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CN-121991013-A - Method for extracting rehmannia pigment compound from rehmannia

CN121991013ACN 121991013 ACN121991013 ACN 121991013ACN-121991013-A

Abstract

The application provides a method for extracting a rehmannia pigment compound from rehmannia, belonging to the fields of natural product chemistry and pharmaceutical chemistry. The preparation method comprises the steps of carrying out normal phase chromatography on a crude rehmannia pigment product to obtain a primary purification component containing a target component, carrying out reverse phase chromatography on the primary purification component to obtain a further purified sub-component, carrying out size exclusion chromatography on the sub-component to obtain a secondary component, and carrying out semi-preparative high performance liquid chromatography and/or preparative high performance liquid chromatography separation on the secondary component to obtain the rehmannia pigment compound. The method has stable process, can obtain the monomer compound with the purity of more than 99 percent, provides a high-quality material basis for subsequent development and application, and solves the problem that the prior art cannot obtain the high-purity rehmannia pigment monomer.

Inventors

  • DING GANG
  • ZHONG LIWEN
  • CHEN LIN

Assignees

  • 中国医学科学院药用植物研究所

Dates

Publication Date
20260508
Application Date
20251231

Claims (10)

  1. 1. A method for extracting a rehmannia pigment compound from rehmannia, comprising the steps of: Carrying out normal phase chromatography on the crude rehmannia pigment product to obtain a primary purification component containing a target component; subjecting the primary purified fraction to reverse phase chromatography to obtain a further purified sub-fraction; And (3) performing semi-preparative high performance liquid chromatography and/or preparative high performance liquid chromatography separation on the secondary component to obtain the rehmannia pigment compound.
  2. 2. The method according to claim 1, wherein the normal phase chromatography is normal phase silica gel column chromatography using a methylene chloride-methanol system as a mobile phase, the reverse phase chromatography is reverse phase ODS column chromatography using a water-methanol system as a mobile phase.
  3. 3. The method according to claim 1, wherein the size exclusion chromatography is performed on a sephadex LH-20 column and methanol or a dichloromethane-methanol mixed solvent is used as a mobile phase.
  4. 4. The method of claim 1, wherein the semi-preparative high performance liquid chromatography and/or preparative high performance liquid chromatography separation comprises: separating the secondary components by semi-preparative high performance liquid chromatography, wherein the semi-preparative high performance liquid chromatography adopts a reversed phase column and an acetonitrile-water mobile phase; wherein, in the acetonitrile-water mobile phase, the volume percentage concentration of acetonitrile in the water solution is 57-74%; and the semi-preparative high performance liquid chromatography is eluted in an isocratic or gradient elution mode.
  5. 5. The method according to claim 1, wherein the semi-preparative high performance liquid chromatography and/or preparative high performance liquid chromatography separation comprises separating the secondary component by chiral column semi-preparative high performance liquid chromatography, wherein the mobile phase of the chiral column semi-preparative high performance liquid chromatography is n-hexane-isopropanol mixed solvent and elution is performed by gradient elution.
  6. 6. The method of claim 2, further comprising mixing the crude rehmannia pigment with 60-100 mesh silica gel to obtain a mixed crude rehmannia pigment before normal phase chromatography of the crude rehmannia pigment, wherein the stationary phase of the normal phase silica gel column is 100-200 mesh silica gel; And the dichloromethane-methanol system is used as a mobile phase, and the mixed solvent of dichloromethane and methanol is used for carrying out gradient elution on the crude rehmannia pigment after sample stirring, wherein the gradient elution of the mixed solvent of dichloromethane and methanol is started from pure dichloromethane to pure methanol.
  7. 7. The method according to claim 2, wherein the reversed-phase ODS column chromatography uses a water-methanol system for gradient elution of the preliminary purification component, wherein the gradient elution volume ratio of the water-methanol system is changed from 9:1 to pure methanol.
  8. 8. The method according to claim 5, wherein the gradient elution mode comprises increasing the volume ratio of isopropanol in n-hexane from 10% to 13% and increasing the volume ratio from 13% to 30% after 10-30 minutes.
  9. 9. The method of claim 1, wherein prior to subjecting the crude rehmannia glutinosa dye to normal phase chromatography, further comprising: pretreating rehmannia root raw materials to obtain phloem of rehmannia root; Extracting the phloem by adopting a first organic solvent to obtain an extracting solution; Concentrating the extract to obtain an extract; Dispersing the extract in a polar solvent, performing liquid-liquid extraction by using a second organic solvent which is not miscible with the polar solvent, separating to obtain a second organic solvent phase containing the rehmannia pigment, and removing the solvent to obtain a crude product of the rehmannia pigment.
  10. 10. The method of claim 9, wherein the first organic solvent is ethyl acetate, the polar solvent is water, and the second organic solvent is dichloromethane.

Description

Method for extracting rehmannia pigment compound from rehmannia Technical Field The application relates to the fields of natural product chemistry and pharmaceutical chemistry, in particular to a method for extracting a rehmannia pigment compound from rehmannia. Background With the increasing importance of healthy life style, active ingredients derived from natural products are favored because of their wide sources and high safety. Rehmannia as a traditional Chinese medicinal material, its extract has been demonstrated to have various biological activities. In the prior art, rehmannia root is usually used for simply extracting rhizome, for example, extracting rhizome by adopting solvents such as water or ethanol and the like to obtain crude extract with complex components, and the crude extract is used for preparing health care products or cosmetics. For example, there are technical proposals disclosing cold water extraction of rehmannia root to obtain crude extract for cosmetics. However, the structure, content and biological activity of each chemical component in the crude extract are not clear, which leads to difficulty in controlling the quality of the product and limits the high-value development of the crude extract in the fields of medicines and the like. At present, no report on obtaining a high-purity rehmannia pigment monomer compound through systematic separation and purification from rehmannia has been made, and the precise chemical structure and biological activity of the rehmannia pigment monomer compound are unknown, which seriously hinders deep research and precise application of the rehmannia pigment substance basis. Disclosure of Invention The application aims to provide a method for extracting a rehmannia pigment compound from rehmannia, which aims to solve the technical problems that the chemical structure and the biological activity of the rehmannia pigment single compound are unknown and the deep development and the application of the rehmannia pigment single compound are limited due to the lack of a method for systematically separating and purifying the rehmannia pigment single compound from the rehmannia in the prior art. In order to solve the technical problems, the application provides a method for extracting a rehmannia pigment compound from rehmannia, which comprises the following steps of carrying out normal phase chromatography on a rehmannia pigment crude product to obtain a primary purification component containing a target component, carrying out reverse phase chromatography on the primary purification component to obtain a further purified sub-component, carrying out size exclusion chromatography on the sub-component to obtain a secondary component, and carrying out semi-preparative high performance liquid chromatography and/or preparative high performance liquid chromatography separation on the secondary component to obtain the rehmannia pigment compound. Alternatively, the normal phase chromatography is normal phase silica gel column chromatography and adopts a methylene dichloride-methanol system as a mobile phase, and the reversed phase chromatography is reversed phase ODS column chromatography and adopts a water-methanol system as a mobile phase. Alternatively, the size exclusion chromatography is performed on a sephadex LH-20 column with methanol or a dichloromethane-methanol mixed solvent as the mobile phase. Optionally, the semi-preparative high performance liquid chromatography and/or preparative high performance liquid chromatography separation comprises separating the secondary component by semi-preparative high performance liquid chromatography, wherein the semi-preparative high performance liquid chromatography adopts a reversed phase column and an acetonitrile-water mobile phase, the volume percentage concentration of acetonitrile in the mobile phase in an aqueous solution is 57-74%, and the elution is performed by adopting an isocratic or gradient elution mode. Optionally, the semi-preparative high performance liquid chromatography and/or preparative high performance liquid chromatography separation comprises separating the secondary component by chiral column semi-preparative high performance liquid chromatography, wherein the mobile phase is n-hexane-isopropanol mixed solvent, and eluting by gradient elution. Further, in the scheme of adopting normal phase silica gel column chromatography, before the crude rehmannia pigment is subjected to chromatography, the method further comprises the step of mixing the crude rehmannia pigment with 60-100 mesh silica gel, wherein the stationary phase of the normal phase silica gel column is 100-200 mesh silica gel, a dichloromethane-methanol system is adopted as a mobile phase, and specifically, the mixed solvent is used for carrying out gradient elution on the mixed crude product, and the gradient starts from pure dichloromethane to pure methanol. Further, in the scheme using reverse phase ODS column chromatography, the