阅读说明：本技术 新的环阿尔廷型皂苷-9,19-裂环-9,11-烯衍生物及其制备方法和应用 (New cycloartane type saponin-9, 19-seco-9, 11-ene derivative and its preparing method and use ) 是由 王芳 江志波 梁大连 张岱州 段崇刚 王帆 袁振海 李敏 贾婵媛 于 2020-07-10 设计创作，主要内容包括：本发明属于医药技术领域,涉及三个新的环阿尔廷型皂苷-9,10-裂环-9,11-烯衍生物制备方法及其应用；三个新的环阿尔廷型皂苷-9,10-裂环-9,11-烯衍生物,具有如下结构：<Image he="189" wi="700" file="DDA0002578046700000011.GIF" imgContent="drawing" imgFormat="GIF" orientation="portrait" inline="no"></Image>该三个环阿尔廷型皂苷-9,10-裂环-9,11-烯衍生物1具有抗肿瘤活性,可用于抗肿瘤药物的开发和应用。(The invention belongs to the technical field of medicines, and relates to a preparation method and application of three novel cycloartane-type saponin-9, 10-seco-9, 11-ene derivatives; three novel cycloartane-type saponin-9, 10-seco-9, 11-ene derivatives having the structure: the three cycloartane-type saponin-9, 10-seco-9, 11-ene derivative 1 has antitumor activity, and can be used for development and application of antitumor drugs.)

1. Novel cycloartane-type saponin-9, 10-seco-9, 11-ene derivatives characterized by: has the following structure:

2. a process for the preparation of a novel cycloartane-type saponin-9, 10-seco-9, 11-ene derivative as claimed in claim 1, characterized in that:

(1) performing ultrasonic and flash extraction or solvent extraction on the pteris latifolia by using 30-95% ethanol or methanol, recovering an extracting solution under reduced pressure until no alcohol smell exists, dispersing water, extracting with ethyl acetate, recovering a solvent, and collecting ethyl acetate and a water phase extract;

(2) subjecting the water phase extract to nonpolar macroporous resin column chromatography, and performing gradient elution with water and alcohol in sequence to obtain 15-95% alcohol elution parts;

(3) separating the alcohol-washed substance obtained in the step (2) by silica gel column chromatography, reversed phase medium pressure column chromatography or gel column chromatography, and gradient eluting with dichloromethane, chloroform, acetone, methanol, ethyl acetate, ethanol, water or mixed solvent;

(4) and (3) carrying out semi-preparative or preparative HPLC chromatographic separation on the obtained flow component in the step (3), and eluting by using methanol/water, acetonitrile/water or methanol/acetonitrile/water as a mobile phase to obtain the new cycloartane-type saponin-9, 10-seco-9, 11-ene derivative 1-3.

3. A process for preparing a novel cycloartane-type saponin-9, 10-seco-9, 11-ene derivative according to claim 2, characterized in that: the plant source is aerial part of Asplenium ruprechiti Sa.Kurata of family Achilleae.

4. A process for preparing a novel cycloartane-type saponin-9, 10-seco-9, 11-ene derivative according to claim 2, characterized in that: the extraction method in the step (1) is heating reflux extraction or ultrasonic extraction for 1-3 times, the used solvent is 30-95% ethanol, and the ratio of the mass of the camptosorus sibiricus to the volume of the solvent is 1: 5-1: 20; the extraction is carried out for 1-5 times, and the volume ratio of the aqueous phase extract to the solvent is 1: 3-1: 10.

5. A process for preparing a novel cycloartane-type saponin-9, 10-seco-9, 11-ene derivative according to claim 2, characterized in that: the alcohol in the step (2) is ethanol, and the gradient elution concentration of the alcohol is 15-95%; the nonpolar resin is AB-8 macroporous resin and MCI resin, 30-60% of ethanol elution part is separated by AB-8 macroporous resin column chromatography, and 50-80% of ethanol elution part contains the target compound by MCI resin column chromatography.

6. A process for preparing a novel cycloartane-type saponin-9, 10-seco-9, 11-ene derivative according to claim 2, characterized in that: separating by silica gel column chromatography in the step (3), wherein an elution solvent is chloroform or dichloromethane/methanol or ethanol, ethyl acetate/methanol or 95% ethanol with the ratio of 1: 12-3: 7, and the fraction with the mixing ratio of 4: 1-3: 1 contains the target compound; separating by reversed-phase medium-pressure column chromatography, wherein an elution solvent is a mixed solvent of ethanol-water or methanol-water, the volume ratio of the mixed solvent is 0: 100-100: 0, preferably 50: 50-100: 0, and the fraction with the mixing ratio of 60: 40-90: 10 contains the target compound; and (3) separating by gel column chromatography, wherein an elution solvent is methanol, a dichloromethane/methanol mixed solvent is eluted isocratically, and the fraction with the dichloromethane/methanol volume ratio of 1: 1-1: 5 contains the target compound.

7. The process for producing a novel cycloartane-type saponin-9, 10-seco-9, 11-ene derivative according to claim 2, characterized in that: and (4) separating the elution flow fraction in the step (4) by using high performance liquid chromatography, wherein the mobile phase is methanol/water, acetonitrile/water or methanol/acetonitrile/water, and the compound 1-3 is obtained from the flow fraction with the methanol/water mixed solvent ratio of 5: 1-1: 1.

8. The use of a novel cycloartane-type saponin-9, 10-seco-9, 11-ene derivative 1-3 as claimed in claim 1 in the preparation of an anti-tumor medicament.

Technical Field

The invention belongs to the technical field of medicines, relates to separation, preparation and application of saponin compounds, and particularly relates to three novel cycloartane-type triterpenoid saponin derivatives, a preparation method and application thereof in antitumor drugs.

Background

The cycloartane (9,19-cycloartane) compound is a lanostane type tetracyclic triterpene compound. In recent decades, a large number of ring-altenin classes have been discovered. The isolated cycloartane compounds are mainly from the plant kingdom, such as the genera Astragalus (Astragalus) of the Leguminosae family, Cimicifuga (Cimicifuceae) and Thalictrum (Thalictrum) of the Ranunculaceae family, and Ceratopteris (Asplenium) of the Ceratopteridae family.

The cycloartane-type saponin compound is the main component of cimicifuga rhizome and astragalus root. Recent studies have shown that they have a wide variety of pharmacological activities. The document reports that Astragalus oleifolius of the genus Astragalus has the effects of arresting sweating and promoting urination, and has curative effects on nephritis, diabetes, leukemia and uterine cancer, and the main component of the Astragalus oleifolius comprises cycloartane triterpenes; astragalus spinous is used in Egyptian folk to treat hypertension, leukemia and uterine cancer, and the alcohol extract of Astragalus verrucosus has immunity enhancing and regulating effects, and contains cycloartane triterpene as main component.

Although the literature reports that 12 cyclic-altin-type saponins isolated from the 70% ethanol extract of the whole plant of pteris latifolia collected in the northeast region have no cytotoxic activity. In earlier researches, the large-polarity extraction part of the whole herb of the pteris latifolia which is collected from the south of Henan is found to be rich in novel cyclic-argentine-type saponin, the large-polarity extraction part of the whole herb of the pteris latifolia is continuously separated and prepared from the large-polarity extraction part of the whole herb of the pteris latifolia by comprehensively utilizing various separation and identification means and focusing on the acquisition of trace components, 3 cyclic-argentine-type saponin-9, 10-split-ring-9, 11-alkene derivatives with novel structures are identified and the antitumor activity is determined, and reports are not found in the prior art.

Disclosure of Invention

The invention aims to provide three cyclic-Altin type saponin-9, 19-split-ring-9, 11-alkene derivatives with new structures, a preparation method and application thereof.

The invention provides three novel cyclic-argentine-type saponins-9, 19-seco-9, 11-ene derivatives having the structure:

the invention also provides a preparation method of the novel cycloartane saponin-9, 19-seco-9, 11-ene derivative, which comprises the following steps:

(1) performing ultrasonic and flash extraction or solvent extraction on the pteris latifolia by using 30-95% ethanol or methanol, recovering an extracting solution under reduced pressure until no alcohol smell exists, dispersing water, extracting with ethyl acetate, recovering a solvent, and collecting ethyl acetate and a water phase extract;

(2) subjecting the water phase extract obtained in the step (1) to nonpolar macroporous resin column chromatography, and performing gradient elution with water and alcohol in sequence to obtain 15-95% alcohol elution parts;

(3) separating the alcohol-washed substance obtained in the step (2) by silica gel column chromatography, reversed phase medium pressure column chromatography or gel column chromatography, and gradient eluting with dichloromethane, chloroform, acetone, methanol, ethyl acetate, ethanol, water or mixed solvent;

(4) and (3) carrying out semi-preparative or preparative HPLC chromatographic separation on the obtained flow component in the step (3), and eluting by using methanol/water, acetonitrile/water or methanol/acetonitrile/water as a mobile phase to obtain the new cycloartane-type saponin-9, 19-seco-9, 11-ene derivative 1-3.

The invention provides a preparation method of a novel cycloartane saponin-9, 19-seco-9, 11-ene derivative 1-3, the extraction method in step (1) is heating reflux extraction or ultrasonic extraction for 1-3 times, the used solvent is ethanol with the volume of 30-95%, and the ratio of the mass of the camptosorus sibiricus to the volume of the solvent is 1: 5-1: 20; the extraction is carried out for 1-5 times, and the volume ratio of the aqueous phase extract to the solvent is 1: 3-1: 10.

The invention provides a preparation method of the novel cycloartane saponin-9, 19-seco-9, 11-ene derivative 1-3, wherein the alcohol in the step (2) is ethanol, and the gradient elution concentration of the alcohol is 15-95 percent; the nonpolar resin is AB-8 macroporous resin and MCI resin, 30-60% of ethanol elution part is separated by AB-8 macroporous resin column chromatography, and 50-80% of ethanol elution part contains the target compound by MCI resin column chromatography.

The novel cycloartane-type saponin-9, 19-split ring-9, 11-alkene derivative 1-3 is prepared by a method comprising the following steps of (3) separating by silica gel column chromatography, wherein an elution solvent is chloroform or dichloromethane/methanol or ethanol, ethyl acetate/methanol or 95% ethanol with the ratio of 1: 12-3: 7, and a fraction with the mixing ratio of 4: 1-3: 1 contains a target compound; separating by reversed-phase medium-pressure column chromatography, wherein an elution solvent is a mixed solvent of ethanol-water or methanol-water, the volume ratio of the mixed solvent is 0: 100-100: 0, preferably 50: 50-100: 0, and the fraction with the mixing ratio of 60: 40-90: 10 contains the target compound; and (3) separating by gel column chromatography, wherein an elution solvent is methanol, a dichloromethane/methanol mixed solvent is eluted isocratically, and the fraction with the dichloromethane/methanol volume ratio of 1: 1-1: 5 contains the target compound.

According to the preparation method of the novel cycloartane-type saponin-9, 19-seco-9, 11-ene derivative 1-3, the eluted fraction in the step (4) is separated by high performance liquid chromatography, the mobile phase is methanol/water, acetonitrile/water or methanol/acetonitrile/water, and the compound 1-3 is obtained from the fraction with the methanol/water mixed solvent ratio of 5: 1-1: 1.

The invention carries out preliminary test and evaluation on the in vitro anti-tumor activity of the novel compound 1-3, and the selected cell lines are leukemia cells (HL-60) and human liver cancer cells (HepG 2). Therefore, the novel compounds 1-3 prepared by the invention can be applied to the development of antitumor drugs.

The invention provides a method for enriching, preparing and identifying new cycloartane saponin-9, 19-split ring-9, 11-alkene derivatives 1-3 by taking the whole grass of pteris latifolia as a raw material for the first time, and the invention evaluates the anti-tumor activity and explains the application of the derivatives in developing anti-tumor drugs.

Detailed Description

The following examples further illustrate the invention but are not intended to limit the invention thereto.