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CN-121987677-A - Antibacterial active component grading preparation method based on hundred-color mango peel residues and application

CN121987677ACN 121987677 ACN121987677 ACN 121987677ACN-121987677-A

Abstract

The invention relates to a preparation method of antibacterial active components based on a hundred-color mango peel residue, which comprises the steps of crushing the dried mango peel residue, performing enzymolysis pretreatment by adopting a compound enzyme system containing cellulase and pectase, performing ultrasonic auxiliary extraction by adopting a eutectic solvent with a choline chloride-glycerol molar ratio of 1:2, obtaining a crude extract after centrifugal separation, loading the crude extract on a macroporous adsorption resin column, performing gradient elution by adopting an ethanol-water solution containing 1-3% hydroxypropyl-beta-cyclodextrin as a protective eluent, collecting active components, directly cutting the active components into a preparation type high-efficiency liquid chromatography system by adopting a column switching technology, collecting chromatographic peaks with retention time of 14.5-15.5 min, and volatilizing the solvent to obtain the active components of targeted anti-malassezia. The method can be used for preparing medicine or cosmetic for inhibiting malassezia.

Inventors

  • YANG ZHENGZHOU
  • XIE XIAONA
  • WANG HUI
  • LIU JING
  • Ban Guiyuan

Assignees

  • 百色学院

Dates

Publication Date
20260508
Application Date
20260318

Claims (9)

  1. 1. The grading preparation method of the antibacterial active component based on the residue of the trichromatic mango peel is characterized by comprising the following steps of: crushing the dried mango peel residues, and performing enzymolysis pretreatment by adopting a compound enzyme system containing cellulase and pectase; ultrasonic-assisted extraction is carried out on the material subjected to enzymolysis by adopting a eutectic solvent with the choline chloride-glycerol molar ratio of 1:2, and crude extract is obtained after centrifugal separation; loading the crude extract to a macroporous adsorption resin column, performing gradient elution by adopting protective eluent containing beta-cyclodextrin, and collecting eluent in the protective elution stage as an active component; The collected active components are directly cut into a preparation type high performance liquid chromatography system by adopting a column switching technology, chromatographic grade methanol-water solution is used as a mobile phase for isocratic elution, chromatographic peaks with retention time of 14.5-15.5min are collected, and the active components of the targeted anti-malassezia are obtained after the solvent is volatilized; wherein the protective eluent containing beta-cyclodextrin is ethanol-water solution containing 1-3% hydroxypropyl-beta-cyclodextrin.
  2. 2. The method for the fractional preparation of the antibacterial active ingredient based on the residue of the trichromatic mango peel according to claim 1, wherein the gradient elution is a two-step gradient elution comprising a non-protective elution phase and a protective elution phase: The first step, eluting with ethanol-water solution without hydroxypropyl-beta-cyclodextrin, wherein the volume percentage concentration of ethanol is 20-30%, and the eluting volume is 2-4 times of the column volume; and secondly, eluting with ethanol-water solution containing 1-3% of hydroxypropyl-beta-cyclodextrin, wherein the volume percentage concentration of ethanol is 40-70%, the flow rate during elution is 0.5-1.5mL/min, the column temperature is maintained at 25-35 ℃ during elution, and the eluent in the second step is collected as an active component.
  3. 3. The method for preparing the antibacterial active ingredient based on the residue of the trichromatic mango peel according to claim 2, wherein the second step of elution is a multi-step gradient elution: The first step, eluting 1-2 times of column volume by ethanol-water solution which does not contain hydroxypropyl-beta-cyclodextrin and has the volume percentage concentration of 40-50 percent of ethanol; Secondly, eluting 1-2 times of column volume by using ethanol-water solution containing 1-3% of hydroxypropyl-beta-cyclodextrin and 40-50% of ethanol in percentage by volume; and thirdly, eluting with ethanol-water solution containing 1-3% w/v of hydroxypropyl-beta-cyclodextrin and 60-70% of ethanol by volume percentage concentration until the effluent liquid has no characteristic ultraviolet absorption under the detection wavelength of 280 nm.
  4. 4. A process for the fractional preparation of a bacteriostatic active ingredient based on the residue of the trichromatic mango peel according to claim 3, characterized in that in said second elution, the concentration of hydroxypropyl- β -cyclodextrin is graded according to one of the following modes: in one mode, the concentration of hydroxypropyl-beta-cyclodextrin increases from 1% to 3% at a constant rate over time; In the second mode, the concentration of the hydroxypropyl-beta-cyclodextrin is firstly 1% to elute 0.5-1 times of the column volume, then the concentration of the hydroxypropyl-beta-cyclodextrin is 2% to elute 0.5-1 times of the column volume, and finally the concentration of the hydroxypropyl-beta-cyclodextrin is 3% to elute until the effluent liquid has no characteristic ultraviolet absorption under the detection wavelength of 280 nm.
  5. 5. The method for preparing the antibacterial active component based on the residue of the trichromatic mango peel according to claim 4, wherein the hydroxypropyl-beta-cyclodextrin is eluted by adopting a gradient concentration, and the specific steps are as follows: The first step, eluting 1.5 times of column volume by using ethanol-water solution containing 1% hydroxypropyl-beta-cyclodextrin and having the volume percentage concentration of ethanol of 50% at the flow rate of 0.8 mL/min; secondly, eluting 1.5 times of column volume by using an ethanol-water solution containing 2% of hydroxypropyl-beta-cyclodextrin and having the volume percentage concentration of ethanol of 60% at the flow rate of 1.0 mL/min; And thirdly, eluting with an ethanol-water solution containing 3% of hydroxypropyl-beta-cyclodextrin and 70% of ethanol by volume percentage at a flow rate of 1.2 mL/min until the effluent has no characteristic ultraviolet absorption at the detection wavelength of 280 nm.
  6. 6. The method for preparing the antibacterial active ingredient based on the residue of the tricolor peel according to any one of claims 1 to 5, wherein the active ingredient contains flavonoid compounds and phenolic acid compounds, and the active ingredient has a characteristic absorption peak at a retention time of 14.5 to 15.5 min in a high performance liquid chromatogram, and the compound corresponding to the characteristic peak has an inhibitory activity on malassezia.
  7. 7. The method for preparing the antibacterial active ingredient based on the residue of the trichromatic mango peel according to claim 6, wherein the active ingredient is detected by high performance liquid chromatography-mass spectrometry analysis in a positive ion mode, and a characteristic ion peak with a mass-to-charge ratio of m/z 581.2 is detected, and the molecular weight of a compound corresponding to the characteristic ion peak is 580.2.
  8. 8. Use of the hierarchical preparation method of antibacterial active ingredients based on residue of tricolor peel according to claim 6 or 7 for preparing a medicament or cosmetic for inhibiting malassezia.
  9. 9. The use according to claim 8, wherein the malassezia comprises at least one of malassezia furfur, malassezia globosa or malassezia syn.

Description

Antibacterial active component grading preparation method based on hundred-color mango peel residues and application Technical Field The invention relates to the technical field of natural product extraction and separation, in particular to a method for preparing antibacterial active components in a grading manner based on a residue of a hundred-color mango peel and application thereof. Background Mango is a popular tropical fruit worldwide that produces a lot of peel residues during processing into fruit juice, jam, preserved fruit, etc. The hundred-color mango is used as a Chinese geographic marking product, the yield is huge, and the waste of the mango peel is generated, so that not only is the resource wasted, but also a certain pressure is formed on the environment. Researches show that the mango peel is rich in various bioactive components such as polyphenol, flavonoid and the like, and has potential application values such as antioxidation, antibiosis and the like. Therefore, how to efficiently and economically utilize the mango peel residues and separate the functional components with high added value from the residues has become one of the hot spots of current researches. In the prior art, extraction and separation of active ingredients from plant materials such as mango peel and the like, methods such as organic solvent extraction or reflux extraction are generally adopted. However, the traditional methods have the defects of high extraction temperature, long time consumption, high organic solvent consumption, more impurities in the extract and the like. Particularly for structurally unstable, easily oxidized phenolic compounds, high temperature and prolonged operation may lead to degradation and inactivation thereof, thereby affecting the biological activity of the final product. In order to improve the extraction efficiency and the product purity, some researches try to introduce an enzymolysis pretreatment technology, and destroy the cell wall structure of plants by cellulase, pectase and the like so as to promote the release of intracellular active substances. In addition, ultrasonic-assisted extraction, macroporous adsorption resin separation and other techniques are also widely used in the purification process of natural products. However, there are significant disadvantages in the prior art in terms of the preparation of targeted bacteriostatic active ingredients against specific pathogenic bacteria (such as malassezia) in the mango peel residue. Malassezia is a conditional pathogenic fungus commonly found on the surface of human skin and is closely related to various skin diseases such as seborrheic dermatitis, tinea versicolor and the like. The efficient and low-toxicity malassezia-resistant active ingredients are screened and prepared from the mango peel, and the method has important significance for developing novel skin care products or medicines. However, the existing extraction and separation processes generally lack precise protection of target active ingredients, which results in loss of activity in the separation process, and meanwhile, the multi-step separation operation is complex, and high-purity target active ingredients are difficult to obtain due to loss or dilution of the components in the switching process. Therefore, how to establish a preparation method capable of effectively protecting active ingredients and realizing efficient and accurate separation so as to obtain a targeting component with anti-malassezia activity from the residue of the tricolor peel is still a technical problem to be solved in the field. Disclosure of Invention It is an object of the present invention to address at least the above-mentioned drawbacks and to provide at least the advantages to be described later. In order to achieve the purposes and other advantages of the invention, a preparation method of a antibacterial active component based on the residue of the tricolor mango peel is provided, which comprises the following steps of crushing the residue of the dried mango peel, performing enzymolysis pretreatment by adopting a compound enzyme system containing cellulase and pectase, performing ultrasonic-assisted extraction on the material subjected to enzymolysis by adopting a eutectic solvent with the molar ratio of choline chloride to glycerol of 1:2, obtaining a crude extract after centrifugal separation, loading the crude extract on a macroporous adsorption resin column, performing gradient elution by adopting a protective eluent containing beta-cyclodextrin, collecting the eluent in the protective elution stage as an active component, performing isocratic elution by adopting a column switching technology on the collected active component, collecting chromatographic peaks with the retention time of 14.5-15.5 min, and volatilizing the solvent to obtain the active component of the targeted anti-malassezia bacteria, wherein the protective eluent containing beta-cyclodextrin is ethanol-beta-aq