CN120131609A

CN120131609A - Application of N1, N10-dicaffeoyl spermidine in the preparation of anti-aging drugs, health products or functional foods

Info

Publication number: CN120131609A
Application number: CN202510558572.XA
Authority: CN
Inventors: 马学琴; 王伟彪; 高涛; 杨志刚; 赵建军
Original assignee: Ningxia Medical University
Current assignee: Ningxia Medical University
Priority date: 2025-04-30
Filing date: 2025-04-30
Publication date: 2025-06-13

Abstract

The invention discloses an application of N1, N10-dicaffeoylspermidine in preparing anti-aging medicines, health products or functional foods. Belongs to the technical field of biological medicine. Animal experiments prove that the N1, N10-dicaffeoylspermidine has an anti-aging effect, can obviously improve the working memory and the spatial memory capacity of D-galactose induced aging mice, obviously reduce the expression level of P53, P21 and P16 proteins of brain tissue aging markers of the mice, obviously reduce the contents of inflammatory factors 1L-1 beta, 1L-6 and TNF-alpha of the aging mice, and obviously improve the contents of oxidative stress factors GSH and SOD. The N1, N10-dicaffeoylspermidine is used for preparing medicaments, health products and/or functional foods for delaying aging, preventing and/or treating the aging, provides a new choice for aging resistance, and has important practical application value in the fields of medicine and aging resistance.

Description

Application of N1, N10-dicaffeoylspermidine in preparing anti-aging medicines, health products or functional foods

Technical Field

The invention belongs to the technical field of biological medicines, and particularly relates to application of N1, N10-dicaffeoylspermidine in preparation of anti-aging medicines, health products or functional foods.

Background

Current anti-aging interventions mainly include Senolytics (senescent cell scavengers), such as dasatinib+quercetin combinations, nad+ precursors, such as Nicotinamide Mononucleotide (NMN) and Nicotinamide Riboside (NR), mTOR inhibitors, such as rapamycin derivatives, and the like. In addition, natural health products such as resveratrol, curcumin, collagen peptide, etc. are provided. However, the existing medicines have the problems of limited effect, obvious side effect and the like.

Natural products become the core direction for the research and development of anti-aging drugs due to the chemical diversity and low toxicity characteristics. At present, natural compounds such as ginsenoside Rg1, apigenin and the like have been reported to have anti-aging effects. The plant components (such as green tea polyphenol and salidroside) for long-term eating or medical use are verified by history, the risk of toxic and side effects is far lower than that of synthetic medicines, and the method is suitable for long-term intervention.

N1, N10-dicaffeoylspermidine is a monomer active ingredient separated from medicinal and edible traditional Chinese medicines such as medlar and the like by the inventor, the research report on the biological activity of N1, N10-dicaffeoylspermidine is less at present, and the related record of N1, N10-dicaffeoylspermidine for preventing and treating aging is not seen yet.

Disclosure of Invention

In view of the above, the invention provides the application of N1, N10-dicaffeoylspermidine in preparing medicines, health products or functional foods for preventing and treating aging. The N1, N10-dicaffeoylspermidine has obvious effect of relieving aging through early animal experiments. Further, the compound can be developed into a medicament for delaying, preventing and/or treating aging, or a health-care product or functional food for delaying, preventing and/or treating aging, and has great clinical application value.

N1, N10-dicaffeoylspermidine has a molecular formula of C ₂₅H₃₁N₃O₆, a molecular weight of 469.22, and is called (E)-3-(3 ,4-dihydroxyphenyl)-N-(3-((4-((E)-3-(3 ,4- dihydroxyphenyl)acrylamido)butyl)amino)propyl)acrylamide, N1, N10-bis (caffeoyl) spermidine or N1, N10-di-caffeoylspermidine. The chemical structural formula is shown as formula (1):

(1)

In order to achieve the above purpose, the present invention adopts the following technical scheme:

the application of N1, N10-dicaffeoylspermidine in preparing medicines for preventing and treating aging, or in preparing health products or functional foods for preventing and treating aging.

Further, N1, N10-dicaffeoylspermidine can significantly increase the spontaneous alternation reaction rate of the aging mice in the Y maze experiment, i.e. improve the working memory capacity of the mice.

Furthermore, the N1, N10-dicaffeoylspermidine can obviously reduce the escape latency of the aging mice in Morris water maze experiments and increase the number of crossing platforms, namely improve the spatial memory capacity of the mice.

Further, N1, N10-dicaffeoylspermidine can significantly reduce the expression level of P53, P21 and P16 proteins of brain tissue senescence markers of aging mice.

Further, N1, N10-dicaffeoylspermidine can remarkably reduce the contents of brain tissue inflammatory factors 1L-1 beta, 1L-6 and TNF-alpha of aging mice.

Furthermore, N1, N10-dicaffeoylspermidine can obviously improve the content of oxidation stress factors SOD and GSH in brain tissues of the aging mice.

An anti-aging pharmaceutical product comprising N1, N10-dicaffeoylspermidine.

Further, the dosage form is a pharmaceutically acceptable oral dosage form, an external patch or an injection.

An antiaging health product comprises N1, N10-dicaffeoylspermidine.

An antiaging functional food comprises N1, N10-dicaffeoylspermidine.

Compared with the prior art, the technical scheme has the beneficial effects that in the dosage range of 10-25 mg/kg given to experimental animals, N1, N10-dicaffeoylspermidine can obviously improve the working memory and the spatial memory capacity of D-galactose induced aging mice, obviously reduce the protein expression level of P53, P21 and P16 of brain tissue aging markers of the mice, obviously reduce the contents of inflammatory factors 1L-1 beta, 1L-6 and TNF-alpha of the aging mice and obviously improve the contents of oxidative stress factors SOD and GSH. The above results prove that the N1, N10-dicaffeoylspermidine has anti-aging effect and can be used for preparing medicaments, health products and/or functional foods for delaying, preventing and/or treating aging.

Drawings

In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings that are required to be used in the embodiments or the description of the prior art will be briefly described below, and it is obvious that the drawings in the following description are only embodiments of the present invention, and that other drawings can be obtained according to the provided drawings without inventive effort for a person skilled in the art.

FIG. 1 shows the effect of N1, N10-dicaffeoylspermidine on body weight of aging mice (C is the normal group, M is the model group, P is the positive drug piracetam group, NNDCP-H is N1, N10-dicaffeoylspermidine high dose group, NNDCP-L is N1, N10-dicaffeoylspermidine low dose group; ^# P <0.05 compared to the normal group; P <0.05; P <0.01 compared to the model group)

FIG. 2 is a graph showing the effect of Y maze experiments on spontaneous alternation response rates in senescent mice (C is the normal group, M is the model group, P is the positive drug piracetam group, NNDCP-H is N1, N10-dicaffeoylspermidine high dose group, NNDCP-L is N1, N10-dicaffeoylspermidine low dose group; ^#p<0.05,^##p<0.01,^### P <0.001 compared to the normal group; P <0.01, P <0.001 compared to the model group)

FIG. 3 is a chart showing Morris water maze test showing the effect of N1, N10-dicaffeoylspermidine on the escape latency and number of passes through the platform in senescent mice (C is the normal group, M is the model group, P is the positive drug piracetam group, NNDCP-H is N1, N10-dicaffeoylspermidine high dose group, NNDCP-L is N1, N10-dicaffeoylspermidine low dose group; ^### P <0.001 compared to the normal group; P <0.001 compared to the model group)

FIG. 4 is a graph showing the effect of N1, N10-dicaffeoylspermidine on the expression levels of the senescence markers P53, P21 and P16 in brain tissue of senescent mice (C is the normal group, M is the model group, NNDCP-H is N1, N10-dicaffeoylspermidine high dose group, NNDCP-L is N1, N10-dicaffeoylspermidine low dose group; ^#p<0.05,^## P <0.01 compared to the normal group; P <0.05, P <0.01, P <0.001 compared to the model group)

FIG. 5 is a graph showing the effect of N1, N10-dicaffeoylspermidine on inflammatory factor 1L-1 beta, 1L-6 and TNF-alpha levels in brain tissue of aging mice (C is normal, M is model, P is positive drug piracetam, NNDCP-H is N1, N10-dicaffeoylspermidine high dose, NNDCP-L is N1, N10-dicaffeoylspermidine low dose; ^##p<0.01,^### P <0.001 compared to normal; P <0.05, P <0.01, P <0.001 compared to model)

FIG. 6 is a graph showing the effect of N1, N10-dicaffeoylspermidine on the levels of oxidative stress factor SOD, GSH in brain tissue of aging mice (C is the normal group, M is the model group, P is the positive drug piracetam group, NNDCP-H is N1, N10-dicaffeoylspermidine high dose group, NNDCP-L is N1, N10-dicaffeoylspermidine low dose group; P <0.05, P <0.001 compared to model group)

Detailed Description

The following description of the embodiments of the present invention will be made clearly and completely with reference to the accompanying drawings, in which it is apparent that the embodiments described are only some embodiments of the present invention, but not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.

N1, N10-dicaffeoylspermidine used in the following examples is a compound represented by the above formula (1) and can be obtained commercially or experimentally self-made.

The required medicament is a conventional experimental medicament purchased from a commercial channel, and the unrecited experimental method is a conventional experimental method and is not described in detail herein.

Example 1

An antiaging medicine in the form of oral preparation comprises N1, N10-dicaffeoylspermidine.

Example 2

An antiaging medicine in the form of injection comprises N1, N10-dicaffeoylspermidine.

Example 3

An antiaging medicine in the form of topical patch comprises N1, N10-dicaffeoylspermidine.

Example 4

An antiaging health product comprises N1, N10-dicaffeoylspermidine.

Example 5

An antiaging functional food comprises N1, N10-dicaffeoylspermidine.

The following animal experiments further illustrate the effects of examples 1 to 5 above:

1. Design of animal experiment

D-galactose induced aging mouse model was used. After administration of N1, N10-dicaffeoylspermidine by intragastric administration, the anti-aging effect of N1, N10-dicaffeoylspermidine was evaluated by the body weight of mice, Y maze test, morris water maze test, the expression level of proteins of senescence markers P53, P21 and P16, the contents of inflammatory factors 1L-1 beta, 1L-6 and TNF-alpha, the contents of oxidative stress factors SOD and GSH, and the like.

2. Experimental procedure

1. Experimental animal

Healthy SPF-class C57BL/6 mice, male, body weight 20-30 g, were taken, supplied by the university of Ningxia medical laboratory animal center (laboratory animal use license SYSK (Ning) 2020-0001), were kept in an SPF-class barrier environment, were lighted 12 h, were dark 12 h, and were fed freely with water.

2. Experimental method

D-galactose induced aging mouse model was used. After 3 days of adaptive feeding, the 40 mice were randomly divided into 5 groups, normal group (normal saline), model group (normal saline), piracetam positive drug group (468 mg/kg/d), N1, N10-dicaffeoylspermidine low dose group (NNDCP-L, 10 mg/kg/d), N1, N10-dicaffeoylspermidine high dose group (NNDCP-H, 25 mg/kg/d), 8 animals each. Piracetam and N1, N10-dicaffeoylspermidine are dissolved in physiological saline and administered by lavage at 10 mL/kg. Mice in groups other than the normal group induced aging model by daily subcutaneous injection of D-galactose (200 mg/kg) at fixed time points, normal group mice were injected subcutaneously and lavaged with equal volumes of physiological saline. The subcutaneous injection and the gastric lavage are carried out once a day, and the injection is continuously molded for 6 weeks.

3. Y maze experiment

After 6 weeks of administration of each group of mice, the effect of N1, N10-dicaffeoylspermidine on the working memory capacity of the aged mice was examined by Y maze test, and evaluation was carried out by calculating the spontaneous alternation response rate of each mouse.

4. Morris water maze experiment

6 Weeks after each group of mice was dosed, the effect of N1, N10-dicaffeoylspermidine on spatial memory capacity of the senescent mice was examined by Morris experiment (5 days of continuous measurement), and evaluated by escape latency and number of passes across the platform.

5. Determination of expression levels of senescence markers P53, P21 and P16 proteins

After 6 weeks of administration and behavioral testing, the mice were sacrificed after taking serum, the whole brain was taken after head breaking on ice, immediately frozen in liquid nitrogen, and transferred to-80 ℃ for later use after the experiment was completed. Weighing a proper amount of brain tissue of a mouse, adding a Lysis Buffer containing 0.1% protease inhibitor and 1% of 100mM PMSF, manually homogenizing for 30-50 times on and off by a glass homogenizer, transferring the tissue homogenate to a centrifuge tube precooled by 1.5 mL, centrifuging at 4 ℃ for 5min, transferring the supernatant to a new precooled centrifuge tube to obtain a whole protein extract, quantifying the protein (BCA method), adding 20% of 5X SDS-PAGE protein loading Buffer and purified water into the whole protein extract to prepare protein loading liquid with the same total protein concentration, loading SDS-PAGE gel, electrophoresis, transferring a membrane by a wet transfer method, respectively incubating primary antibodies of P53, P21 and P16 proteins overnight after membrane sealing, incubating secondary antibodies, and finally ECL developing, exposing and photographing.

6. 1L-1 beta, 1L-6, TNF-alpha, SOD and GSH content determination

After 6 weeks of administration and behavioral testing, the mice were sacrificed after taking serum, the whole brain was taken after head breaking on ice, immediately frozen in liquid nitrogen, and transferred to-80 ℃ for later use after the experiment was completed. Taking mouse brain tissue, grinding uniformly by a tissue grinder to prepare brain tissue homogenate, centrifuging at 4 ℃ for 10min at 5000 xg/min, collecting supernatant, and detecting the contents of 1L-1 beta, 1L-6, TNF-alpha, SOD and GSH in the supernatant by using a kit.

7. Data processing

The data processing adopts imageJ and GRAPHPAD PRISM 9.5.5 to carry out data analysis and statistics, protein bands are analyzed by using imageJ, and the content expression condition of each group of target proteins is expressed by the ratio of the gray value of the target protein band to the gray value of the reference protein band.

The results are all expressed as mean ± standard deviation (x±s), the comparison of the differences between groups is data processed using one-way ANOVA, the differences are statistically significant with p <0.05 as the standard.

3. Experimental results

1. Effects on mouse body weight

The effect of N1, N10-dicaffeoylspermidine on the body weight of mice is shown in FIG. 1.

The mice in the model group had significantly reduced body weight (p < 0.05) compared to the control group after 6 weeks of dosing. Compared with the model group, the piracetam positive drug group has an ascending trend, but the difference has no statistical significance, and the weight of the mice in the N1, N10-dicaffeoylspermidine group is obviously increased (p <0.05 or p < 0.01), so that the N1, N10-dicaffeoylspermidine has obvious improvement effect on the weight reduction of the mice caused by D-galactose induced aging mice modeling.

2. Effects on spontaneous alternative response rates in mice

The results of the Y maze experiment examining the effect of N1, N10-dicaffeoylspermidine on spontaneous alternation rate of aged mice are shown in FIG. 2.

The results show that compared with the control group, the spontaneous alternation rate of the mice in the model group is obviously reduced (p < 0.001), which indicates that the aging and modeling of the mice are successful, and the D-galactose can obviously damage the working memory capacity of the mice. The spontaneous alternation rates were significantly increased in both the positive drug group and the N1, N10-dicaffeoylspermidine group mice (p <0.001 or p < 0.01) compared to the model group. The positive medicine and N1, N10-dicaffeoylspermidine are proved to have obvious protection effect on the working memory capacity injury of aged mice.

3. Influence on escape latency and number of crossing platforms of mice

The Morris water maze experiment examined the effect of N1, N10-dicaffeoylspermidine on the escape latency and the number of platform crossing times of aged mice, and the results are shown in FIG. 3.

The results show that model mice have an increased escape latency and significantly reduced number of platform crossings (p < 0.001) compared to control mice. Compared to the model group, the positive drug group mice had a significantly increased escape latency (p < 0.001), the N1, N10-dicaffeoylspermidine group mice had a significantly decreased escape latency, the N1, N10-dicaffeoylspermidine high dose group mice had a significantly increased number of passes (p < 0.001), and the N1, N10-dicaffeoylspermidine low dose group mice had a tendency to have an increased number of passes through the platform. The above description shows that N1, N10-dicaffeoylspermidine can improve spatial memory in aging mice to some extent.

4. Effect on expression levels of senescence markers P53, P21 and P16 proteins in mouse brain tissue

The effect of N1, N10-dicaffeoylspermidine on the expression level of the senescence markers P53, P21 and P16 proteins in mouse brain tissue is shown in FIG. 4.

The results show that compared with the control group, the expression level of the senescence markers P53, P21 and P16 in the brain tissue of the mice in the model group is obviously improved (P <0.05 or P < 0.01), which indicates that the senescence model of the D-galactose mice is successfully established. The expression levels of P53, P21 and P16 in brain tissue of mice in the N1, N10-dicaffeoylspermidine group were all significantly reduced to different extents compared to the model group (P <0.05, P <0.01 or P < 0.001). The results indicate that N1, N10-dicaffeoylspermidine can improve D-galactose induced aging in mice.

5. Effects on the levels of inflammatory factors 1L-1 beta, 1L-6 and TNF-alpha in mouse brain tissue

The effect of N1, N10-dicaffeoylspermidine on the levels of inflammatory factors 1L-1 beta, 1L-6 and TNF-alpha in brain tissue of aged mice is shown in FIG. 5.

The results showed that the levels of 1L-1. Beta. And TNF-. Alpha.in the brain tissue of mice in the model group were significantly increased (p <0.01 or p < 0.001) compared to the control group, suggesting that the mechanism of D-galactose-induced aging in mice may be associated with inflammation of brain tissue. The levels of 1L-1β, 1L-6 and TNF- α were significantly reduced in the brain tissue of mice from both the positive drug group and the N1, N10-dicaffeoylspermidine group compared to the model group (p <0.05, p <0.01 or p < 0.001). The results indicate that the anti-aging mechanism of positive drugs and N1, N10-dicaffeoylspermidine may be related to its reduction of the content of 1L-1 beta, 1L-6 and TNF-alpha in brain tissue, reducing inflammation.

6. Influence on the content of SOD and GSH of mice brain tissue oxidative stress factor

The effect of N1, N10-dicaffeoylspermidine on the content of SOD and GSH in brain tissue of aged mice is shown in FIG. 6.

The results show that compared with the control group, the content of SOD and GSH in the brain tissue of the mice in the model group has a reduced tendency, which suggests that the mechanism of aging of the mice caused by D-galactose is possibly related to the oxidative stress of the brain tissue. The content of SOD in the brain tissue of the mice in the positive drug group tended to be increased compared with the model group, while the content of SOD in the brain tissue of the mice in the N1, N10-dicaffeoylspermidine group was significantly increased (p < 0.05), and the content of GSH in the brain tissue of the mice in the positive drug group and the N1, N10-dicaffeoylspermidine group was significantly increased (p <0.05 or p < 0.001). The results show that the positive drugs and N1, N10-dicaffeoylspermidine have the action mechanism of resisting aging of mice, which is possibly related to increasing the content of SOD and GSH in brain tissues, thereby reducing oxidative stress.

During the whole animal administration process, the death phenomenon of the mice is avoided, and the mice have no abnormal appearance and behavior.

In the present specification, each embodiment is described in a progressive manner, and each embodiment is mainly described in a different point from other embodiments, and identical and similar parts between the embodiments are all enough to refer to each other.

The previous description of the disclosed embodiments is provided to enable any person skilled in the art to make or use the present invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the generic principles defined herein may be applied to other embodiments without departing from the spirit or scope of the invention. Thus, the present invention is not intended to be limited to the embodiments shown herein but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. Application of N1,N10-dicaffeoylspermidine in the preparation of medicines for preventing and treating aging, or in the preparation of health products or functional foods for preventing and treating aging.

2. The use according to claim 1, characterized in that N1,N10-dicaffeoylspermidine can significantly increase the spontaneous alternation reaction rate of aging mice in a Y-maze test, that is, improve their working memory ability.

3. The use according to claim 1, characterized in that N1,N10-dicaffeoylspermidine can significantly reduce the escape latency of aging mice in the Morris water maze test and increase the number of times they cross the platform, that is, improve their spatial memory ability.

4. The use according to claim 1, characterized in that N1,N10-dicaffeoylspermidine can significantly reduce the expression levels of P53, P21 and P16 proteins, which are aging markers in the brain tissue of aging mice.

5. The use according to claim 1, characterized in that N1,N10-dicaffeoylspermidine can significantly reduce the content of inflammatory factors 1L-1β, 1L-6 and TNF-α in the brain tissue of aging mice.

6. The use according to claim 1, characterized in that N1, N10-dicaffeoylspermidine can significantly increase the content of oxidative stress factors SOD and GSH in brain tissue of aging mice.

7. An anti-aging drug, characterized in that it comprises N1,N10-dicaffeoylspermidine.

8. The drug according to claim 7, characterized in that the dosage form is an oral dosage form, an external patch or an injection dosage form permitted in pharmaceutical science.

9. An anti-aging health product, characterized in that it comprises N1,N10-dicaffeoylspermidine.

10. An anti-aging functional food, characterized in that it comprises N1,N10-dicaffeoylspermidine.