阅读说明：本技术 一种丹酚酸a类似物及其作为抗氧化剂的用途 (Salvianolic acid A analogue and application thereof as antioxidant ) 是由 谢志忠 曹轩 丘金梅 吴忱昊 牛盾 于 2019-10-14 设计创作，主要内容包括：本发明提供了一类新的具有抗氧化活性的丹酚酸A类似物,其可通过上调CAT和SOD等抗氧化酶的表达以及增加非酶抗氧化物质GSH的产生,直接或间接地清除ROS,体现了较好的抗氧化能力,拓宽了现有抗氧化剂的范围,并可作为先导化合物继续优化。同时本发明化合物即使在低浓度(10μM/kg·天)下,也可以保护肝脏免受CCl<Sub>4</Sub>诱导的氧化应激损伤,具有良好的成药前景。(The invention provides a novel salvianolic acid A analogue with antioxidant activity, which can up-regulate the expression of antioxidase such as CAT, SOD and the likeAnd increase the production of non-enzymatic antioxidant GSH, and directly or indirectly scavenge ROS, thus embodying better antioxidant capacity, widening the range of the existing antioxidant, and being capable of being continuously optimized as a lead compound. At the same time, the compound of the invention can protect the liver from CCl even at low concentration (10 mu M/kg-day) 4 The induced oxidative stress injury has good patent medicine prospect.)

1. A compound of formula I, or a pharmaceutically acceptable salt thereof, having the structure:

wherein: r is selected from C₁-C₆Alkyl radical, C₁-C₆Alkenyl radical, C₁-C₆A haloalkyl group.

2. A compound of formula I according to claim 1, characterized in that: r is selected from C₁-C₄An alkyl group.

3. A compound of formula I according to claim 1, having the structure:

4. a process for the preparation of a compound of formula I according to claim 1, which reaction scheme is as follows:

5. a process according to claim 4 for the preparation of a compound of formula I, comprising the steps of:

the method comprises the following steps: performing Vilsmeier reaction on resveratrol in POCl3 and DMF to obtain aldehyde-based resveratrol (1);

step two: continuously introducing dry hydrogen chloride into the methyldopa in an alcohol solution ROH, heating and refluxing for reaction, and carrying out post-treatment to obtain methyldopate (2);

step three: dissolving the aldehyde resveratrol prepared in the step one and the methyldopate prepared in the step two in an organic solvent, refluxing and stirring for reaction, and performing post-treatment after the reaction to obtain the resveratrol methyldopa Schiff base (3)

Step four: dissolving the resveratrol methyldopa Schiff base prepared in the third step into an organic solvent, adding a reducing agent at normal temperature for reaction, and performing post-treatment after the reaction to obtain the compound shown in the formula I.

6. A process according to claim 5 for the preparation of compounds of formula I, characterized in that:

in the first step: resveratrol and POCl₃Molar volume ofThe ratio is 1:0.4-0.7, preferably 1: 0.5-0.6; the reaction temperature is 0-60 ℃, and room temperature is preferred; the reaction time is 0.5 to 2 hours, preferably 1 hour.

7. A process according to claim 5 or 6 for the preparation of compounds of formula I, characterized in that:

the reaction temperature in the second step is from room temperature to reflux temperature; the reaction time is 3-8 hours.

8. A process according to claim 5 or 6 for the preparation of compounds of formula I, characterized in that:

in the third step, the molar ratio of the aldehyde-based resveratrol (1) to the methyldopate (2) is 1:1-2, preferably 1:1.4-1.8, and most preferably 1: 1.6; the organic solvent is selected from one or more of methanol, ethanol, tetrahydrofuran and DMF, and most preferably methanol.

9. A process according to claim 5 or 6 for the preparation of compounds of formula I, characterized in that:

in the fourth step, the reducing agent is selected from one or more of sodium borohydride, lithium aluminum hydride, sodium cyanoborohydride, Pd/C and Raney nickel, and sodium borohydride is most preferred.

10. A pharmaceutical composition comprising a compound of formula I as described in any one of claims 1-3, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

11. Use of a compound of formula I as defined in any one of claims 1 to 3 or a pharmaceutical composition as defined in claim 9 for the preparation of an antioxidant.

Technical Field

The invention relates to the field of pharmaceutical chemistry, in particular to a salvianolic acid A analogue, a preparation method and application thereof.

Background

More and more studies show that anti-oxidation is an important step in preventing aging, because free radicals or oxidants break down cells and tissues, affect metabolic functions, and cause various health problems. Reactive oxygen Radicals (ROS), which contain one or more unpaired electrons in their outer orbitals, have short lifetimes and high chemical reactivity and have long been considered as harmful molecules, although recent studies have shown that they also play important role in physiological responses in certain molecules (oxid. Excessive free radicals directly damage sensitive tissues and are associated with a variety of pathological events, such as neurodegenerative diseases (biomed. pharmacother.2016,78:39-49), myocardial infarction (eur. heart j.,2017,38(41): 3105-. Oxidative stress refers to a severe imbalance between ROS production in the body and the antioxidant capacity of the cell. Superoxide dismutase (SOD), Catalase (CAT) and glutathione peroxidase-1 (GPx-1)) and non-enzymatic antioxidants (e.g. reduced Glutathione (GSH)) constitute an antioxidant stress network in vivo to minimize oxidative damage (Oxid Med celllangev.2016: 6235641; oxid Med Cell Longgev.2016: 6043038). It has been shown that a high intake of natural antioxidant-rich foods, such as plant polyphenols, reduces the risk of diseases associated with oxidative stress (CellBiochem. Biophys.,2015,73(3): 649-664).

Salvia miltiorrhiza, a well-known traditional Chinese herbal medicine, has been in clinical use for over 2000 years in China. Salvianolic Acid A (SAA) is one of the main water-soluble polyphenol compounds extracted from dried root and rhizome of Salvia miltiorrhiza. It has been shown to be a strong antioxidant, reducing oxidative stress in vivo and having a broad protective effect on liver fibrosis (Liver.2001,21(6): 384-) -390, ischemia reperfusion injury (Liver.2016,21(6): 384-) -390), Alzheimer's disease (Pharm.biol.,2016,54(1):18-24), however, the SAA content in Salvia miltiorrhiza is low (< 0.05%) and it cannot be widely used in clinical trials due to its chemical structural instability caused by the polyphthalanol moiety (Arevie w.journal of Ethnopharmacology,2018,225: 18-30).

Resveratrol (RES) is a natural trans-stilbene compound that has not attracted interest until the 1990's. Growth studies involving the biological activity of RES have been conducted over the last two decades and have shown that RES has a number of health-promoting effects in both animals and humans (Neurochem int, 2015,89: 75-82). The mechanism by which RES exerts such a wide range of beneficial effects is not clear. In some studies, RES, like other polyphenolic compounds, has a powerful ability to scavenge free radicals, and the antioxidant capacity of RES is considered to be critical to its biological effects (Recent Pat food NuodNutr Agric.,2013,5(2): 144-.

Resveratrol is a molecule recognized as having antioxidant activity, but it is associated with Cu in the presence of copper ions²⁺In which phenolic hydroxyl group is oxidized to phenoxy radical, and Cu²⁺Conversion to cu (i). Cu (I) can form a complex with DNA, and the complex is degraded under the action of resveratrol phenoxy radical, so that the resveratrol shows pro-oxidation function (biochem. Soc. T.2007,35, 1156-1160). However, studies have shown that the solubility of resveratrol is not ideal, which limits the absorption of the molecule, and that the bioavailability of resveratrol in humans by oral administration is very low (far below 1%) (Molecules 2014,19, 17154-17172). In addition, the polyphenol hydroxyl structure in the resveratrol molecule is easily subjected to covalent modification by enzymes in the liver and intestinal tract during metabolism to form sulfate or glucoside (Ann.N.Y.Acad.Sci.2011,1215, 9-15; Ann.N.Y.Acad.Sci.2011,1215,48-59), so that the physiological function and the pharmacological action of the phenolic hydroxyl are partially counteracted.

In view of the above-mentioned shortcomings of resveratrol in practical applications, appropriate structural modification of resveratrol molecules has become an important research topic. In the derivation research of resveratrol structure, it is common to diversify the functional groups on the benzene ring in resveratrol, for example, introduce other groups (such as nitro group, halogen) on the benzene ring to replace hydroxyl (j.org.chem.2006,71, 4246-; the resveratrol of which the phenolic hydroxyl group is protected by methyl can be introduced into aldehyde group through Vilsmeier reaction, and the introduced aldehyde group is further connected with other structural fragments through Aldol condensation, so that the variety of the resveratrol derivative is widened (J.Med.chem.2013,62, 222-231); furthermore, the replacement of the 4' -hydroxyl group by other groups has also been reported in the prior art; or etherification or esterification of phenolic hydroxyl groups to improve the related activity of resveratrol. Inspired by the fact that resveratrol has wide biological activity, and the knowledge that the phenolic hydroxyl group of resveratrol has a little deficiency, people gradually depart from the research idea of exploring or modifying the phenolic hydroxyl group of resveratrol so as to obtain better biological activity, and begin to carry out various functional groups on the stilbene which is the core structure of resveratrol, so that the research on derivatization of the resveratrol structure which is based on the stilbene structure, based on various functional groups and aims at various biological activities is realized, and the research type is more common. However, the improvement on the antioxidant performance of resveratrol rarely relates to the improvement in the above modification, and especially, similar research for organically combining the structures of resveratrol and salvianolic acid A with strong antioxidant activity has not been reported yet.

In the invention, an analog E-DRS of salvianolic acid A with a resveratrol structure is designed and synthesized from RES and methyldopanic acid through esterification, acylation and aldimine condensation reaction, and the compound has good antioxidation in vitro and in vivo. Provides a new direction for the improvement of the target specificity, the side effect alleviation, the drug delivery mode improvement and the absorption and metabolism efficiency of the resveratrol in the follow-up research.

Disclosure of Invention

The technical problem to be solved by the invention is as follows: provides a salvianolic acid A analogue which has good antioxidant activity; and can be used as lead compound for various diseases.

In a first aspect of the invention, there is provided a compound of formula I:

wherein: r is selected from C₁-C₆Alkyl radical, C₁-C₆Alkenyl radical, C₁-C₆A haloalkyl group;

preferably, R is selected from C₁-C₄An alkyl group;

most preferably, the compound of formula I has the following structure (i.e., compound E-DRS):

in another aspect of the invention, there is provided a process for the preparation of a compound of formula I, the synthetic route for which is as follows:

the reaction steps are as follows:

the method comprises the following steps: prepared from resveratrol in POCl₃Performing Vilsmeier reaction in DMF to obtain aldehyde-based resveratrol (1);

step two: continuously introducing dry hydrogen chloride into the methyldopa in an alcohol solution ROH, heating and refluxing for reaction, and carrying out post-treatment to obtain methyldopate (2);

step three: dissolving the aldehyde resveratrol prepared in the step one and the methyldopate prepared in the step two in an organic solvent, refluxing and stirring for reaction, and performing post-treatment after the reaction to obtain the resveratrol methyldopa Schiff base (3)

Step four: dissolving the resveratrol methyldopa Schiff base prepared in the third step into an organic solvent, adding a reducing agent at normal temperature for reaction, and performing post-treatment after the reaction to obtain the compound shown in the formula I.

Preferably, in step one: the molar volume ratio of the resveratrol to the POCl3 is 1:0.4-0.7, preferably 1: 0.5-0.6; the reaction temperature is 0-60 ℃, and room temperature is preferred; the reaction time is 0.5 to 2 hours, preferably 1 hour.

Preferably, the reaction temperature in step two is from room temperature to reflux temperature; the reaction time is 3-8 hours.

Preferably, the molar ratio of the aldehyde-based resveratrol (1) to the methyldopate (2) in the third step is 1:1-2, preferably 1:1.4-1.8, and most preferably 1: 1.6; the organic solvent is selected from one or more of methanol, ethanol, tetrahydrofuran and DMF, and most preferably methanol.

In the fourth step, the reducing agent is selected from one or more of sodium borohydride, lithium aluminum hydride, sodium cyanoborohydride, Pd/C and Raney nickel, and sodium borohydride is most preferred.

In another aspect of the present invention, a pharmaceutical composition is provided, which comprises a compound represented by formula I or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier or excipient.

Another aspect of the invention relates to the use of a compound of formula I or a pharmaceutical composition comprising the same for the preparation of an antioxidant.

Defining:

in certain embodiments, the pharmaceutically acceptable form is a pharmaceutically acceptable salt, which is well known in the art. Examples of pharmaceutically acceptable salts are such as hydrochloric, hydrobromic, phosphoric, sulfuric, perchloric, acetic, oxalic, maleic, tartaric, citric, succinic or malonic, acetic, propionic, glycolic, pyruvic, oxalic, lactic, trifluoroacetic, methanesulfonic, ethanesulfonic, p-toluenesulfonic, salicylic, and the like.

"pharmaceutically acceptable carrier" or "pharmaceutically acceptable excipient" includes any and all solvents, dispersion media, coating agents, isotonic and absorption delaying agents and the like. Pharmaceutically acceptable carriers or excipients do not destroy the pharmacological activity of the disclosed compounds and are non-toxic when administered in a dose sufficient to deliver a therapeutic amount of the compound. The use of such media and agents for pharmaceutically active substances is well known in the art.

Compared with the prior art, the invention has the beneficial effects that:

the invention provides a novel salvianolic acid A analogue containing a resveratrol structure and having antioxidant activity, which can directly remove ROS and indirectly resist oxidation by increasing the expression of antioxidase such as CAT, SOD and the like and improving the generation of a non-enzymatic antioxidant substance GSH, broaden the range of the existing antioxidant and can be continuously optimized as a lead compound.

The compound has good in-vitro and in-vivo antioxidant activity, overcomes the defect of bioavailability of resveratrol in a living body in the prior art, and has good patent medicine prospect.

The compounds of the present invention, even at low concentrations (10. mu.M/kg-day, E-DRSL group), can protect the liver from CCl₄Induced oxidative stress damage.

Drawings

Figure 1 is compound E-DRS (R ═ CH)₂CH₃) Is/are as follows¹H NMR spectrum.

Figure 2 is compound E-DRS (R ═ CH)₂CH₃) IR spectrum of (a).

Figure 3 is compound E-DRS (R ═ CH)₂CH₃) Is/are as follows¹³C NMR spectrum.

Figure 4 is compound E-DRS (R ═ CH)₂CH₃) MS (ESI) spectrum of (A).

FIG. 5(A) shows the total antioxidant capacity of different substances detected by the FRAP method. All compounds were evaluated for antioxidant activity at the same test concentration of 0.125 mg/ml. FIGS. 5(B) and 5(C) show superoxide anion scavenging ability (B) and DPPH radical scavenging activity (C), respectively. Wherein: E-DRSD is a compound of the present application (R ═ CH)₂CH₃) RES is resveratrol, SAA is salvianolic acid A, Rutin is Rutin, and Vit C is ascorbic acid. IC (integrated circuit)₅₀(. mu.M) indicates superoxide anion scavenging ability and DPPH radical scavenging activity. Data are presented as mean ± SEM (n ═ 3) · P<0.01,***P<0.001vs.E-DRS group,#P>0.05vs.SAA group,

>0.05vs.E-DRS group。

Detailed Description

The present invention will be described in detail with reference to examples. In the present invention, the following examples are intended to better illustrate the present invention and are not intended to limit the scope of the present invention.

Resveratrol (98%), methyldopa, salvianolic acid A (99.5%), rutin, and ascorbic acid were purchased from Changsha Mike. Other chemicals were of analytical grade unless otherwise indicated.