CN113121353B - Ferulic acid dimer with antioxidant activity and preparation method and application thereof - Google Patents

Abstract

The present invention provides a ferulic acid dimer with antioxidant activity and a preparation method and application thereof. Ferulic acid dimer compound of the present invention 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-dihydro The structural formula of naphthalene-2-carboxylic acid is shown in formula (I), which is obtained from the dried or fresh product of the fruit, stem or leaf of rice. It was confirmed by in vitro pharmacological experiments that the compound 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-dihydronaphthalene-2 ‑Carboxylic acid has good superoxide radical scavenging ability, and its PSC value is 1.47±0.57 μmol TE/μmol, indicating that it has good antioxidant activity. Described ferulic acid dimer compound 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-dihydro Naphthalene-2-carboxylic acid is expected to be used as a lead compound to develop new antioxidant drugs.

Description

A kind of ferulic acid dimer with antioxidant activity and its preparation method and application

technical field

The invention belongs to the technical field of natural medicines, in particular to a new ferulic acid dimer compound 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methyl) Oxycarbonyl)-3,4-dihydronaphthalene-2-carboxylic acid and its preparation method and application in the preparation of antioxidant medicines.

Background technique

Studies have shown that cancer, aging, neurodegenerative diseases and cardiovascular diseases are inextricably linked with the production of excess free radicals (highly reactive molecules). Continued accumulation of reactive oxygen species (ROS) and reactive nitrogen species (RNS) in the body initiates oxidative stress (a long-term imbalance between the production of highly reactive molecules and their anti-oxidative effects leading to tissue damage) , thereby impairing the defense system of the organism. These active molecules can directly oxidize and damage DNA, proteins, lipids, activate a variety of intracellular signaling pathways related to insulin resistance and impaired β-cell function, and even interfere with physiological reduction signaling to affect cell proliferation and differentiation. Antioxidants are a class of substances that slow down or inhibit oxidation, which can inhibit the formation of free radicals, scavenge free radicals or repair damaged biomolecules, thereby effectively preventing and alleviating the damage caused by oxidative stress to the body. Phenolic substances with polyphenolic hydroxyl structures, such as flavonoids, phenolic acids, and proanthocyanidins, exhibit good antioxidant activity and have become an important source of new antioxidants.

Rice (Citrusgrandis L.Osbeck) is a genus of Poaceae, and its fruit is the staple food of nearly half of the world's population. It has been reported that rice is rich in phenolics, and a considerable part of phenolics exists in the form of binding. , these phenolic substances are expected to become new natural antioxidants. 6-Hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-diol with antioxidant activity in rice has not been found so far. Research report on the composition of hydronaphthalene-2-carboxylic acid.

SUMMARY OF THE INVENTION

The purpose of the present invention is to provide a new ferulic acid dimer component 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl) -3,4-dihydronaphthalene-2-carboxylic acid, and a preparation method thereof, and application of the same in the preparation of medicines with antioxidative effect.

The novel ferulic acid dimer component of the present invention is 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4 -Dihydronaphthalene-2-carboxylic acid, or a pharmaceutically acceptable salt thereof, or an esterified derivative thereof, the structure of which is represented by formula (I):

The novel ferulic acid dimer component of the present invention is 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4 -Dihydronaphthalene-2-carboxylic acid was isolated and obtained by the inventors for the first time from dried or fresh products of various tissue parts of rice (Oryza sativa L.), and the specific parts can be brown rice, bran layer and polished rice layer.

The novel ferulic acid dimer component provided by the present invention is 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4- Dihydronaphthalene-2-carboxylic acid was confirmed by in vitro pharmacological experiments to have good superoxide radical (PSC) scavenging ability, and its PSC value was 1.47±0.57μmol TE/μmol. The new ferulic acid dimer component 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-di Hydronaphthalene-2-carboxylic acid, or a pharmaceutically acceptable salt thereof, or an esterified derivative thereof can be used to prepare antioxidant drugs.

The novel ferulic acid dimer component of the present invention 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-di Hydronaphthalene-2-carboxylic acid, or its pharmaceutically acceptable salt, or its esterified derivative can be combined with a preparation or a pharmaceutically acceptable excipient or carrier to prepare a drug with antioxidant activity that can be used to relieve oxidative stress or pharmaceutical composition. The medicine or pharmaceutical composition can be in wettable powder, tablet, granule, capsule, oral liquid, drop pill, injection, aerosol and other dosage forms; it can also adopt controlled-release or sustained-release dosage forms known in the modern pharmaceutical industry or Nanoformulations.

Therefore, the present invention also provides an antioxidant drug, which comprises the above-mentioned new ferulic acid dimer component 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy- 3-(methoxycarbonyl)-3,4-dihydronaphthalene-2-carboxylic acid, or a pharmaceutically acceptable salt thereof, or an esterified derivative thereof, as an active ingredient.

The antioxidant drug also includes formulations or pharmaceutically acceptable excipients or carriers, such as wettable powder, tablet, granule, capsule, oral liquid, drop pill, injection, aerosol and other dosage forms; The controlled-release or sustained-release dosage forms or nano-formulations well known in the modern pharmaceutical industry are used.

The present invention also provides a new ferulic acid dimer component 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4 -The preparation method of dihydronaphthalene-2-carboxylic acid, which is obtained from rice (Oryza sativa L.) plants, including dried or fresh products of fruit, stem or leaf of rice. Preferably, it is obtained from dry or fresh products of various tissue parts of rice fruit, and the specific parts can be brown rice, bran layer and/or polished rice.

The novel ferulic acid dimer component of the present invention 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-di The preparation method of hydronaphthalene-2-carboxylic acid, the specific steps are preferably as follows: the rice is pulverized and then leached with an ethanol aqueous solution, the extract is separated and the residue is collected, the extraction residue is added to a NaOH solution and extracted under a nitrogen environment, and then the pH is adjusted to 1, and the Extract with ethyl acetate, collect the ethyl acetate phase and concentrate under reduced pressure to remove the organic solvent to obtain the extract. The extract is subjected to normal phase silica gel column chromatography, eluted with a gradient of chloroform/methanol from 100:0 to 60:40 by volume, and collected. Component F2 eluted with a volume ratio of chloroform/methanol of 90:10. Component F2 was chromatographed on an ODS-A medium pressure column, eluted with a gradient of methanol/water alcohol from a volume ratio of 30:70 to 90:10, and collected methanol/water. Fraction F2-8 eluted with a water volume ratio of 70:30, fraction F2-8 was purified by high performance liquid phase separation to obtain 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methyl Oxy-3-(methoxycarbonyl)-3,4-dihydronaphthalene-2-carboxylic acid.

The invention adopts the extraction and separation of high-efficiency antioxidant (drug)-ferulic acid dimer compound 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy- 3-(methoxycarbonyl)-3,4-dihydronaphthalene-2-carboxylic acid has controllable preparation process conditions, convenient extraction, environmental friendliness and potential good economic benefits, and the obtained new dihydroferulic acid Polymer Compound 6-Hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-dihydronaphthalene-2-carboxylic acid Properties Stable and easy to store. It has good antioxidant activity and is expected to be used as a lead compound to develop new antioxidant drugs.

Description of drawings

Figure 1 is the compound 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-dihydronaphthalene-2-carboxylic acid ¹ H NMR spectrum of ;

Figure 2 is the compound 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-dihydronaphthalene-2-carboxylic acid The ¹³ C NMR spectrum of ;

Figure 3 is the compound 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-dihydronaphthalene-2-carboxylic acid ¹ H ^-1 H COSY spectrum of ;

Figure 4 is the compound 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-dihydronaphthalene-2-carboxylic acid The HSQC spectrum of ;

Figure 5 is the compound 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-dihydronaphthalene-2-carboxylic acid HMBC spectrum of ;

Figure 6 is the compound 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-dihydronaphthalene-2-carboxylic acid HR-EI-MS spectrum.

Detailed ways

The following examples are further descriptions of the present invention, not limitations of the present invention. Simple improvements to the present invention according to the essence of the present invention all belong to the scope of protection of the present invention.

Example 1: Ferulic acid dimer compound 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-di Preparation of Hydronaphthalene-2-Carboxylic Acid

1.1 Plant origin and identification

Fruit samples of rice (Oryza sativa L.), a plant material for extraction, were purchased from Harbin, Heilongjiang Province.

1.2 Extraction and separation

The rice fruit (rice) was crushed and extracted with 95% ethanol aqueous solution for 48 hours. The extract was separated and the residue was collected. The extraction residue was added with 2M NaOH solution (1:20, w/v g/ml) and extracted under nitrogen atmosphere. After 4 hours, the pH was adjusted to 1 with 6M HCl, extracted with ethyl acetate (1:1, v/v) for 5 times, and the ethyl acetate phase was collected and concentrated under reduced pressure to remove the organic solvent to obtain an extract. The extract was subjected to normal-phase silica gel column chromatography, eluted with a gradient of chloroform/methanol from 100:0 to 60:40 in a volume ratio, and the fraction F2 eluted with a chloroform/methanol volume ratio of 90:10 was collected. A medium pressure column chromatography, gradient elution with methanol/water alcohol from 30:70 to 90:10 by volume, fraction F2-8 and subfraction F2-8 eluted with methanol/water volume ratio of 70:30 were collected Using 45% methanol aqueous solution (v/v) as the mobile phase and 6 mL/min as the flow rate, the compound 6- Hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-dihydronaphthalene-2-carboxylic acid (3 mg, tR 128 min) .

1.3 Structural identification of new ferulic acid dimer compounds

Figure 1 is the compound 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-dihydronaphthalene-2-carboxylic acid ¹ H NMR spectrum of ; Figure 2 is the compound 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-dihydro ¹³ C NMR spectrum of naphthalene-2-carboxylic acid; Figure 3 is the compound 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)- ¹ H- ¹ H COSY spectrum of 3,4-dihydronaphthalene-2-carboxylic acid; Figure 4 is the compound 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy The HSQC spectrum of -3-(methoxycarbonyl)-3,4-dihydronaphthalene-2-carboxylic acid; Figure 5 is the compound 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7 - HMBC spectrum of -methoxy-3-(methoxycarbonyl)-3,4-dihydronaphthalene-2-carboxylic acid; Figure 6 is the compound 6-hydroxy-4-(4-hydroxy-3-methoxybenzene yl)-7-methoxy-3-(methoxycarbonyl)-3,4-dihydronaphthalene-2-carboxylic acid HR-EI-MS spectrum; 6-hydroxy-4-(4-hydroxy-3- Methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-dihydronaphthalene-2-carboxylic acid, pale yellow oil; UV(MeOH)λmax nm(logε)225( 4.34), 289(3.89), 326(3.87); HRESIMS m/z 401.1250[M+H] ⁺ (calcd for C21H21O8 ⁺ , 401.1231, error-1.9mDa) and 423.1070[M+Na] ⁺ (calcd for C21H20NaO8+ 423.1050, error-2.0mDa); ¹ H (500MHz) and ¹³ C (125MHz) NMR data are shown in Table 1 below.

Table 1. Compound 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-dihydronaphthalene-2-carboxylic acid NMR data (in CD ₃ OD)

According to the comprehensive analysis of the above UV, mass spectrometry and one-dimensional and two-dimensional nuclear magnetic resonance data, the structural formula of the compound is analytically deduced as shown in formula (I), named 6-hydroxy-4-(4-hydroxy-3-methoxy phenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-dihydronaphthalene-2-carboxylic acid:

Example 2: Ferulic acid dimer compound 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-di Antioxidant activity detection of hydronaphthalene-2-carboxylic acid

1. Instruments and materials

Dimethyl sulfoxide was produced by Cambridge Company, USA; Trolox reagent, fluorescein, 2,2'-azo(2-methylpropionamidine) hydrochloride (AAPH) were purchased from Sigma Company, USA; microplate reader was Genois microplate reader (TecanGENios, Swizerland).

2. Evaluation of PSC activity

The PSC antioxidant capacity of the compounds was determined by fluorescence microplate reader assay. Trolox standard and various samples to be tested were dissolved and diluted to the appropriate concentration with 75mM phosphate buffer (PH=7.4), 80μL of DCFH-DA with a concentration of 2.48mM was added to 900μL of 1.0mM KOH and hydrolyzed for 5 minutes to remove diacetate ( DA) part to obtain an activated DCFH solution. 100 μL of pure compound or extract (appropriately diluted in 75 mM phosphate buffer (pH 7.4)) was transferred to a 96-well plate and 100 μL of DCFH was added. The 96-well plate was loaded into a spectrofluorometer plate holder and the reaction was started by mixing the solution in each well by shaking at 1200 rpm for 20 s, followed by an additional 50 μL of 200 mM ABAP. The reaction was carried out at 37°C, and the fluorescence was monitored with a spectrofluorometer at 485 nm excitation and 538 nm emission, every 2 min for 40 min. Trolox (vitamin E) served as a positive control. Each experiment was replicated three times, and the PSC values of the test samples were calculated according to the Trolox standard curve, and the results were expressed in Trolox equivalents (μmol TE/μmol).

PSC(unit)=1-(SA/CA)

where SA is the AUC of the sample or standard and CA is the AUC of the buffer only control reaction.

3. The experimental data are shown in Table 2:

Table 2. Compound 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-dihydronaphthalene-2-carboxylic acid antioxidant activity

4. Experimental conclusion:

This experiment shows that the ferulic acid dimer compound 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-di Hydronaphthalene-2-carboxylic acid has good antioxidant activity, and it is expected to be developed for the preparation of antioxidant drugs, with wide application potential and promising prospects.

The above are only the preferred embodiments of the present invention. It should be noted that the above preferred embodiments should not be regarded as limitations of the present invention, and the protection scope of the present invention should be based on the scope defined by the claims. For those skilled in the art, without departing from the spirit and scope of the present invention, several improvements and modifications can also be made, and these improvements and modifications should also be regarded as the protection scope of the present invention.

Claims (4)

1. A ferulic acid dimer compound 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-di The preparation method of hydronaphthalene-2-carboxylic acid is characterized in that, the concrete steps are: a. Preparation of total extract: pulverize the rice and then extract with ethanol aqueous solution, separate the extract to collect the residue, add NaOH solution to the extraction residue and extract under nitrogen atmosphere, then adjust the pH to 1, extract with ethyl acetate, and collect the ethyl acetate The extract is obtained by concentrating under reduced pressure to remove the organic solvent; b. Separation and purification: The extract was subjected to normal phase silica gel column chromatography, eluted with a gradient of chloroform/methanol from a volume ratio of 100:0 to 60:40, and the fraction F2 eluted with a volume ratio of chloroform/methanol of 90:10 was collected. Fraction F2 adopts ODS-A medium pressure column chromatography, eluted with methanol/water gradient from volume ratio 30:70 to 90:10, and collects fraction F2-8 eluted with methanol/water volume ratio of 70:30. F2-8 was purified by HPLC to obtain 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-dihydro Naphthalene-2-carboxylic acid; The ferulic acid dimer 6-hydroxy-4-(4-hydroxy-3-methoxyphenyl)-7-methoxy-3-(methoxycarbonyl)-3,4-dihydronaphthalene The structure of -2-carboxylic acid is shown in formula (I):

Formula (I). 2. preparation method according to claim 1 is characterized in that, described ethanol aqueous solution is the ethanol aqueous solution of volume fraction 95%. 3. The preparation method according to claim 1 or 2, wherein the rice comprises dried or fresh products of fruit, stem or leaf of rice. 4. The preparation method according to claim 3, wherein the rice is rice fruit, comprising bran layer and/or polished rice.

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