Method for separating and purifying total flavonoids from abelmoschus manihot flowers

Abstract

The invention discloses a method for separating and purifying total flavonoids from abelmoschus manihot flowers. The method comprises the following steps: 1) an extraction process; (2) pretreating the feed liquid; 3) a flocculation process; 4) macroporous adsorption resin extraction; (5) dealcoholization; and 6) drying. The method comprises the following steps: extracting an abelmoschus manihot flower material with an ethanol salt solution, filtering, taking a treated soak solution, concentrating, dealcoholizing, diluting with water, and adding sodium lignin sulfonate and chitosan for flocculation reaction to obtain a column loading solution; selecting macroporous resin for separation and purification, carrying out rotary evaporation on the desorbed mixed solution to recover ethanol, and carrying out spray drying or freeze-drying to obtain a high-content flavone product; extraction and separation are carried out by utilizing the property of flavone, an extraction and separation method with high content and high yield is obtained, the use of a large amount of organic solvents is reduced, and environmental protection is facilitated.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of plant extraction, and particularly relates to a method for separating and purifying total flavonoids from Malva sylvestris flowers. BACKGROUND

[0002] Malva sylvestris is an annual herbaceous plant of Malvaceae Malva, which is a plant with medicinal and edible values, is praised as "plant giant panda" and "life lifesaver" in the biological world, and has great development value. Malva sylvestris is a plant with extremely rich medicinal values, which is recorded in Shennong Bencao Jing as having the effects of clearing damp-heat and relieving inflammation and pain, and is used for treating five types of stranguria and edema, and treating scalds caused by hot water.

[0003] Malva sylvestris contains flavonoids, unsaturated fatty acids, organic selenium, zinc, iron, dietary fiber, collagen, polysaccharides, organic phenolic acid and various trace elements, and is a very valuable medicinal and edible material. Among more than 200 Malva plants, Malva sylvestris has the highest utilization value in terms of edible, medicinal and health care functions.

[0004] Especially, the flowers of Malva sylvestris contain a large amount of flavonoids, including hyperoside, gossypetin-8-O-beta-D-glucuronide, isoquercitrin, quercetin-3-O-beta-D-glucopyranoside, quercetin-3-O-robinin, myricetin and quercetin, which have significant antioxidant, anti-inflammatory, anti-aging and anticancer activities.

[0005] In addition, Malva sylvestris also performs well in regulating human endocrine, enhancing immunity and improving cardiovascular microcirculation.

[0006] In terms of extraction process, domestic researchers use Soxhlet extraction, ultrasonic extraction, microwave-assisted extraction and other methods to efficiently separate and extract total flavonoids from Malva sylvestris, and optimize the extraction conditions through orthogonal experiments to improve the extraction efficiency.

[0007] Chinese patent CN 110590882B discloses a method for simultaneously separating and purifying six flavonoid compounds from Malva sylvestris flowers. First, microwave-assisted extraction is used, then solvent extraction is used to preliminarily separate flavonoids, and finally polyamide preparation chromatography is used to separate each flavonoid single component. This method can obtain products with high content, but the processing capacity is too small, the cost is high, and a large amount of solvent is used, so it can only be used in the laboratory stage.

[0008] Chinese patent CN202310421299.7 discloses a Malva sylvestris flower total flavonoid effective part extract and an industrialized mass production preparation process. The method includes alcohol extraction, concentration, oil removal by freezing, appropriate dilution according to relative density, and direct column macroporous adsorption resin. This method is based on relative density and cannot effectively indicate the content of flavonoids. The energy consumption for oil removal by freezing is too large, and the production is not suitable for use.

[0009] Chinese patent CN202410109763.3 discloses a method for mechanically and chemically assisted extraction of total flavonoids from hibiscus sabdariffa flowers, using zirconium beads as grinding medium and adding chemical additives for ultrasonic extraction. This method can help improve the extraction rate, but the operation is complex and not suitable for mass production.

[0010] Chinese patent CN202111150320.1 discloses a method for simultaneously extracting polysaccharides and flavonoids from hibiscus sabdariffa flowers. Ethanol / water system is used for reflux extraction of hibiscus sabdariffa flowers, and polysaccharides are obtained by alcohol precipitation, and flavonoids are obtained by concentrating the supernatant. This method requires two steps of concentration, which causes great loss of flavonoids and complex operation.

[0011] Therefore, to solve the problems of the prior art, such as the use of a large amount of organic solvent for extracting hibiscus sabdariffa flowers, serious pollution, high recovery cost, low resin adsorption capacity, poor anti-pollution ability, and low yield, it is an urgent technical problem for those skilled in the art to provide an extraction and separation method with high content and high yield, which can reduce the use of a large amount of organic solvent and is conducive to environmental protection. SUMMARY

[0012] Therefore, the present application provides a method for separating and purifying total flavonoids from hibiscus sabdariffa flowers.

[0013] To solve the above technical problems, the present application adopts the following technical solutions:

[0014] A method for separating and purifying total flavonoids from hibiscus sabdariffa flowers, comprising the following steps:

[0015] 1) Extraction process: hibiscus sabdariffa flowers are added to an ethanol salt solution, soaked, filtered, the obtained filter cake is added to an ethanol salt solution again, soaked, filtered, and the two soaking solutions are combined;

[0016] 2) Pretreatment of the feed liquid: the soaking solution obtained in step 1) is subjected to reduced pressure distillation to recover ethanol, obtaining a concentrated solution; the concentrated solution is diluted with water and stirred;

[0017] 3) Flocculation process: the soaking solution obtained in step 2) is added with an acid to adjust the pH of the system, then sodium lignosulfonate aqueous solution is added for flocculation reaction, stirred, allowed to stand, filtered, the obtained filtrate is adjusted to the pH of the system with an acid, then chitosan aqueous solution is added for flocculation reaction, stirred, allowed to stand, filtered or centrifuged, obtaining a treated liquid;

[0018] 4) Macroporous adsorption resin extraction: the treated liquid obtained in step 3) is subjected to dynamic adsorption, water washing, pre-elution and elution using a macroporous adsorption resin, obtaining an eluted mixed solution;

[0019] 5) Dealcoholization: the purified liquid obtained in step 4) is subjected to rotary evaporation to remove organic solvents, obtaining a residue;

[0020] 6) drying: drying the residue obtained in step 5).

[0021] Preferably, in step 1), 1 part of Malveflora is added to 8-10 parts of the salt solution, soaked at 20-50℃ for 4-8h, and filtered;

[0022] The obtained filter cake is added to 6-8 parts of the salt solution, soaked at 20-50℃ for 4-8h, and filtered;

[0023] The salt in the salt solution is sodium acetate, sodium carbonate, sodium bicarbonate, sodium phosphate, etc. weak base salt; the mass concentration of the salt solution is 0.1-0.5%; the concentration of the ethanol solution is 80-95%.

[0024] Preferably, in step 2), the distillation is carried out at 40-50℃ under reduced pressure.

[0025] Preferably, in step 2), the concentrated solution is diluted with 10-20 times of water, stirred for 0.5-1h;

[0026] The volume of the concentrated solution is 10-20% of the volume of the soaking solution.

[0027] Preferably, in step 3), the pH of the system is adjusted to 7.0±0.5 by adding acid, and then sodium lignosulfonate aqueous solution is added for flocculation reaction, the amount of which is 0.5-2% of the volume of the soaking solution, stirred, and placed for 10-30min.

[0028] Preferably, in step 3), the pH of the system is adjusted to 3.0-5.5 by adding acid, and then chitosan aqueous solution is added for flocculation reaction, the amount of which is 0.5-2% of the volume of the soaking solution, stirred, and placed for 0.5-2h.

[0029] Preferably, in step 3), the acid is formic acid, acetic acid, oxalic acid, hydrochloric acid, sulfuric acid;

[0030] The concentration of the sodium lignosulfonate aqueous solution and the chitosan aqueous solution is 1%.

[0031] Preferably, in step 4), the column loading amount for dynamic adsorption is 80-120g / L, and the flow rate is 2-4BV / h,

[0032] The adsorption amount of the resin is 70-80g / L;

[0033] The resin is a medium-polar resin;

[0034] The amount of water used for washing is 2-5BV, and the flow rate is 1-4BV / h;

[0035] The pre-dissolving agent used for pre-dissolution is an alcohol ketone aqueous solution with a mass concentration of 1-10%.

[0036] wherein the alcohol ketone is one or more of methanol, ethanol, acetone and isopropanol, the pre-elution agent is used in an amount of 1-3 BV, and the flow rate is 1-3 BV / h;

[0037] The elution agent used in the elution process is an alcohol ketone aqueous solution with a mass concentration of 20-50%;

[0038] wherein the alcohol ketone is one or more of methanol, ethanol, acetone and isopropanol;

[0039] The elution agent is used in an amount of 3-8 BV, and the elution flow rate is 0.1-2.0 BV / h.

[0040] Preferably, in the step 5), the obtained purified liquid is subjected to rotary evaporation at 30-60℃ to remove the organic solvent.

[0041] Preferably, in the step 6), the drying method is freeze-drying or spray drying.

[0042] The present application has the following technical effects relative to the prior art:

[0043] ①The method for separating and purifying total flavonoids from Malva sylvestris flowers of the present application improves the extraction rate of flavonoids from raw materials by adding high-concentration ethanol and weak base salt, reduces the alcohol precipitation step, further reduces the concentration step, and reduces the degradation rate of flavonoids in the concentration process;

[0044] ②The method for separating and purifying total flavonoids from Malva sylvestris flowers of the present application improves the adsorption capacity of the resin for effective substances by the isoelectric point method, thereby reducing the production cost;

[0045] ③The method for separating and purifying total flavonoids from Malva sylvestris flowers of the present application removes organic phenolic acid substances by overloading, thereby improving the final flavonoid content of the product;

[0046] ④The method for separating and purifying total flavonoids from Malva sylvestris flowers of the present application can effectively remove some non-polar macromolecular tannins represented by palm oil, volatile oil substances such as long-chain hydrocarbons, and further improve the product content, because the flavonoid molecule is small and has strong polarity, and the medium-polar resin with a large specific surface and small pore structure is selected;

[0047] ⑤The method for separating and purifying total flavonoids from Malva sylvestris flowers of the present application solves the problem of low yield and great waste of raw materials in the traditional process, and the resin yield of the method of the present application can reach more than 95%;

[0048] ⑥The method for separating and purifying total flavonoids from Malva sylvestris flowers of the present application can obtain a total flavonoid product with high content by gradient elution of organic solvents in the purification process. BRIEF DESCRIPTION OF DRAWINGS

[0049] Figure 1 This is a flowchart of the present invention;

[0050] Figure 2 This is a standard curve diagram for detecting rutin in this invention. Detailed Implementation

[0051] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0052] like Figure 1 As shown, this invention discloses a method for separating and purifying total flavonoids from golden sunflower flowers, comprising the following steps:

[0053] 1) Extraction process: By weight, add 1 part of golden sunflower flowers to 8-10 parts of ethanol salt solution, soak at 20-50℃ for 4-8 hours, filter, add the obtained filter cake to 6-8 parts of ethanol salt solution, soak at 20-50℃ for 4-8 hours, filter, and combine the two soaking solutions.

[0054] 2) Pretreatment of the solution: The soaking solution obtained in step 1) is distilled under reduced pressure at 40-50℃ to recover ethanol and obtain a concentrated solution. The concentrated solution is diluted with 10-20 times water and stirred for 0.5-1h.

[0055] 3) Flocculation process: The soaking solution obtained in step 2) is added to adjust the pH of the system to 7.0±0.5 with acid, and then sodium lignosulfonate aqueous solution is added to carry out the flocculation reaction. The amount added is 0.5-2% of the volume of the soaking solution. Stir, let stand for 10-30 minutes, filter, adjust the pH of the filtrate to 3.0-5.5 with acid, and then add chitosan aqueous solution to carry out the flocculation reaction. The amount added is 0.5-2% of the volume of the soaking solution. Stir, let stand for 0.5-2 hours, filter or centrifuge to obtain the treated solution;

[0056] 4) Macroporous adsorption resin extraction: The treatment solution obtained in step 3) is subjected to dynamic adsorption, water washing, pre-decomposition and desorption using macroporous adsorption resin to obtain a desorption mixture;

[0057] The loading rate for the dynamic adsorption is 80-120 g / L, and the flow rate is 2-4 BV / h.

[0058] The resin adsorption capacity is 70-80 g / L.

[0059] The amount of water used for washing is 2-5 BV, and the flow rate is 1-4 BV / h.

[0060] The pre-degrading agent used is an aqueous solution of alcohol and ketone with a mass concentration of 1-10%, wherein the alcohol and ketone are one or more of methanol, ethanol, acetone and isopropanol, the amount of pre-degrading agent is 1-3 BV, and the flow rate is 1-3 BV / h;

[0061] The eluent used in the eluent process is an aqueous solution of an alcohol-ketone with a mass concentration of 20-50%, wherein the alcohol-ketone is one or more of methanol, ethanol, acetone and isopropanol; the amount of eluent used is 3-8 BV, and the eluent flow rate is 0.1-2.0 BV / h;

[0062] 5) De-alcoholization: The purified solution obtained in step 5) is removed by rotary evaporation at 30~60℃ to remove the organic solvent, and the residue is obtained;

[0063] 6) Drying: Dry the residue obtained in step 6).

[0064] The macroporous adsorption resin column volume used in Examples 1-4 of this application is 20 mL.

[0065] The macroporous adsorption resin column used in Example 5 of this application has a volume of 100L.

[0066] Detection method: Ultraviolet spectrophotometry

[0067] like Figure 2 As shown, the preparation of the reference solution is as follows: Accurately weigh 12.5 mg of rutin reference standard into a 25 ml volumetric flask, dissolve and dilute to volume with 50% ethanol (sonication may be necessary to dissolve), cool, and prepare a reference solution with a concentration of 0.5 mg / mL. The preparation of the standard curve is as follows: Accurately measure 1 ml, 2 ml, 3 ml, 4 ml, 5 ml, and 6 ml of the reference solution into separate 25 ml volumetric flasks, add water to each to 6 ml, add 1 ml of 5% sodium nitrite solution, mix well, let stand for 6 minutes, add 1 ml of 10% aluminum nitrate solution, shake well, let stand for 6 minutes, add 10 ml of 4% sodium hydroxide solution, then add water to the mark, shake well, let stand for 15 minutes, using the corresponding reagent as a blank, measure the absorbance at a wavelength of 510 nm using ultraviolet-visible spectrophotometry, and plot the standard curve with absorbance as the ordinate and concentration as the abscissa.

[0068] Preparation of the test solution: Weigh 0.15 g of this product accurately, place it in a 25 ml volumetric flask, add 50% ethanol to make up to volume, stopper tightly, shake well, sonicate for 5 minutes, cool, filter, accurately measure 2 ml of the filtrate, place it in a 25 ml volumetric flask, dilute with water to the mark, shake well, and use as the test solution.

[0069] Example 1:

[0070] (1) Extraction process: 50 g of Hibiscus coccineus flowers was added to 500 mL of 90% ethanol solution containing 0.2% sodium carbonate, and soaked at 40°C for 6 h. The filter cake was filtered, and 400 mL of 90% ethanol solution containing 0.2% sodium carbonate was added to the filter cake, and soaked at 40°C for 4 h. The filter cake was filtered, and the two soaking solutions were combined. The mass ratio of flavonoids in the soaking solution to the material was 4.66%;

[0071] (2) Pretreatment of the material solution: the obtained soaking solution was distilled under reduced pressure at 40°C to recover ethanol, and a concentrated solution was obtained. The concentrated solution was diluted with 650 mL of water and stirred for 0.5 h.

[0072] (3) Flocculation process: the obtained material solution was added with oxalic acid to adjust the pH of the system to 7.0, and then 0.7 mL of 1% sodium lignosulfonate aqueous solution was added for flocculation reaction. After stirring and standing for 30 min, the solution was filtered. The obtained filtrate was adjusted to pH 3.5 with oxalic acid, and then 0.7 mL of 1% chitosan aqueous solution was added for flocculation reaction. After stirring and standing for 1 h, the solution was filtered to obtain a treated solution. The flavonoid concentration of the treated solution was 2.777 mg / mL, and the volume was 755 mL;

[0073] (4) Macroporous adsorption resin extraction: the obtained treated solution was subjected to dynamic adsorption on a macroporous adsorption resin LK2MG column. The flow rate was 2.5 BV / h, the water washing amount was 4 BV, the flow rate was 2 BV / h, the pre-elution agent was 3% ethanol aqueous solution, the pre-elution agent amount was 2 BV, the flow rate was 1.5 BV / h, the elution agent was 40% ethanol aqueous solution, the elution agent amount was 6 BV, and the elution flow rate was 0.8 BV / h;

[0074] (5) Dealcoholization: the obtained purified solution was subjected to rotary evaporation at 40°C to remove the organic solvent, and a residue was obtained;

[0075] (6) Drying: the residue obtained in step 6) was subjected to freeze-drying to obtain a product with a mass of 1.87 g. The product content was 80.35%, and the resin elution yield was 96.66%.

[0076] Example 2:

[0077] (1) Extraction process: 50 g of Hibiscus coccineus flowers was added to 500 mL of 90% ethanol solution containing 0.2% sodium carbonate, and soaked at 40°C for 6 h. The filter cake was filtered, and 400 mL of 90% ethanol solution containing 0.2% sodium carbonate was added to the filter cake, and soaked at 40°C for 4 h. The filter cake was filtered, and the two soaking solutions were combined. The mass ratio of flavonoids in the soaking solution to the material was 4.66%;

[0078] (2) Pretreatment of the material solution: the obtained soaking solution was distilled under reduced pressure at 40°C to recover ethanol, and a concentrated solution was obtained. The concentrated solution was diluted with 650 mL of water and stirred for 0.5 h.

[0079] (3) flocculation process: the obtained liquor, adding hydrochloric acid to adjust the system pH to 7.0, then adding 1.13 mL of 1% lignin sulfonate sodium aqueous solution to carry out flocculation reaction, stirring, standing for 30 min, filtering, the obtained filtrate is adjusted to pH 4.0 with oxalic acid, then adding 1.13 mL of 1% chitosan aqueous solution to carry out flocculation reaction, stirring, standing for 1 h, filtering to obtain a treated liquor, the treated liquor has a flavone concentration of 2.354 mg / mL and a volume of 1000 mL;

[0080] (4) macroporous adsorption resin resin extraction: the obtained treated liquor is subjected to dynamic adsorption on macroporous adsorption resin LK37, the flow rate is 3.0 BV / h, the water washing amount is 3 BV, the flow rate is 3.0 BV / h, the pre-elution agent is 7% methanol aqueous solution, the pre-elution agent amount is 3 BV, the flow rate is 1.0 BV / h, the elution agent is 45% methanol aqueous solution, the elution agent amount is 5 BV, and the elution flow rate is 1.0 BV / h;

[0081] (5) dealcoholization: the obtained purified liquor is subjected to rotary evaporation at 45℃ to remove the organic solvent, and a residue is obtained;

[0082] (6) drying: the residue obtained in step 6) is subjected to freeze-drying to obtain a product with a mass of 1.76 g, a product content of 78.56%, and a resin elution yield of 93.67%.

[0083] Example 3

[0084] (1) extraction process: 50 g of Hibiscus sabdariffa flowers is added into 450 mL of 95% ethanol solution containing 0.08% sodium bicarbonate, and soaked at 55℃ for 8 h, then filtered; the obtained filter cake is added into 400 mL of 95% ethanol solution containing 0.15% sodium bicarbonate, and soaked at 55℃ for 4 h, then filtered; the two soaking solutions are combined, and the flavone content in the soaking solution is 4.35% of the mass of the material;

[0085] (2) liquor pretreatment: the obtained soaking solution is subjected to vacuum distillation at 45℃ to recover ethanol, and a concentrated solution is obtained. The concentrated solution is diluted with 550 mL of water, and stirred for 1 h.

[0086] (3) flocculation process: the obtained liquor is added with sulfuric acid to adjust the system pH to 7.0, then added with 1.42 mL of 1% lignin sulfonate sodium aqueous solution to carry out flocculation reaction, stirring, standing for 30 min, filtering, the obtained filtrate is adjusted to pH 4.0 with sulfuric acid, then added with 1.42 mL of 1% chitosan aqueous solution to carry out flocculation reaction, stirring, standing for 1 h, filtering to obtain a treated liquor, the treated liquor has a flavone concentration of 3.263 mg / mL and a volume of 600 mL;

[0087] (4) Macroporous adsorption resin resin extraction: the obtained treatment liquid is subjected to dynamic adsorption on a macroporous adsorption resin 860021, the flow rate is 4.0 BV / h, the water washing amount is 2 BV, the flow rate is 4.0 BV / h, the pre-elution agent is a 5% isopropanol aqueous solution, the pre-elution agent amount is 3 BV, the flow rate is 1.0 BV / h, the elution agent is a 30% isopropanol aqueous solution, the elution agent amount is 3 BV, and the elution flow rate is 1.0 BV / h;

[0088] (5) Dealcoholization: the obtained purified liquid is subjected to rotary evaporation at 45°C to remove the organic solvent, and a residue is obtained;

[0089] (6) Drying: the obtained residue is subjected to freeze-drying drying, 1.752 g of product is obtained, the product content is 82.21%, and the resin elution yield is 95.81%.

[0090] Example 4:

[0091] (1) Extraction process: 50 g of Hibiscus sabdariffa flowers is added into 500 mL of 80% ethanol solution containing 0.12% sodium phosphate, and is soaked at 35°C for 5 h, and is filtered, and the obtained filter cake is added into 400 mL of 85% ethanol solution containing 0.12% sodium acetate, and is soaked at 35°C for 3 h, and is filtered, and the two soaking liquids are combined, and the flavonoids in the soaking liquid account for 5.31% of the mass of the material;

[0092] (2) Liquid pretreatment: the obtained soaking liquid is subjected to distillation under reduced pressure at 40°C to recover ethanol, and a concentrated liquid is obtained. The concentrated liquid is diluted with 1200 mL of water, and is stirred for 1 h.

[0093] (3) Flocculation process: the obtained liquid is added with hydrochloric acid to adjust the system pH to 7.0, and is added with 1.13 mL of 1% sodium lignosulfonate aqueous solution to perform flocculation reaction, is stirred, is left to stand for 30 min, is filtered, the obtained filtrate is added with oxalic acid to adjust the system pH to 4.0, and is added with 1.13 mL of 1% chitosan aqueous solution to perform flocculation reaction, is stirred, is left to stand for 1 h, is filtered, and a treatment liquid is obtained, the flavonoid concentration of the treatment liquid is 1.732 mg / mL, and the volume is 1380 mL;

[0094] (4) Macroporous adsorption resin resin extraction: the obtained treatment liquid is subjected to dynamic adsorption on a macroporous adsorption resin LK2MG, the flow rate is 3.5 BV / h, the water washing amount is 2.5 BV, the flow rate is 3.5 BV / h, the pre-elution agent is a 2% ethanol aqueous solution, the pre-elution agent amount is 4 BV, the flow rate is 1.0 BV / h, the elution agent is a 45% ethanol aqueous solution, the elution agent amount is 5 BV, and the elution flow rate is 0.5 BV / h;

[0095] (5) Dealcoholization: the obtained purified liquid was rotary evaporated at 45°C to remove the organic solvent, to obtain a residue;

[0096] (6) Drying: the obtained residue was freeze-dried to obtain a product with a mass of 1.98 g, a product content of 78.89%, and a resin elution yield of 96.22%.

[0097] Example 5:

[0098] (1) Extraction process: 240 kg of Hibiscus sabdariffa flowers was added into 2.4 m3 of 85% ethanol solution containing 0.5% sodium acetate, and was countercurrently extracted at 25°C for 6 h, and was filtered. The obtained filter residue was added into 1.9 m3 of 85% ethanol solution containing 0.5% sodium acetate, and was countercurrently extracted at 25°C for 6 h, and was filtered by a disc centrifuge. The two soaking liquids were combined, and the flavonoids in the soaking liquid accounted for 5.10% of the mass of the material;

[0099] (2) Pretreatment of the material liquid: the obtained soaking liquid was single-effect evaporated at 45°C to recover ethanol, to obtain a concentrated liquid. The concentrated liquid was diluted with 3.2 m3 of water, and was stirred for 1 h.

[0100] (3) Flocculation process: the obtained material liquid was added with hydrochloric acid to adjust the pH of the system to 7.0, and was added with 40 L of 1% sodium lignosulfonate aqueous solution to perform a flocculation reaction. After stirring and standing for 30 min, the system was filtered by a disc centrifuge. The obtained filtrate was added with hydrochloric acid to adjust the pH of the system to 4.0, and was added with 40 L of 1% chitosan aqueous solution to perform a flocculation reaction. After stirring and standing for 1 h, the system was filtered by a disc centrifuge, to obtain a treated liquid. The flavonoid concentration of the treated liquid was 3.06 mg / mL, and the volume was 3.6 m3;

[0101] (4) Macroporous adsorption resin extraction: the obtained treated liquid was subjected to dynamic adsorption on a macroporous adsorption resin LK37. The flow rate was 3.0 BV / h, the water washing amount was 3 BV, the flow rate was 3.0 BV / h, the pre-elution agent was 8% ethanol aqueous solution, the pre-elution agent amount was 3 BV, the flow rate was 1.0 BV / h, the elution agent was 40% ethanol aqueous solution, the elution agent amount was 5 BV, and the elution flow rate was 1.0 BV / h;

[0102] (5) Dealcoholization: the obtained purified liquid was rotary evaporated at 45°C to remove the organic solvent, to obtain a residue;

[0103] (6) Drying: the residue obtained in step 6) was freeze-dried to obtain a product with a mass of 9.91 kg, a product content of 82.33%, and a resin elution yield of 95.53%.

[0104] The above merely describes preferred embodiments of the present application, and is not intended to limit the technical scope of the present application in any way. Any minor modification, equivalent change and modification made to the above embodiments according to the technical essence of the present application shall still fall within the technical scope of the present application.

Claims

1. A method for isolation and purification of total flavonoids from Hibiscus mutabilis flowers, characterized by, It comprises the following steps: 1) extraction process: adding malve flower into ethanolic salt solution, soaking, filtering, adding the obtained filter cake into ethanolic salt solution again, soaking, filtering, and combining the two soaking solutions; 2) feed liquid pretreatment: distilling the soaking solution obtained in step 1) under reduced pressure to recover ethanol, and obtaining concentrated solution; Dilute the concentrated solution with water and stir; 3) flocculation process: adjusting the pH of the soaking solution obtained in step 2) by adding acid, then adding sodium lignosulfonate aqueous solution for flocculation reaction, stirring, standing, filtering, adjusting the pH of the obtained filtrate by adding acid, then adding chitosan aqueous solution for flocculation reaction, stirring, standing, filtering or centrifuging, and obtaining treated solution; 4) macroporous adsorption resin extraction: performing dynamic adsorption, water washing, pre-elution and elution on the treated solution obtained in step 3) with macroporous adsorption resin to obtain elution mixture; 5) dealcoholization: removing organic solvent from the purified solution obtained in step 4) by rotary evaporation to obtain residue; 6) drying: drying the residue obtained in step 5).

2. The method as claimed in claim 1, wherein the total flavonoids are isolated and purified from the flowers of A. auriculatum. In step 1), 1 part of malve flower is added into 8-10 parts of ethanolic salt solution, soaked at 20-50℃ for 4-8h, and filtered; The obtained filter cake is added into 6-8 parts of ethanolic salt solution again, soaked at 20-50℃ for 4-8h, and filtered; In the salt solution, the salt is sodium acetate, sodium carbonate, sodium bicarbonate, sodium phosphate and other weak base salts; the mass concentration of the salt solution is 0.1-0.5%; and the concentration of the ethanol solution is 80-95%.

3. The method as claimed in claim 1, wherein the total flavonoids are isolated and purified from the flowers of A. auriculatum. In step 2), distillation is performed under reduced pressure at 40-50℃.

4. The method as claimed in claim 1, wherein the total flavonoids are isolated and purified from the flowers of A. auriculatum. In step 2), the concentrated solution is diluted with 10-20 times of water, and stirred for 0.5-1h; The volume of the concentrated solution is 10-20% of the volume of the soaking solution.

5. The method for separating and purifying total flavonoids from golden sunflower according to claim 1, characterized in that, In step 3), the pH of the system is adjusted to 7.0±0.5 by adding acid, then sodium lignosulfonate aqueous solution is added for flocculation reaction, the addition amount is 0.5-2% of the volume of the soaking solution, stirring, standing for 10-30min.

6. The method as claimed in claim 1, wherein the total flavonoids are isolated and purified from the flowers of Aerva sanguinea (L.) Juss. In step 3), the pH of the system is adjusted to 3.0-5.5 by adding acid to the obtained filtrate, then chitosan aqueous solution is added for flocculation reaction, the addition amount is 0.5-2% of the volume of the soaking solution, stirring, standing for 0.5-2h.

7. The method for separating and purifying total flavonoids from golden sunflower according to claim 1, characterized in that, In step 3), the acid is formic acid, acetic acid, oxalic acid, hydrochloric acid, sulfuric acid; The concentration of sodium lignosulfonate aqueous solution and chitosan aqueous solution is 1%.

8. The method for separating and purifying total flavonoids from golden sunflower according to claim 1, characterized in that, In step 4), the column loading amount of dynamic adsorption is 80-120g / L, and the flow rate is 2-4BV / h, The adsorption amount of resin adsorption is 70-80g / L; The resin is a medium-polar resin; The amount of water washing is 2-5BV, and the flow rate is 1-4BV / h; The pre-elution agent used in pre-elution is an alcohol ketone aqueous solution with a mass concentration of 1-10%; The alcohol ketone is one or more of methanol, ethanol, acetone and isopropanol, the pre-elution agent is used in an amount of 1-3BV, and the flow rate is 1-3BV / h; The eluent used in the elution process is an alcohol ketone aqueous solution with a mass concentration of 20-50%; The alcohol ketone is one or more of methanol, ethanol, acetone and isopropanol; The amount of eluent is 3-8BV, and the elution flow rate is 0.1-2.0BV / h.

9. The method for separating and purifying total flavonoids from golden sunflower according to claim 1, characterized in that, In the step 5), the obtained purified solution is subjected to rotary evaporation at 30-60°C to remove the organic solvent.

10. The method for separating and purifying total flavonoids from golden sunflower according to claim 1, characterized in that, In the step 6), the drying method is freeze-drying or spray-drying.

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