis tissue to pretreatment dispersion buffer, and perform high-pressure homogenization twice at 0~4℃ and 80MPa for 15 min each time to obtain DNA lysate. Centrifuge at 0~4℃ and 6000r / min for 12 min to separate the precipitate. (2) Disperse the precipitate in step (1) in 6 times the mass of the enzyme hydrolysis buffer, add trypsin and RNase A for enzymatic hydrolysis. The concentration of trypsin after addition is 5 g / L, the concentration of RNase A after addition is 0.06 g / L, the enzymatic hydrolysis temperature is 37℃, the enzyme hydrolysis time is 3.5 h, and the enzyme hydrolysate is obtained. Inactivate the enzyme at 65℃ for 8 min. (3) After step (2), DNase I was added to the enzyme hydrolysate after enzyme inactivation. The concentration of DNase I after addition was 0.1 g / L. The reaction was carried out at 55℃ for 3 h, and then cooled to 45℃ for 1 h. After the reaction was completed, EDTA-2Na was added to terminate the enzyme digestion reaction. The concentration of EDTA-2Na after addition was 18 mmol / L. The supernatant was obtained by centrifugation at 0~4℃ and 12000 r / min for 12 min. (4) Add 2 times the volume of ethanol to the supernatant of step (3) for precipitation, centrifuge at 10000r / min for 10min, collect the precipitate, freeze dry to obtain PDRN from salmon testes.

[0025] Comparative Example 1 Unlike Example 1, Comparative Example 1 used the same solution for pretreatment dispersion buffer and enzymatic digestion buffer, with the following composition: 0.10 mol / L Tris, 0.15 mol / L NaCl, 0.01 mol / L EDTA, and pH adjusted to 7.8 with hydrochloric acid; the remaining steps and parameter settings were the same as in Example 1, and PDRN derived from salmon testes was prepared.

[0026] Comparative Example 2 Unlike Example 1, sodium citrate was not added to the pretreatment dispersion buffer in Comparative Example 1. The remaining steps and parameter settings were the same as in Example 1, and PDRN derived from salmon testes was prepared.

[0027] Comparative Example 3 Unlike Example 1, sodium citrate was not added to the enzyme digestion buffer in Comparative Example 3. The remaining steps and parameter settings were the same as in Example 1, and PDRN derived from salmon testes was prepared.

[0028] Comparative Example 4 Unlike Example 1, in Comparative Example 4, step (3) was reacted at 52°C for 5 hours, and the remaining steps and parameter settings were the same as in Example 1, to prepare PDRN derived from salmon testes.

[0029] Comparative Example 5 Unlike Example 1, in Comparative Example 5, step (3) was reacted at 42°C for 5 hours, and the remaining steps and parameter settings were the same as in Example 1, to prepare PDRN derived from salmon testes.

[0030] Performance testing The yield (product mass / salmon testis tissue) and purity (OD) of PDRN derived from salmon testes prepared in Examples 1-3 and Comparative Examples 1-5 were compared. 260 / OD 280 OD 260 and OD 280The absorbance and molecular weight distribution of high-purity salmon testis-derived PDRN at wavelengths of 260 nm and 280 nm were tested, and the results are shown in Table 1 below: Table 1 Performance test results of PDRN derived from salmon testes As shown in Table 1, the method of this invention for extracting PDRN from salmon testes yields over 85% PDRN with a molecular weight concentrated between 40 and 60 kDa, while the proportions of PDRN with molecular weights less than 40 kDa and greater than 60 kDa are relatively small. Furthermore, the method of this invention exhibits a relatively high yield, and the purity of all samples meets the requirements. Only Comparative Example 5 shows slightly lower purity, which may be related to the presence of a small amount of impurities in the high molecular weight PDRN. The increased content of PDRN with molecular weights less than 40 kDa in Comparative Examples 1-3 may be due to the absence of sodium citrate in the pretreatment dispersion buffer and / or the enzyme digestion buffer, which fails to inhibit the enzyme activity inherent in the raw materials, leading to spontaneous degradation into smaller PDRN molecules. Additionally, the high content of smaller PDRN molecules can also cause component loss, affecting the yield. Comparative Examples 4 and 5 employ a one-step enzymatic digestion reaction; excessive or insufficient digestion can affect the yield, purity, and molecular weight distribution of PDRN.

[0031] Application Examples A functional cosmetic composition is formulated as follows: 1 wt% active ingredient (PDRN derived from salmon testes), 10 wt% glycerin, 10 wt% sodium hyaluronate (500-1000 kDa), 0.1 wt% EDTA-2Na, with the balance being water. The functional cosmetic composition was prepared according to the above formulation using PDRN derived from salmon testes prepared in Examples 1-3 and Comparative Examples 1-5 as the active ingredient; and human efficacy tests were conducted. Test conditions: From May 8, 2025 to June 8, 2025, 80 female volunteers aged 2 to 50 years with dry, dull skin and obvious facial and eye wrinkles (level 2 to 4) were selected. Eight test groups were set up, including Examples 1 to 3 and Comparative Examples 1 to 5 (functional cosmetic compositions), with 10 people in each group. After cleansing their faces in the morning and evening, they applied the functional cosmetic compositions prepared with Examples 1 to 3 and Comparative Examples 1 to 5 as active ingredients. No other cosmetics were used during the test period, which lasted for 4 weeks.

[0032] (1) The skin stratum corneum moisture content was tested at 0, 14 and 28 days using a skin stratum corneum moisture content tester. The higher the value, the higher the stratum corneum moisture content. The results are shown in Table 2 below: Table 2 Results of Facial Skin Moisture Content Test (2) Facial skin elasticity (ISE value) was tested at 0, 7, 14 and 28 days using a skin elasticity tester. The higher the value, the better the skin condition. The results are shown in Table 3 below: Table 3. Results of Facial Skin Elasticity (ISE Value) Test (3) Eye wrinkle area test: The area of ​​eye wrinkles was analyzed and recorded using the VISIA-CR instrument and image processing system. The smaller the wrinkle area, the better the skin condition. The results are shown in Table 4. Table 4. Area of ​​wrinkles around the eyes (mm) 2 Test Result Table As shown in Tables 2-4 above, the functional cosmetic compositions corresponding to Examples 1-3 exhibit good technical effects in improving facial stratum corneum water content, facial skin elasticity, and eye wrinkles. The poorer improvement effects of Comparative Examples 1-4 compared to Example 1 may be related to their higher content of low molecular weight (<40kDa) PDRN. Although low molecular weight PDRN is beneficial for facial absorption, it is quickly lost and difficult to accumulate in the dermis, resulting in poor long-term repair effects. Comparative Example 5 has a high PDRN content >60kDa, leading to poor transdermal penetration. Most of it adheres to the extracellular matrix, resulting in poor absorption by skin cells and a poor effect on facial skin improvement. Therefore, PDRN with a molecular weight of 40-60kDa has a better effect on improving skin condition.

[0033] The above embodiments are merely preferred embodiments of the present invention and should not be construed as limiting the scope of protection of the present invention. Any non-substantial changes and substitutions made by those skilled in the art based on the present invention shall fall within the scope of protection claimed by the present invention.

Claims

1. A method for extracting high-purity PDRN from salmon testes, characterized in that, Includes the following steps: (1) Remove blood vessels and fat impurities from frozen salmon testis tissue, perform cryogenic pulverization at -20~-25℃, add to pretreatment dispersion buffer, perform high pressure homogenization at 80~120MPa to obtain DNA lysate, and centrifuge to obtain precipitate. (2) Disperse the precipitate from step (1) in a special buffer for enzymatic hydrolysis, add protease and RNase A for enzymatic hydrolysis to obtain the hydrolysate, and then perform enzyme inactivation treatment; (3) After the enzyme inactivation in step (2), DNase I was added to the enzyme hydrolysate for enzymatic digestion. After the enzyme digestion reaction was completed, the stop solution was added to terminate the enzyme hydrolysis reaction, and the supernatant was obtained by centrifugation. (4) Add 2 to 3 times the volume of ethanol to the supernatant of step (3) for precipitation, centrifuge to collect the precipitate, and obtain PDRN from salmon testes.

2. The method for extracting high-purity PDRN from salmon testes according to claim 1, characterized in that, The pretreatment dispersion buffer described in step (1) includes the following concentrations of substances: 0.05~0.15 mol / L Tris, 0.1~0.2 mol / L NaCl, 0.001~0.1 mol / L EDTA, and 15~20 mmol / L sodium citrate, with the pH adjusted to 7.5~8.0 using hydrochloric acid.

3. The method for extracting high-purity PDRN from salmon testes according to claim 1, characterized in that, The enzyme-specific buffer solution mentioned in step (2) includes the following concentrations of substances: 0.04~0.06 mol / L Tris, 0.01~0.05 mol / L NaCl, 3~4 mmol / L MgCl2, 0.4~0.7 mmol / L CaCl2, 0.05~0.1 mol / L EDTA, and 20~30 mmol / L sodium citrate, with the pH adjusted to 7.3~7.5 using hydrochloric acid.

4. The method for extracting high-purity PDRN from salmon testes according to claim 1, characterized in that, The protease in step (2) is one or more of trypsin, papain and proteinase K. The concentration of the protease after addition is 2~5 g / L, the concentration of RNase A after addition is 0.05~0.1 g / L, the enzymatic hydrolysis temperature is 35~45℃, and the enzymatic hydrolysis time is 3~5 h.

5. The method for extracting high-purity PDRN from salmon testes according to claim 1, characterized in that, In step (3), the concentration of DNase I after addition is 0.05~0.1g / L, and the enzyme digestion reaction conditions are 50~55℃ for 3~4h, and then the temperature is lowered to 40~45℃ for 0.5~2h.

6. The method for extracting high-purity PDRN from salmon testes according to claim 1, characterized in that, In step (1), the mass ratio of frozen salmon testis tissue to pretreatment dispersion buffer is 1:6~10. The tissue is homogenized under high pressure at 0~4℃ 1~2 times, each time for 5~15 min.

7. The method for extracting high-purity salmon testes-derived PDRN according to claim 1, characterized in that, In step (2), the mass ratio of the precipitate and the enzyme digestion buffer is 1:5~8; in step (1), the centrifugation conditions are 0~4℃, 6000~8000r / min, 10~15min; in step (3), the centrifugation conditions are 0~4℃, 10000~12000r / min, 10~15min; in step (4), the centrifugation conditions are 8000~10000r / min, 8~10min.

8. The method for extracting high-purity salmon testes-derived PDRN according to claim 1, characterized in that, In step (2), the enzyme inactivation treatment conditions are 60~70℃ for 8~10min; in step (3), EDTA-2Na is added to terminate the enzyme digestion reaction, and the concentration of EDTA-2Na after addition is 15~20mmol / L.

9. The application of high-purity salmon testis-derived PDRN prepared by the preparation method according to any one of claims 1 to 8 in improving skin condition.

10. The application according to claim 9, characterized in that, The high-purity salmon testis-derived PDRN is used to prepare pharmaceuticals or cosmetics, especially cosmetics with functions such as moisturizing, increasing facial elasticity, firming skin, and anti-wrinkle.

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