阅读说明：本技术 一种木芙蓉叶总黄酮的提取制备方法 (Method for extracting and preparing total flavonoids of cotton rose leaves ) 是由 王平 蒋锦燕 江瑜 于 2019-07-31 设计创作，主要内容包括：本发明提供的一种木芙蓉叶总黄酮的提取制备方法,包括以下步骤：(1)将木芙蓉叶原料烘干、粉碎、过筛；(2)将过筛后的木芙蓉叶粉末与低共熔溶剂混合均匀,经超声辅助提取,抽滤得澄清的含有总黄酮的木芙蓉叶提取液,旋转蒸发浓缩后得到木芙蓉叶总黄酮。作为优选,所述步骤(1)中的过筛为过《中华人民共和国药典》规定的4号筛。作为优选,所述步骤(2)中,过筛后的木芙蓉叶原料与低共熔溶剂按照料液比计重量体积比为1:5～1:35的比例混合。(The invention provides an extraction and preparation method of total flavonoids in cotton rose leaves, which comprises the following steps: (1) oven drying folium Hibisci Mutabilis, pulverizing, and sieving; (2) mixing sieved folium Hibisci Mutabilis powder with eutectic solvent, ultrasonic-assisted extracting, vacuum filtering to obtain clear folium Hibisci Mutabilis extractive solution containing total flavonoids, and rotary evaporating for concentrating to obtain folium Hibisci Mutabilis total flavonoids. Preferably, the sieving in the step (1) is a No. 4 sieve specified in pharmacopoeia of the people's republic of China. Preferably, in the step (2), the sieved cotton rose hibiscus leaf raw material and the eutectic solvent are mixed according to a weight-volume ratio of the raw material to the eutectic solvent of 1: 5-1: 35.)

1. A method for extracting and preparing total flavonoids of cotton rose leaves is characterized by comprising the following steps:

(1) oven drying folium Hibisci Mutabilis, pulverizing, and sieving;

(2) mixing sieved folium Hibisci Mutabilis powder with eutectic solvent, ultrasonic-assisted extracting to obtain eutectic solvent system containing folium Hibisci Mutabilis total flavone, filtering to obtain clear extractive solution, and evaporating to concentrate to obtain folium Hibisci Mutabilis total flavone;

wherein the eutectic solvent is selected from a mixed solution of choline chloride, ethylene glycol and water, a mixed solution of choline chloride, glycerol and water, a mixed solution of choline chloride, 1, 3-butanediol and water, a mixed solution of choline chloride, oxalic acid and water, a mixed solution of choline chloride, citric acid and water, a mixed solution of choline chloride, urea and water, a mixed solution of choline chloride, glucose and water or a mixed solution of choline chloride, acetamide and water.

2. The method for extracting and preparing total flavonoids from cotton rose leaves according to claim 1, wherein in the step (2), the sieved cotton rose leaf raw material and the eutectic solvent are mixed according to a material-liquid ratio of 1: 5-1: 35.

3. The method for extracting and preparing total flavonoids from cotton rose hibiscus leaves as claimed in claim 1, wherein the molar ratio of choline chloride to ethylene glycol in the mixed solution of choline chloride, ethylene glycol and water in step (2) is 1:2, the molar ratio of choline chloride to glycerol in the mixed solution of choline chloride, glycerol and water is 1:2, the molar ratio of choline chloride to 1, 3-butanediol in the mixed solution of choline chloride, 1, 3-butanediol and water is 1:2, the molar ratio of choline chloride to oxalic acid in the mixed solution of choline chloride, oxalic acid and water is 1:2, the molar ratio of choline chloride to citric acid in the mixed solution of choline chloride, citric acid and water is 1:2, the molar ratio of choline chloride to urea in the mixed solution of choline chloride, urea and water is 1:2, the molar ratio of choline chloride to glucose in the mixed solution of choline chloride, glucose and water is 1:2, the molar ratio of choline chloride to acetamide in the mixed solution of choline chloride, acetamide and water is 1: 2.

4. The method for extracting total flavonoids from cotton rose hibiscus leaves as claimed in claim 1, wherein the water content of the eutectic solvent in the step (2) is not higher than 80%.

5. The method for extracting and preparing total flavonoids from cotton rose hibiscus leaves as claimed in claim 1, wherein in the step (2), the ultrasonic-assisted extraction time is 10-60 min, and the ultrasonic-assisted extraction temperature is 20-80 ℃.

6. The method for extracting and preparing total flavonoids from cotton rose leaves according to claim 1, wherein in the step (2), the suction filtration solution of the cotton rose leaf extract is subjected to a color reaction with sodium nitrite, and the total flavonoids concentration is calculated by measuring the absorbance with an ultraviolet spectrophotometer.

7. The method for extracting and preparing total flavonoids from cotton rose leaves according to claim 1 or 2, wherein the weight volume ratio of the sieved cotton rose leaf raw material to the eutectic solvent is 1:30 according to the material-liquid ratio of the material-liquid ratio.

8. The method for extracting total flavonoids from hibiscus mutabilis leaves as claimed in claim 1 or 4, wherein the eutectic solvent has a water content of 40%.

9. The method for extracting total flavonoids from hibiscus mutabilis leaves as claimed in claim 1 or 5, wherein the extraction time is 30min and the extraction temperature is 60 ℃.

10. A method for extracting and preparing total flavonoids of cotton rose leaves is characterized by comprising the following steps:

(1) drying folium Hibisci Mutabilis in a vacuum forced air drying oven at 50 deg.C, pulverizing, and sieving with No. 4 sieve specified in pharmacopoeia of the people's republic of China;

(2) uniformly mixing the sieved cotton rose leaf powder with a mixed solution eutectic solvent of choline chloride, ethylene glycol and water, wherein the mixed solution eutectic solvent contains 40% of water and the molar ratio of the choline chloride to the ethylene glycol is 1:2, the weight-volume ratio of the choline chloride to the ethylene glycol is 1:30, performing ultrasonic-assisted extraction at the temperature of 60 ℃ for 30min to obtain a eutectic solvent system containing the total flavonoids in the cotton rose leaves, performing suction filtration to obtain a clear extracting solution, and performing rotary evaporation and concentration to obtain the total flavonoids in the cotton rose leaves.

Technical Field

The invention belongs to the field of medicinal material extraction processes, and particularly relates to a method for extracting and preparing total flavonoids in cotton rose leaves.

Background

Hibiscus mutabilis leaves are a traditional Chinese medicine in China, are called Hibiscus mutabilis leaves and antifrost leaves, have long-term origins and are recorded in numerous ancient medical books. The characteristics of the traditional Chinese medicine are recorded in the Ben Cao Tu Jing and the Ben Cao gang mu, and the traditional Chinese medicine is pungent, mild and nontoxic in taste, and has the pharmacological effects of cooling blood, detoxifying, reducing swelling, relieving pain and the like. Modern pharmacological tests prove that the effective extract components of the cotton rose hibiscus leaves can inhibit insects and bacteria in vitro, resist nonspecific inflammation, protect kidney ischemia reperfusion injury and the like. Wherein, the flavonoid glycoside is one of the main active ingredients, mainly comprises rutin, kaempferol-3-O-beta-rutinoside, kaempferol-3-O-beta-robinia diglucoside and the like, and has obvious clinical effect. The modern literature reports that the extraction methods of the total flavonoids in the leaves of the cotton rose comprise a water extraction method, an organic solvent extraction method, an ultrasonic method, a microwave-assisted method and the like. Due to the defects of low efficiency, organic solvent residue, large loss of extracted components and the like of the traditional extraction method, the application of a plurality of novel solvent extraction methods in the total flavonoids in cotton rose leaves needs to be researched urgently.

In recent years, along with the development and innovation of science and technology, people are increasingly conscious of environmental protection, and more attention is paid to the application of green chemistry. The low-toxicity, pollution-free, environment-friendly and safe green chemical reagent is more and more sought by students, and the eutectic solvent is used as a novel medium solvent, has unique dissolving capacity, can selectively dissolve inorganic or organic materials and other components, has polarity, and has good application prospect in the extraction and separation process of traditional Chinese medicinal materials and the like. The eutectic solvent is an ionic liquid consisting of a hydrogen bond acceptor (such as quaternary ammonium salt and the like) and a hydrogen bond donor (such as saccharides, amino acid and the like), has the characteristics of low melting point, simple composition, greenness, safety, low price, easy degradation and the like, and is proved to be an efficient extracting agent adapting to modern requirements in relevant experimental reports. Moreover, the eutectic solvents are various in variety, different eutectic solvent systems can be designed and combined by changing different proportions of quaternary ammonium salt and the like and hydrogen bond donors, the properties are changed, and the variability proves the special potential of the eutectic solvent system which is different from the ordinary reagent. Loli et al (CN201711061313.8) were prepared by the expression [ N (CH)₃)₃CH2CH2OH]+[X·zY]Extraction of C-flavonoid glycosides from globeflower with choline eutectic solventThe choline eutectic solvent provided by the invention can be used for efficiently and selectively extracting three important flavone C-glycosides in globeflower, and compared with the traditional methanol solvent extraction, the choline eutectic solvent not only can improve the extraction rate, but also has few impurities in the extract, so that a silica gel column is not required to be repeatedly used for separation in the purification process, the separation and purification steps are greatly reduced, and the separation and purification cost is reduced.

Zhao Longshan et al (CN201810149224.7) extracted flavonoid glycoside compounds from herba Epimedii with eutectic solvent, especially choline eutectic solvent, as extractant and water as diluent to reduce system viscosity under ultrasonic enhancement, and found that the extraction method has high extraction efficiency of flavonoid glycoside components, and the extraction efficiency is close to 100%. Besides mild extraction conditions and simple operation, the method also has the characteristics of non-explosive, non-volatile, low loss, low energy consumption and the like of the extracting agent, and can be used for efficient green large-scale production of flavonoid glycoside compounds and realizing comprehensive utilization of epimedium herb resources. Hertel R.M. et al (WO/2012/145522), Naik P.K. et al (2016, Fluid Phase Equilibria, 423, 146-. The general flavone of cotton rose hibiscus leaves is the main active ingredient in cotton rose hibiscus leaves, such as rutin, kaempferol-3-O-beta-rutinoside, kaempferol-3-O-beta-robinia diglucoside and the like.

Disclosure of Invention

The invention provides an extraction and preparation method of total flavonoids in cotton rose leaves, which comprises the following steps:

(1) oven drying folium Hibisci Mutabilis, pulverizing, and sieving;

(2) mixing sieved folium Hibisci Mutabilis powder with eutectic solvent, ultrasonic-assisted extracting, vacuum filtering to obtain clear folium Hibisci Mutabilis extractive solution containing total flavonoids, and rotary evaporating for concentrating to obtain folium Hibisci Mutabilis total flavonoids. Preferably, the sieving in the step (1) is a No. 4 sieve specified in pharmacopoeia of the people's republic of China. Preferably, in the step (2), the sieved cotton rose hibiscus leaf raw material and the eutectic solvent are mixed according to a weight-volume ratio of the raw material to the eutectic solvent of 1: 5-1: 35.

Preferably, in the step (2), the material-liquid ratio of the sieved cotton rose hibiscus leaf raw material to the eutectic solvent is 1:30 by weight.

Preferably, the eutectic solvent in the step (2) is selected from a group consisting of a mixed solution of choline chloride, ethylene glycol and water, a mixed solution of choline chloride, glycerol and water, a mixed solution of choline chloride, 1, 3-butanediol and water, a mixed solution of choline chloride, oxalic acid and water, a mixed solution of choline chloride, citric acid and water, a mixed solution of choline chloride, urea and water, a mixed solution of choline chloride, glucose and water, and a mixed solution of choline chloride, acetamide and water.

Preferably, the molar ratio of choline chloride to ethylene glycol in the mixed solution of choline chloride, ethylene glycol and water is 1:2, the molar ratio of choline chloride to glycerol in the mixed solution of choline chloride, glycerol and water is 1:2, the molar ratio of choline chloride to 1, 3-butanediol in the mixed solution of choline chloride, 1, 3-butanediol and water is 1:2, the molar ratio of choline chloride to oxalic acid in the mixed solution of choline chloride, oxalic acid and water is 1:2, the molar ratio of choline chloride to citric acid in the mixed solution of choline chloride, citric acid and water is 1:2, the molar ratio of choline chloride to urea in the mixed solution of choline chloride, urea and water is 1:2, the molar ratio of choline chloride to glucose in the mixed solution of choline chloride, glucose and water is 1:2, the mol ratio of choline chloride to acetamide in the mixed solution of choline chloride, acetamide and water is 1: 2.

Preferably, in the step (2), the eutectic solvent has a water content of not higher than 80%.

Preferably, in the step (2), the eutectic solvent has a water content of 40%.

Preferably, in the step (2), the ultrasonic-assisted extraction time is 10-60 min, and the ultrasonic-assisted extraction temperature is 20-80 ℃.

Preferably, in the step (2), the ultrasound-assisted extraction time is 30min, and the ultrasound-assisted extraction temperature is 60 ℃.

Preferably, in the step (2), the suction filtration solution of the cotton rose hibiscus leaf extract is treated with NaNO₂-Al(NO₃)₃-NaOH, measuring absorbance by using an ultraviolet spectrophotometer, and calculating the concentration of total flavonoids.

The invention provides an extraction preparation method of total flavonoids in cotton rose leaves, which has the advantages of green and safe extraction solvent, mild reaction conditions, simple operation, high extraction efficiency and the like, is wide in application range, and can be used for extracting active ingredients from traditional Chinese medicinal materials.

Drawings

FIG. 1: rutin standard curve

Detailed Description

The inventor finds that the method for extracting and preparing the total flavonoids in the leaves of the cotton rose has the advantages of green and safe extraction solvent, mild reaction conditions, simple operation, high extraction efficiency and the like, has wide application range, and can be used for extracting active ingredients from Chinese medicinal materials.

The inventor adds sodium nitrite into a flavone standard sample solution with known concentration to perform color reaction, then measures the ultraviolet absorbance, makes a standard curve, and then uses the standard curve to quantify the concentration of total flavonoids in cotton rose leaves obtained by extraction without experimental conditions, thereby obtaining the optimal experimental conditions and using the best embodiment to prove the superiority of the extraction preparation method.

The present invention will be described in detail with reference to specific examples below: