阅读说明：本技术 一种化合物及其制备方法和应用 (Compound and preparation method and application thereof ) 是由 肖伟 倪付勇 温建辉 谢雪 李明 吴云 王振中 于 2018-08-24 设计创作，主要内容包括：本发明涉及一种植物源化合物及其制备方法和应用。本发明对川芎进行研究,得到了一种能得到新的活性组分,其结构式如下,经实验表明,其对LPS刺激A7r5细胞产生NO有明显的抑制作用,该化合物具有良好的抗炎作用；该化合物对经典途径的补体激活有明显的抑制作用,具有良好的抗补体活性。<Image he="253" wi="700" file="DDA0001776799940000011.GIF" imgContent="drawing" imgFormat="GIF" orientation="portrait" inline="no"></Image>(The invention relates to a plant source compound and a preparation method and application thereof. The invention researches the ligusticum wallichii to obtain a new active component with the structural formula shown in the specification, and experiments show that the compound has an obvious inhibition effect on the stimulation of LPS to A7r5 cells to generate NO, and has a good anti-inflammatory effect; the compound has obvious inhibition effect on the activation of the complement in the classical pathway and has good anti-complement activity.)

1. A compound having the structure of formula I:

2. a process for the preparation of a compound according to claim 1, comprising the steps of:

step (1): reflux-extracting rhizoma Ligustici Chuanxiong with ethanol water solution, and concentrating the extractive solution to obtain extract;

step (2): mixing the extract obtained in the step (1) with water, sequentially extracting with petroleum ether, ethyl acetate and n-butanol, and concentrating the ethyl acetate extraction part to obtain an ethyl acetate extract;

and (3): performing column chromatography separation on the ethyl acetate extract obtained in the step (2), performing gradient elution by using a dichloromethane-methanol mixed solvent, wherein the dichloromethane-methanol volume ratio of the gradient elution is (50-1): 1, and collecting a component obtained by separation with the dichloromethane-methanol volume ratio of (15-1): 1;

and (4): purifying the component parts obtained in the step (3) by gel column chromatography, eluting by an organic solvent, collecting and separating;

and (5): and (4) carrying out chromatographic separation and purification on the components separated in the step (4), eluting by using a 15-45% organic solvent water solution, and treating the eluent to obtain the compound shown in the formula (I).

3. The preparation method according to claim 2, wherein in the step (1), the weight ratio of the ligusticum wallichii to the ethanol aqueous solution is 1: (10-20).

4. The method of claim 2, wherein the gel used in the gel column chromatography is selected from the group consisting of Sephadex LH-20, Sephadex G15 and Sephadex G50.

5. The method according to claim 2, wherein in the step (4), the organic solvent is selected from methanol, ethanol, 50% methanol-water or 50% ethanol-water; the chromatographic separation and purification in the step (5) is selected from high performance liquid chromatography, medium and low pressure preparative chromatography or normal pressure column chromatographic separation.

6. Use of a compound of claim 1 for the preparation of an anti-inflammatory or anti-complement drug.

7. Use according to claim 6, the anti-inflammatory agent being selected from agents directed against inflammation induced or triggered by LPS.

8. The use according to claim 6, wherein the anticomplementary drug is selected from the group consisting of drugs that inhibit the complement that hemolyzes hemolysin-sensitized sheep red blood cells.

9. An anti-inflammatory drug or an anti-complement drug comprising the compound of claim 1.

10. An anti-inflammatory agent or an anti-complement agent according to claim 9, wherein the anti-inflammatory agent is selected from an oral administration form, an injection administration form, or an external administration form.

Technical Field

The invention relates to the technical field of medicines, and particularly relates to a plant source compound and a preparation method and application thereof.

Background

The traditional Chinese medicine rhizoma Ligustici Chuanxiong (Ligusticum chuanxiong Hort) is dried rhizome of Umbelliferae rhizoma Ligustici Chuanxiong, and is mainly produced in Yunnan, Guizhou, Sichuan and other places, namely fructus Syzygii Aromatici, herba Pyrolae, rhizoma Ligustici Chuanxiong, and rhizoma Ligustici Chuanxiong.

Chuan Xiong has the actions of activating blood and moving qi, dispelling wind and alleviating pain, which are recorded in Shen nong Ben Cao Jing (Shen nong's herbal). It is mainly used for irregular menstruation, dysmenorrhea, amenorrhea, headache, arthralgia due to wind-cold-dampness, etc. caused by blood stasis and qi stagnation.

The rhizoma Ligustici Chuanxiong has complicated chemical components and wide bioactivity, and comprises volatile oil, alkaloids, polysaccharide, etc. The rhizoma Ligustici Chuanxiong volatile oil mainly comprises peptides, terpenes, organic acids and esters thereof. The alkaloid components in rhizoma Ligustici Chuanxiong mainly include tetramethylrazine, rye alkali, trimethylamine, adenine, adenosine, choline, uracil, etc., wherein ligustrazine is the main effective component.

Disclosure of Invention

The technical problem to be solved by the invention is to provide a compound, a preparation method and application thereof, wherein the prepared compound has good anti-inflammatory activity or anti-complement activity.

Specifically, the invention relates to a compound having the structure of formula I:

the invention also provides a preparation method of the compound, which is characterized by comprising the following steps:

step (1): reflux-extracting rhizoma Ligustici Chuanxiong with ethanol water solution, and concentrating the extractive solution to obtain extract; the ethanol aqueous solution is preferably 60-80% ethanol aqueous solution, more preferably 75% ethanol aqueous solution,

step (2): mixing the extract obtained in the step (1) with water, sequentially extracting with petroleum ether, ethyl acetate and n-butanol, and concentrating the ethyl acetate extraction part to obtain an ethyl acetate extract;

and (3): performing column chromatography separation on the ethyl acetate extract obtained in the step (2), performing gradient elution by using a dichloromethane-methanol mixed solvent, wherein the dichloromethane-methanol volume ratio of the gradient elution is (50-1): 1, and collecting a component obtained by separation with the dichloromethane-methanol volume ratio of (10-6): 1;

and (4): purifying the component parts obtained in the step (3) by gel column chromatography, eluting by an organic solvent, collecting and separating;

and (5): and (4) carrying out chromatographic separation and purification on the components separated in the step (4), eluting by using a 15-45% organic solvent water solution, and treating the eluent to obtain the compound shown in the formula (I).

Further, in the step (1), the weight ratio of the ligusticum wallichii to the ethanol water solution is 1: (10-20).

Specifically, the gel used in the gel column chromatography is selected from Sephadex LH-20, Sephadex G15 or Sephadex G50.

Further, in the step (4), the organic solvent is selected from methanol, ethanol, 50% methanol-water or 50% ethanol-water.

Specifically, the chromatographic separation and purification in the step (5) is selected from high performance liquid chromatography, medium and low pressure preparative chromatography or normal pressure column chromatographic separation. The solvent in step (5) may be selected from methanol, ethanol, acetonitrile, etc.

The invention also provides application of the compound shown in the formula I in preparing anti-inflammatory drugs. The inflammation may be caused by burns, chemical irritation, physical injury, toxins, and the like. The anti-inflammatory agent is preferably directed against inflammation induced or caused by LPS (lipopolysaccharide).

The invention also provides application of the compound shown in the formula I in preparation of an anticomplement medicament. This complement over-activation is caused by foreign invaders, the removal of damaged or dead cells and tissues in the body. The anticomplementary drug is preferably a complement that inhibits hemolysis induced by sensitized 2% sheep red blood cells.

The invention also provides an anti-inflammatory drug or an anti-complement drug, which comprises the compound shown in the formula I.

Specifically, the anti-inflammatory drug or the anti-complement drug may be selected from an oral administration dosage form, an injection administration dosage form, or an external administration preparation.

The invention researches the ligusticum wallichii to obtain a new active component which has obvious consistent effect on stimulating A7r5 cells to generate NO by LPS and has good anti-inflammatory effect. In addition, the compound is found to have obvious inhibition effect on complement activation by measuring the in vitro classical anticomplementary activity of the compound, the effect is stronger than that of a positive drug heparin, and the compound has good anticomplementary activity.

Drawings

FIG. 1 is an ESI-MS diagram of a compound prepared according to example 1 of the present invention;

FIG. 2 is a diagram of the compound prepared in example 1 of the present invention¹H NMR chart;

FIG. 3 is a photograph of a compound prepared in example 1 of the present invention¹³C NMR chart;

FIG. 4 is a HSQC plot of the compound prepared in example 1 of the present invention;

FIG. 5 is a diagram of HMBC for the compound prepared in example 1 of the present invention;

FIG. 6 is a photograph of a compound prepared in example 1 of the present invention¹H-¹H COSY picture.

Detailed Description

The following detailed description of the present invention, taken in conjunction with the accompanying drawings and examples, is provided to enable the invention and its various aspects and advantages to be better understood. However, the specific embodiments and examples described below are for illustrative purposes only and are not limiting of the invention.

It is specifically noted that similar alternatives and modifications will be apparent to those skilled in the art, which are also intended to be included within the present invention. It will be apparent to those skilled in the art that the techniques of the present invention may be implemented and applied by modifying or appropriately combining the methods and applications described herein without departing from the spirit, scope, and content of the present invention.

Secondly, it is to be noted that the concentrations referred to in the present invention are in volume percent (v/v). All percentages, ratios, proportions, or parts are by weight unless otherwise specified. In addition, if the specific conditions are not indicated, the invention is carried out according to the conventional conditions or the conditions suggested by the manufacturer, and the used raw material drugs or auxiliary materials and the used reagents or instruments are the conventional products which can be obtained commercially.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. In addition, any methods and materials similar or equivalent to those described herein can be used in the practice of the present invention. The invention takes the chuanxiong rhizome as the raw material, and the chuanxiong rhizome can be the chuanxiong rhizome known by the technicians in the field, and the invention has no special limitation on the above.

Preparation of Compounds of formula (I)

1. Medicinal materials and reagents

The reagents were analytically pure except for the chromatographic purities of methanol and acetonitrile used in the liquid chromatography. The rhizoma Ligustici Chuanxiong is purchased from Bozhou, Anhui.

2. Example 1

10kg of ligusticum wallichii medicinal material is crushed, then 10 times of 75% ethanol is added for reflux extraction for 2 times, each time lasts for 2 hours, filtration is carried out, and ethanol is recovered from filtrate and concentrated to obtain extract. Adding water to the extract for suspension, sequentially extracting with petroleum ether, ethyl acetate and n-butanol, taking an ethyl acetate extraction part, recovering ethyl acetate under reduced pressure, concentrating to obtain an ethyl acetate extract, performing silica gel column chromatography on the ethyl acetate extract, performing gradient elution with dichloromethane-methanol (50:1, 20:1, 10:1, 8:1, 6:1, 3:1 and 1:1) every 500ml, receiving one fluid fraction, collecting a dichloromethane-methanol (10:1) component, performing reduced pressure concentration, purifying with a Sephadex LH-20 gel column, performing methanol elution with one fluid fraction every 5ml, performing reduced pressure concentration, performing high performance liquid chromatography separation, and eluting with 20% acetonitrile water solution to obtain 7mg of the compound shown in the formula (I).

3. Example 2

15kg of ligusticum wallichii medicinal material is crushed, then is added with 75% ethanol with the amount of 15 times of the weight of the medicinal material for reflux extraction for 3 times, each time lasts for 1.5h, the medicinal material is filtered, and the ethanol is recovered from the filtrate and concentrated to obtain an extract. Adding water to the extract for suspension, sequentially extracting with petroleum ether, ethyl acetate and n-butanol, taking the ethyl acetate extraction part, recovering ethyl acetate under reduced pressure, concentrating to obtain an ethyl acetate extract, performing silica gel column chromatography on the ethyl acetate extract, performing gradient elution with dichloromethane-methanol (50:1, 20:1, 10:1, 8:1, 6:1, 3:1 and 1:1) every 500ml, receiving one fluid fraction, collecting the dichloromethane-methanol (8:1) component, performing reduced pressure concentration, purifying with a Sephadex G15 gel column, performing ethanol elution, collecting one fluid fraction every 8ml, performing reduced pressure concentration, performing high performance liquid chromatography separation, and eluting with 20% acetonitrile water solution to obtain 6mg of the compound shown in the formula (I).

4. Example 3

5kg of ligusticum wallichii medicinal material is crushed, then 20 times of 75% ethanol is added for reflux extraction for 2 times, each time lasts for 2 hours, filtration is carried out, and the ethanol is recovered from the filtrate and concentrated to obtain an extract. Suspending the extract in water, sequentially extracting with petroleum ether, ethyl acetate and n-butanol, taking the ethyl acetate extraction part, recovering ethyl acetate under reduced pressure, concentrating to obtain ethyl acetate extract, subjecting the ethyl acetate extract to silica gel column chromatography, gradient eluting with dichloromethane-methanol (50:1, 20:1, 10:1, 8:1, 6:1, 3:1 and 1:1) with one fraction per 500ml, collecting dichloromethane-methanol (6:1) component, concentrating under reduced pressure, purifying with Sephadex LH-20 gel column, eluting with 50% methanol-water, collecting one fraction per 5ml, concentrating under reduced pressure, separating with normal pressure column chromatography, eluting with 30% methanol water solution to obtain 8mg of the compound shown in formula I.

Structural identification of compounds