Method for improving solubility of quercetin and application thereof

Abstract

The application discloses a method for improving solubility of quercetin and application thereof, and belongs to the technical field of chemistry and plant extraction. The method for improving solubility of quercetin comprises the following steps: S1, mixing choline chloride with water and uniformly stirring and heating to obtain a choline chloride aqueous solution; and S2, mixing quercetin with the choline chloride aqueous solution and performing ultrasonic treatment to obtain a high-concentration quercetin solution. The method for improving solubility of quercetin is simple and easy to operate, is green and has little influence on the environment, and is simple and efficient in solvent preparation process, and the solubility is significantly improved. The method successfully solves the problem of poor solubility of quercetin, can be widely applied to research work of extracting quercetin from plants, and provides strong technical support for industrialized production of quercetin.

Description

Technical Field

[0001] This invention belongs to the field of chemical and plant extraction technology, specifically relating to a method for improving the solubility of quercetin and its application. Background Technology

[0002] Quercetin, a naturally occurring polyhydroxyflavonoid (3,3',4',5,7-pentahydroxyflavone), is widely found in the human diet, particularly in onions, wine, berries, and tea. It possesses a variety of important physiological functions, including antioxidant and free radical scavenging, anti-inflammatory, antiviral, antitumor, antidiabetic, cardiovascular protection, and immunomodulation. However, the low solubility of quercetin monomers in common solvents (such as water and ethanol) significantly limits the effectiveness of its extraction and purification from plants. Therefore, improving the solubility of quercetin is crucial for ensuring efficient extraction and purification, thereby expanding its potential applications.

[0003] In recent years, researchers have primarily employed hot ethanol extraction or alkaline water extraction methods for the extraction and purification of quercetin. While quercetin exhibits relatively high solubility in hot ethanol, the time-consuming filtration and separation process leads to quercetin precipitation and loss in the extraction residue, resulting in incomplete extraction. Furthermore, the alkaline dissolution and acid precipitation process generates acidic and alkaline wastewater, causing environmental pollution. Therefore, developing a novel, efficient, low-cost, environmentally friendly, and easily industrially applicable solvent to enhance the solubility of quercetin monomers and applying it to the extraction and purification of quercetin from plants is of significant practical importance. Summary of the Invention

[0004] In view of this, the purpose of this invention is to provide a method for improving the solubility of quercetin and its application. This method successfully solves the problem of poor solubility of quercetin and can be widely used in research on the extraction of quercetin from plants. It is also green, environmentally friendly, and suitable for industrial production.

[0005] To achieve the above objectives, the present invention provides the following technical solution:

[0006] In a first aspect, the present invention provides a method for improving the solubility of quercetin, comprising the following steps:

[0007] Quercetin was dissolved using an aqueous solution of choline chloride as a solvent.

[0008] Furthermore, the specific steps are as follows:

[0009] S1. Mix choline chloride with water and heat and stir until homogeneous to obtain an aqueous solution of choline chloride;

[0010] S2. Quercetin and choline chloride aqueous solution are mixed and heated and sonicated to obtain a high-concentration quercetin solution.

[0011] Choline chloride is a quaternary ammonium salt with good biodegradability and is relatively environmentally friendly. Currently, choline chloride is the most common hydrogen bond acceptor in the preparation of eutectic solvents (DESs).

[0012] Preferably, the concentration of the choline aqueous solution in S1 is 10-70%.

[0013] Further preferably, the concentration of the choline aqueous solution in S1 is 60-70%.

[0014] Preferably, the heating temperature in S1 is 80-100℃; and the stirring time is 10-12h.

[0015] Preferably, the heating temperature in S2 is 50-100℃; the ultrasonic time is 1-5h; and the ultrasonic power is 300-500w.

[0016] Further preferably, the heating temperature in S2 is 70°C.

[0017] Secondly, the present invention provides the application of the above-described method in the extraction of quercetin from plant raw materials.

[0018] As a preferred method, the extraction method of quercetin from plant raw materials is as follows:

[0019] Plant raw materials are mixed with a 50-70% aqueous solution of choline chloride and extracted by heating and stirring. The supernatant is then separated by centrifugation. Water is added to the supernatant to reduce the concentration of choline chloride to below 10% to precipitate crude quercetin. The crude quercetin is then purified with ethanol to obtain the finished quercetin product.

[0020] Preferably, the mass ratio of the plant material to the choline chloride aqueous solution is 1g:(4-10)ml.

[0021] Preferably, the heating and stirring temperature is 65-75℃; the stirring frequency is 150 r / min; and the stirring time is 0.5-2 h.

[0022] Preferably, the purity of the quercetin product is ≥93%.

[0023] It contains at least the following beneficial technical effects:

[0024] The method for improving quercetin solubility proposed in this invention is simple and easy to operate, and the solvent preparation process is simple and efficient. This method successfully solves the problem of poor quercetin solubility and can be widely used in research on the extraction of quercetin from plants, providing strong technical support for the industrial production of quercetin. Attached Figure Description

[0025] Figure 1 The graph shows the total energy, temperature, density, and potential energy parameters of quercetin in a 70% ChCl solution at simulated equilibrium.

[0026] Figure 2 These are snapshots of quercetin clusters at 5 ns in different solvents.

[0027] Figure 3 The hydrogen bond changes during the simulation of quercetin in different solvents. Detailed Implementation

[0028] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0029] It should be understood that the terminology used in this invention is merely for describing particular embodiments and is not intended to limit the invention. Furthermore, with respect to numerical ranges in this invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Every smaller range between any stated value or intermediate value within a stated range, and any other stated value or intermediate value within said range, is also included in this invention. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0030] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar or equivalent to those described herein may be used in the implementation or testing of this invention. All references to this specification are incorporated by way of citation to disclose and describe methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail.

[0031] Various modifications and variations can be made to the specific embodiments described in this specification without departing from the scope or spirit of the invention, as will be apparent to those skilled in the art. Other embodiments derived from this specification will also be obvious to those skilled in the art. This application specification and embodiments are merely exemplary.

[0032] The terms “include,” “including,” “have,” “contain,” etc., used in this article are all open-ended terms, meaning that they include but are not limited to.

[0033] Unless otherwise specified, "room temperature" and "normal temperature" in this invention refer to 25±2℃.

[0034] Unless otherwise specified, all raw materials or instruments used in the following embodiments of the present invention are commercially available.

[0035] Example 1

[0036] Prepare aqueous solutions of choline chloride at different concentrations:

[0037] Choline chloride was mixed with water and heated and stirred at 80°C for 10 hours to obtain an aqueous solution of choline chloride. 10 ml of aqueous solutions of choline chloride with concentrations of 10% wt, 20% wt, 30% wt, 40% wt, 50% wt, 60% wt, and 70% wt were prepared respectively.

[0038] Seven portions of excess quercetin solid solute were accurately weighed and slowly added to pre-prepared choline chloride aqueous solutions of different concentrations. The solutions were then sonicated at 70°C for 1 hour, followed by immediate centrifugation at 4000 r / min for 5 min. The supernatant was collected and diluted to the concentration range of the standard curve. The quercetin content was determined by HPLC analysis, and the solubility of quercetin at that temperature could be calculated (Table 1).

[0039] Detection results:

[0040] Determination of quercetin content in aqueous solutions of choline chloride at different mass concentrations in Example 1

[0041] (1) Establishment of the standard curve

[0042] Accurately weigh quercetin and dilute to 10 mL to obtain a standard stock solution with a quercetin concentration of 0.5 mg / mL. Take 0.2, 0.4, 0.6, 0.8, and 1 mL of the stock solution, dilute to 10 mL respectively, and filter through a 0.45 μm organic filter membrane to obtain five standards of different concentrations. The standards were determined by HPLC, with the following HPLC conditions: C18 reversed-phase column, mobile phase of acetonitrile-0.1% formic acid solution, flow rate of 1.0 mL / min, column temperature of 30℃, injection volume of 20 μL, and detection at a wavelength of 254 nm. A standard curve was established with mass (μg) as the abscissa (x) and peak area as the ordinate (y). Specifically, the quercetin standard curve is: y = 3729.6x - 6.1283.

[0043] (2) Determination of quercetin content

[0044] After diluting the supernatant obtained in Example 1 by 10 times, it was filtered through a 0.45 μm organic filter membrane and detected by HPLC in step (1). The peak area of ​​the detection result was substituted into the standard curve in step (1) to obtain the solubility of quercetin in this solution (Table 1).

[0045] Table 1. Solubility of quercetin in aqueous solutions of choline chloride at different mass concentrations (n ​​= 5, 70℃)

[0046] Choline chloride aqueous solution (%) Quercetin solubility (mg / ml) 10 0.46±0.01 20 0.52±0.02 30 0.74±0.05 40 1.33±0.01 50 2.22±0.01 60 12.29±0.85 70 58.34±0.81

[0047] As shown in Table 1, the solubility of quercetin gradually increases with the increase of the concentration of choline chloride aqueous solution. At 70%, the solubility of quercetin increases sharply. When it exceeds 70%, choline chloride and water are in a highly viscous state, which is not conducive to the dissolution of the solute.

[0048] Example 2

[0049] Solubility of quercetin at different temperatures

[0050] Accurately weigh 6 portions of excess quercetin solid solute and slowly add them to 10 ml of a pre-prepared 70% (mass ratio) choline chloride aqueous solution. Dissolve the solid by sonication (power 400w) for 1 hour (temperatures set at 50℃, 60℃, 70℃, 80℃, 90℃, and 100℃ respectively) to promote solid dissolution. After reaching dissolution equilibrium, immediately centrifuge at 4000 r / min for 5 min, collect the supernatant, dilute it to the concentration range of the standard curve, and determine the quercetin content by HPLC analysis. The solubility of quercetin at this temperature can then be calculated, as shown in Table 2.

[0051] Table 2. Solubility of quercetin in 70% choline chloride aqueous solution at different temperatures (n=5)

[0052] Temperature (°C) Quercetin solubility (mg / ml) 50 19.33±16.11 60 56.12±2.51 70 60.61±5.39 80 60.61±10.09 90 59.57±15.39 100 60.11±19.99

[0053] Comparative Example 1

[0054] Accurately weigh two portions of excess quercetin solid solute and slowly add them to 10 ml of pure ethanol, methanol, and acetone, respectively. Sonicate for 1 hour (at 70℃) to promote solid dissolution. Then immediately centrifuge at 4000 r / min for 5 min, collect the supernatant, dilute to the concentration range of the standard curve, and determine the quercetin content using HPLC analysis. The solubility of quercetin at this temperature can then be calculated, as shown in Table 3.

[0055] Table 3. Solubility of quercetin in common solvents (n = 5, 70℃)

[0056] solvent Anhydrous ethanol methanol acetone Quercetin solubility (mg / ml) 8.06±0.95 18.06±1.24 16.98±0.16

[0057] Example 3

[0058] Solubility experiment of quercetin under optimal conditions

[0059] Accurately weigh an excess of quercetin solid solute and slowly add it to 10 ml of a pre-prepared 70% (mass ratio) choline chloride aqueous solution. Sonicate for 2 hours (70℃) to promote solid dissolution. After reaching dissolution equilibrium, centrifuge at 4000 r / min for 5 min, collect the supernatant, dilute to the concentration range of the standard curve, and determine the quercetin content using HPLC analysis. The solubility of quercetin at this temperature can then be calculated to be 60.68 mg / ml.

[0060] Example 4

[0061] Same as Example 3, except that the aqueous solution of choline chloride was 60%. Analytical calculations showed that the solubility of quercetin was 13.26 mg / ml.

[0062] Example 5

[0063] Experiment on the extraction of quercetin from plant materials using choline chloride aqueous solution.

[0064] Weigh 20 g of the enzymatically hydrolyzed Sophora japonica bud residue (patent number 202311264631.X) and add it to an extraction tank. Add 100 mL of a prepared choline chloride aqueous solution (70%) to the extraction tank, set the heating temperature to 70℃, and extract at a constant temperature with stirring at 150 r / min for 2 h. During this process, the choline chloride aqueous solution fully dissolves quercetin in the Sophora japonica buds into the solution through osmosis and diffusion. After extraction, perform solid-liquid centrifugation at 4000 r / min for 15 min to separate the solid residue from the extract. Collect the supernatant to obtain a crude extract containing quercetin. Based on the difference in quercetin solubility in aqueous solutions of choline chloride of different concentrations, water was added to promote the mass concentration of choline chloride in the extract to 20%, resulting in the precipitation of a large amount of quercetin monomer. After centrifugation and drying, crude quercetin with a purity of 90.26% was obtained. The crude quercetin was then further processed with ethanol to obtain a finished quercetin product with a purity of up to 95%.

[0065] Example 6

[0066] Example of the mechanism by which choline chloride aqueous solution enhances the solubility of quercetin

[0067] This patent uses molecular dynamics simulations to investigate the microscopic dissolution behavior of quercetin monomers in aqueous solutions of choline chloride and ethanol at different concentrations, as well as their interactions with the solvents, revealing the mechanism by which choline chloride aqueous solution enhances the solubility of quercetin. The main focus of the study was the interaction between quercetin monomers and three solvents (aqueous solution of choline chloride, ethanol, and water), including microscopic dissolution behavior, the number of hydrogen bonds, the average lifetime of hydrogen bonds, and interaction energies. Figure 1The simulation parameters (total energy, temperature, density, and potential energy) for 70% choline chloride are shown to change over time, with the values ​​of total energy, temperature, density, and potential energy gradually becoming constant. These results indicate that the entire system reaches equilibrium after 5 ns of simulation.

[0068] Figure 2 These are snapshots of quercetin monomer clusters at 5 ns in different solvent systems. Figure 3 The hydrogen bond changes of quercetin during simulations in different solvents are shown (red: quercetin-quercetin; green: quercetin-choline / ethanol; blue: quercetin-water). The results show that the molecular configuration of quercetin after 5 ns equilibrium differs significantly in different solvent systems. In a pure water system, quercetin molecules rapidly aggregate in the initial stage of the simulation, eventually forming a stable multi-molecular cluster structure. When ethanol or choline chloride is introduced as a co-solvent, the dispersibility of quercetin exhibits a significant concentration dependence—when the co-solvent concentration is increased to 70%, the quercetin molecules in the system are completely monodisperse. This phenomenon is closely related to the unique "amphiphilic solvation" effect of quercetin: its hydrophobic flavonoid skeleton and hydrophilic rhamnose group together constitute an amphiphilic structure, allowing the solvent molecule to stabilize its aromatic ring structure through hydrophobic interactions and anchor the glycoside group through a hydrogen bond network. It is worth noting that in choline chloride solutions with high ionic strength, the ionic atmosphere formed by delocalized charges can effectively shield intermolecular dipole-dipole interactions, thereby inhibiting aggregation behavior and significantly promoting the dispersion and dissolution of quercetin monomers.

[0069] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A method for improving the solubility of quercetin, characterized in that, Includes the following steps: S1. Mix choline chloride with water and heat and stir until homogeneous to obtain an aqueous solution of choline chloride; S2. Quercetin and choline chloride aqueous solution are mixed and heated and sonicated to obtain a high-concentration quercetin solution; The concentration of the choline chloride aqueous solution in S1 is 70%; The heating temperature in S1 is 80-100℃; the stirring time is 10-12h; The heating temperature in S2 is 50-100℃; the ultrasonic time is 1-5h; and the ultrasonic power is 300-500w.

2. The application of the method of claim 1 in the extraction of quercetin from plant materials.

3. The application according to claim 2, characterized in that, The method for extracting quercetin from plant materials is as follows: Plant raw materials are mixed with a 70% choline chloride aqueous solution and extracted by heating and stirring. The supernatant is then separated by centrifugation. Water is added to the supernatant to reduce the concentration of choline chloride to below 10% to precipitate crude quercetin. The crude quercetin is then purified with ethanol to obtain the finished quercetin product.

4. The application according to claim 3, characterized in that, The ratio of the plant material to the choline chloride aqueous solution is 1g:(4-10)ml.

5. The application according to claim 3, characterized in that, The heating and stirring temperature is 65~75℃; the stirring frequency is 150r / min; and the stirring time is 0.5-2h.

6. The application according to claim 3, characterized in that, The purity of the quercetin product is ≥93%.

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