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CN-122010887-A - Method for preparing high-purity isoquercitrin by utilizing polygonum hydropiper

CN122010887ACN 122010887 ACN122010887 ACN 122010887ACN-122010887-A

Abstract

The invention provides a method for preparing high-purity isoquercitrin from polygonum orientale, which comprises the steps of sequentially carrying out water extraction, impurity removal and alcohol extraction on polygonum orientale raw materials to obtain an alcohol extract, decoloring the alcohol extract, adjusting the pH value to 9-11 to precipitate and remove tannins, concentrating under reduced pressure to obtain a crude product of total flavonoids, carrying out acid hydrolysis on the crude product, separating and purifying by adopting a high-pressure preparative liquid chromatography, carrying out on-line monitoring by utilizing an ultraviolet detector, obtaining a fraction rich in isoquercitrin by accurately controlling start and stop points collected by the fraction, and carrying out reduced pressure concentration, water washing and drying to obtain a product. According to the method, the pretreatment and chromatographic separation conditions are optimized, so that the product purity and the yield are synchronously improved, the obtained isoquercitrin HPLC purity reaches more than 98%, the process operation cost is low, the environment is friendly, and the method is suitable for large-scale production.

Inventors

  • LI XIAOJIE
  • YANG JIANGANG
  • LI YINPING
  • NI YAHUI
  • ZHANG QIAO
  • WANG WEI

Assignees

  • 陕西医药控股医药研究院有限公司

Dates

Publication Date
20260512
Application Date
20251219

Claims (10)

  1. 1. A method for preparing high-purity isoquercitrin from polygonum hydropiper, which is characterized by comprising the following steps: pretreating and extracting raw materials, namely sequentially carrying out water extraction, impurity removal and alcohol extraction on the polygonum capitatum raw materials, and collecting an alcohol extract; Removing impurities and enriching total flavonoids, namely decoloring the alcohol extract, regulating the pH value to 9-11, standing for precipitation, removing precipitates, concentrating the filtrate under reduced pressure, separating out and collecting crude products of the total flavonoids; Acid hydrolysis, namely hydrolyzing the crude product of the total flavone by using an acidified alcohol reagent to obtain hydrolysis liquid; chromatographic separation and purification, namely taking the hydrolysate as a sample loading liquid, and adopting high-pressure liquid chromatography to perform separation and purification; the method comprises the steps of taking a chromatographic column with octadecylsilane chemically bonded silica as a filler as a stationary phase, and taking a methanol-phosphoric acid aqueous solution as a mobile phase for elution, adopting an ultraviolet detector for online monitoring an elution process, controlling a fraction collector to start collecting when a chromatographic peak starts to rise according to a monitored isoquercitrin chromatographic peak signal, and stopping collecting when the chromatographic peak falls to one fifth to one sixth of the peak height to obtain an elution fraction rich in isoquercitrin; and (3) post-treatment, namely concentrating, washing and drying the eluted fraction rich in isoquercitrin to obtain the high-purity isoquercitrin.
  2. 2. The method of claim 1, wherein the step of determining the position of the substrate comprises, In the raw material pretreatment and extraction steps, water is used as a solvent, the liquid-solid ratio is 10-15 mL/g, the extraction is carried out for 1-3 times at 90-100 ℃ for 1-3 hours each time, the alcohol extraction is carried out by using ethanol solution with the mass fraction of more than 75% as the solvent, the liquid-solid ratio is 6-12 mL/g, and the extraction is carried out for 2-3 times at 60-85 ℃ for 1.5-2 hours each time.
  3. 3. The method of claim 1, wherein the step of determining the position of the substrate comprises, In the impurity removal and total flavone enrichment step, activated carbon with the mass of 0.3% -1.0% is added into the alcohol extract, stirring and adsorbing are carried out for 15-60 minutes at the temperature of 40-80 ℃, and the pH value of the decolorized liquid is adjusted to 9-11 by adopting a sodium hydroxide solution with the concentration of 0.5-1.0 mol/L, and the decolorized liquid is kept stand for 8-12 hours.
  4. 4. The method of claim 1, wherein the step of determining the position of the substrate comprises, And before the acid hydrolysis step, the method further comprises the step of washing the crude product of the total flavone with water to pH 6-8.
  5. 5. The method of claim 1, wherein the step of determining the position of the substrate comprises, In the acid hydrolysis step, the acidified alcohol reagent is a solution prepared by mixing methanol and hydrochloric acid solution with the mass fraction of 3% -15% according to the volume ratio of (30-60) to (40-70), and the hydrolysis condition is that the ratio of total flavone crude product to acidified alcohol reagent feed liquid is 1:20-40 g/mL, and the hydrolysis is carried out for 0.5-2.0 hours at 50-80 ℃.
  6. 6. The method of claim 1, wherein the step of determining the position of the substrate comprises, In the chromatographic separation and purification step, the methanol-phosphoric acid aqueous solution is formed by mixing methanol and phosphoric acid aqueous solution with the mass fraction of 0.2% -0.8% according to the volume ratio of (55-65) to (35-45), and the eluting flow rate is 10-200 mL/min.
  7. 7. The method of claim 1, wherein the step of determining the position of the substrate comprises, In the chromatographic separation and purification step, the particle size of octadecylsilane chemically bonded silica gel serving as a filler of the chromatographic column is 5-20 mu m, the pore diameter is 40-70A, the column length of the chromatographic column is 10-30 cm, and the diameter is 2-15 cm.
  8. 8. The method of claim 1, wherein the step of determining the position of the substrate comprises, In the chromatographic separation and purification step, a mode of sample loading in a separated mode is adopted, and the sample loading amount of the hydrolysate is 10-20 mL each time.
  9. 9. The method of claim 1, wherein the step of determining the position of the substrate comprises, In the post-treatment step, the washing temperature is 80-100 ℃, and the drying temperature is 50-80 ℃.
  10. 10. The method of claim 1, wherein the step of determining the position of the substrate comprises, And after the post-treatment step, the method further comprises a chromatographic column regeneration step of activating and regenerating the chromatographic column by using hydrochloric acid solution with the mass fraction of 3% -7% when the column efficiency of the chromatographic column is reduced, and then flushing the chromatographic column by using acetonitrile water solution with the mass fraction of 2% -5%.

Description

Method for preparing high-purity isoquercitrin by utilizing polygonum hydropiper Technical Field The invention relates to the technical field of natural product extraction and separation, in particular to a method for preparing high-purity isoquercitrin by utilizing polygonum hydropiper. Background Isoquercitrin is a flavonoid compound with important medicinal value, and widely exists in various plants, wherein the polygonum aviculare is one of main natural sources. At present, the conventional method for extracting isoquercitrin from plants generally has the problems of complex process, low separation efficiency, difficulty in considering product purity and yield, and the like, and is difficult to realize large-scale and low-cost production of high-purity isoquercitrin. Therefore, the development of an extraction and purification process which is efficient, stable and suitable for industrial production has important practical significance and application requirements. In the prior art, although various chromatographic separation methods are used for purifying natural products, when the method is applied to large-scale preparation of high-purity isoquercitrin from polygonum hydropiper, common technical bottlenecks of lengthy pretreatment steps, easy pollution of chromatographic columns, high separation cost, difficulty in simultaneously ensuring high purity, high yield and the like are often faced. In particular to a complete process scheme which can integrate raw material pretreatment, selective impurity removal and high-efficiency chromatographic separation systematically and realize precise control and continuous operation. Disclosure of Invention The invention aims to provide a method for preparing high-purity isoquercitrin by utilizing polygonum aviculare, which at least solves the problem that low-cost and high-efficiency large-scale production of high-purity isoquercitrin cannot be realized in the prior art. In order to achieve the above object, the present invention provides a method for preparing high purity isoquercitrin from polygonum aviculare, which is characterized by comprising the following steps: pretreating and extracting raw materials, namely sequentially carrying out water extraction, impurity removal and alcohol extraction on the polygonum capitatum raw materials, and collecting an alcohol extract; Removing impurities and enriching total flavonoids, namely decoloring an alcohol extract, regulating the pH value to 9-11, standing for precipitation, removing precipitates, concentrating filtrate under reduced pressure, separating out and collecting a crude product of total flavonoids; Acid hydrolysis, namely hydrolyzing the crude product of the total flavone by using an acidified alcohol reagent to obtain hydrolysis liquid; chromatographic separation and purification, namely taking the hydrolysate as a sample loading liquid, and adopting high-pressure liquid chromatography to perform separation and purification; the method comprises the steps of taking a chromatographic column with octadecylsilane chemically bonded silica as a filler as a stationary phase, and taking a methanol-phosphoric acid aqueous solution as a mobile phase for elution, adopting an ultraviolet detector for online monitoring an elution process, controlling a fraction collector to start collecting when a chromatographic peak starts to rise according to a monitored isoquercitrin chromatographic peak signal, and stopping collecting when the chromatographic peak falls to one fifth to one sixth of the peak height to obtain an elution fraction rich in isoquercitrin; and (3) post-treatment, namely concentrating, washing and drying the eluted fraction rich in isoquercitrin to obtain the high-purity isoquercitrin. In the raw material pretreatment and extraction steps, water is used as a solvent, the liquid-solid ratio is 10-15 mL/g, the extraction is carried out for 1-3 times at 90-100 ℃ for 1-3 hours each time, the alcohol extraction is carried out by using ethanol solution with the mass fraction of more than 75% as the solvent, the liquid-solid ratio is 6-12 mL/g, and the extraction is carried out for 2-3 times at 60-85 ℃ for 1.5-2 hours each time. The method comprises the steps of washing the finished product of the medicinal material formed by processing the root or/and the stem (with the skin) of the polygonum hydropiper with water in advance, airing to obtain the raw material, and leaching the raw material from the polygonum hydropiper in water to remove water-soluble impurities such as saccharides, part of tannins and the like contained in the raw material. In the steps of impurity removal and total flavone enrichment, activated carbon with the mass of 0.3% -1.0% is added into the alcohol extract, stirring and adsorption are carried out for 15-60 minutes at the temperature of 40-80 ℃, and the pH value of the decolorized liquid is adjusted to 9-11 by adopting a sodium hydroxide solution with the concentration of 0.5-1.0 mol