A food-grade o / w quercetin microemulsion, a preparation method and application thereof

Abstract

The application discloses a food-grade O / W quercetin microemulsion and a preparation method and application thereof, and each component and weight percentage are as follows: 10%-42% of a surfactant, 2%-32% of an oil phase solution, and the balance of water; wherein the oil phase solution is composed of isopropyl myristate and ethanol, and the weight ratio of isopropyl myristate to ethanol is 1:1-4. The O / W microemulsion preparation method is simple, is a good carrier of active substances or functional substances represented by quercetin which has poor stability and water solubility, so that the stability of the active substances or functional substances in pharmaceutical preparations is improved, and the sustained release behavior is improved.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical science, specifically relating to a food-grade O / W quercetin microemulsion, its preparation method, and its application. Background Technology

[0002] A microemulsion is a dispersion system consisting of two immiscible liquids, one of which is dispersed in the other as tiny droplets. Microemulsions offer many advantages, such as high bioavailability, high solubility, high stability, and low toxicity, and are therefore widely used in pharmaceuticals, cosmetics, and other fields.

[0003] Oil-in-water (O / W) microemulsions are microemulsions with oil as the dispersed phase and water as the continuous phase. They can effectively encapsulate bioactive substances, protecting them from external environmental influences and prolonging their action time. O / W microemulsions typically require the addition of hydrophilic emulsifiers to reduce the surface tension at the water-oil interface, promoting microemulsion formation and stability.

[0004] Quercetin is a naturally occurring flavonoid compound with various biological activities, including antioxidant, anti-inflammatory, antibacterial, and antitumor effects. However, its low solubility in water limits its application in pharmaceuticals and cosmetics. Therefore, using oil-in-water microemulsions as carriers for quercetin is an effective method to improve its solubility and bioavailability.

[0005] Currently, both domestically and internationally, the main methods for overcoming quercetin's susceptibility to oxidation and its insolubility in water involve modifying quercetin or preparing carriers to encapsulate it. For example, patent CN105534897A uses Sophora japonica as a raw material to prepare quercetin, modifies it, and then mixes it with a prepared blank microemulsion, followed by ultrasonic oscillation to obtain a highly water-soluble quercetin microemulsion. Ma Juanjuan et al. prepared quercetin composite colloidal particles and a stable Pickering emulsion delivery system based on the interaction between zein and polysaccharides, phospholipids, and polyphenols. This significantly improved the solubility and photothermal stability of quercetin. (Ma Juanjuan; Study on the Bioactivity and Bioavailability of Quercetin Delivery Carriers Constructed Based on Zein, Doctoral Dissertation, South China University of Technology, 2021, 4: 1-179.). However, these methods involve complex preparation processes and high costs. Summary of the Invention

[0006] Objective of the Invention: The technical problem to be solved by this invention is to address the shortcomings of existing technologies by providing a water-in-oil quercetin microemulsion and its preparation method. By using microemulsification technology to prepare the water-in-oil quercetin microemulsion, the problems of low solubility and poor bioavailability of quercetin are solved, and its stability and efficacy are improved. This emulsion exhibits good stability and sustained-release properties, and can effectively improve the solubility and bioavailability of quercetin.

[0007] To achieve the above objectives, the technical solution adopted by the present invention is as follows:

[0008] A food-grade O / W quercetin microemulsion comprises a blank microemulsion consisting of a surfactant, an oil phase solution, and an aqueous phase, and a quercetin microemulsion encapsulated in the blank microemulsion as a carrier; wherein the oil phase solution comprises oil and a co-surfactant.

[0009] The quercetin microemulsion described above has the following characteristics:

[0010] (1) Appearance: Transparent or translucent, colorless or pale yellow liquid;

[0011] (2) Average particle size: 200–300 nm;

[0012] (3) pH value: 6.0~8.0;

[0013] (4) Drug content: 0.4 mg / g to 1.51 mg / g;

[0014] (5) Drug release: The cumulative release rate of quercetin microemulsion within 24 hours is 25% to 47%.

[0015] Specifically, the quercetin mentioned is a flavonoid compound with the chemical name 3,3',4',5,7-pentahydroxyflavone.

[0016] Specifically, the surfactant is any one of emulsifier EL-35, Tween-80, and emulsifier OP, with emulsifier EL-35 being preferred. EL-35 is a nonionic surfactant, chemically known as polyoxyethylene-35 stearyl alcohol ether, which has the characteristics of low toxicity, low irritation, and low conductivity. It can effectively reduce the surface tension and repulsive forces between micelles in microemulsions, thereby improving the stability and solubilization capacity of microemulsions.

[0017] Specifically, the oil is selected from any one of peanut oil, isopropyl myristate, ethyl butyrate, and ethyl n-octanoate, with isopropyl myristate being preferred. Isopropyl myristate is a non-volatile or low-volatile oil with good biocompatibility and safety, effectively preventing loss and deterioration of the drug during storage and use due to oil volatilization or oxidation.

[0018] Furthermore, the oil phase solution further includes a co-surfactant, which is any one of anhydrous ethanol, polyethylene glycol 400, ethylene glycol, and isopropanol, with anhydrous ethanol being preferred. Anhydrous ethanol reduces interfacial tension, adjusts the polarity of water and oil, and regulates the HLB value. Anhydrous ethanol is a commonly used solubilizer and can increase the solubility of quercetin in the oil phase and the oil phase content in the microemulsion, thereby improving drug loading and release efficiency.

[0019] Preferably, based on weight, the components and their weight percentages are: 10%-42% surfactant, 2%-32% oil phase solution, and the balance being aqueous phase; the oil phase solution is composed of isopropyl myristate and ethanol, with a weight ratio of isopropyl myristate:ethanol = 1:1~4.

[0020] The optimal microemulsion formulation for the blank microemulsion is as follows: 30% surfactant, 20% oil phase solution, and the remainder is aqueous phase.

[0021] Preferably, the oil phase is composed of isopropyl myristate and ethanol in a weight ratio of isopropyl myristate (IPM): ethanol = 1:2.

[0022] Furthermore, the present invention also provides a method for preparing the above-mentioned food-grade O / W microemulsion, specifically comprising:

[0023] Under continuous stirring, the surfactant is added to the mixing vessel by weight percentage, followed by the oil phase solution, and finally the aqueous phase, until the liquid becomes clear and transparent.

[0024] Preferably, the stirring speed is 100 rpm to 500 rpm; the temperature is 50 to 60°C, preferably 55°C; and the pH value is controlled at 6 to 8, preferably 7.4.

[0025] Furthermore, the present invention also provides the application of the above-mentioned food-grade O / W quercetin microemulsion as a carrier in pharmaceutical manufacturing. Beneficial effects

[0026] (1) The quercetin microemulsion prepared in this application has good stability and sustained-release effect, and can be used in pharmaceuticals to increase the solubility and bioavailability of quercetin. The quercetin microemulsion prepared in this invention has a cumulative release rate of 25% to 47% of quercetin within 24 hours, indicating that the microemulsion has a good sustained-release effect, which can prolong the duration of action of the drug in vivo and improve the bioavailability of the drug.

[0027] (2) Compared with the prior art, the quercetin microemulsion prepared in this application has the following advantages:

[0028] High solubility and stability: Quercetin has low solubility in water, which limits its bioavailability and therapeutic efficacy. However, microemulsions can significantly improve the solubility of quercetin in water, making it easier for the body to absorb and utilize.

[0029] The O / W microemulsion preparation method described in this invention is simple and easy to implement; it only requires adding each component in the specified proportion and mixing them thoroughly.

[0030] The O / W microemulsions described in this invention have small particle size (<300nm) and uniform distribution, resulting in good stability of the microemulsion system.

[0031] In summary, the quercetin oil-in-water microemulsion synthesized in this invention has great potential in various application fields such as drug delivery and the cosmetics industry. Attached Figure Description

[0032] The present invention will be further described in detail below with reference to the accompanying drawings and specific embodiments, and the advantages of the present invention in the above and / or other aspects will become clearer.

[0033] Figure 1 This represents the linear relationship between quercetin concentration and ultraviolet absorption.

[0034] Figure 2 The solubility of quercetin in micelle solutions with different surfactants is given.

[0035] Figure 3 The solubility of quercetin in different oils.

[0036] Figure 4 Quercetin content in microemulsions at different temperatures.

[0037] Figure 5 Quercetin content in microemulsions at different pH levels.

[0038] Figure 6 The relationship between quercetin release rate and time in different samples. Detailed Implementation

[0039] The present invention will be described below with reference to specific embodiments. Those skilled in the art will understand that these embodiments are for illustrative purposes only and do not limit the scope of the invention in any way.

[0040] Unless otherwise specified, the experimental methods used in the following examples are conventional methods. Unless otherwise specified, all medicinal materials and reagents used in the following examples are commercially available products.

[0041] Experimental steps:

[0042] Accurately weigh 9.8 mg of quercetin sample and dissolve it in anhydrous ethanol to prepare anhydrous ethanol solutions with concentrations of 1.96, 3.92, 5.88, 7.84, 9.80, and 11.76 μg / mL. The ultraviolet absorption of each solution was measured at 370 nm, and the results are shown below. Figure 1 .

[0043] 1. Selection of surfactants

[0044] The absorbance of sample solutions containing quercetin in EL-35, Tween-80, and emulsifier OP was measured at 370 nm. The solubility of quercetin was calculated, and the results are shown in [Figure number missing]. Figure 2 .Depend on Figure 2 It is known that quercetin has the highest solubility in the emulsion micelle solution EL-35. Because EL-35 has high food safety, wide applicability, and strong emulsifying ability, it was chosen as the surfactant.

[0045] 2. Selection of oil phase

[0046] The solubility of quercetin in peanut oil, ethyl butyrate, isopropyl myristate, and ethyl n-octanoate sample solutions was determined at 370 nm. The results are shown below. Figure 3 .Depend on Figure 3 It is evident that quercetin has low solubility in the commonly used oils mentioned above, and oil solvents alone cannot meet the requirements for drug delivery. Therefore, isopropyl myristate was initially selected.

[0047] 3. Selection of co-surfactants

[0048] The functions of co-surfactants are threefold: first, to reduce interfacial tension; second, to increase interfacial fluidity; and third, to regulate the HLB value. This experiment tested anhydrous ethanol, polyethylene glycol 400, ethylene glycol, and isopropanol. Isopropanol is toxic, highly hydrophilic, and readily forms oil-in-water microemulsions. Long carbon chains and large molecular volumes of co-surfactants can increase resistance to membrane embedding. Considering factors such as short chains, non-toxicity, safety, and microemulsion size, different ratios of co-surfactant and oil were determined by testing the solubility of quercetin in different ratios. When the ratio of anhydrous ethanol to oil was 1:2, 1:1, and 2:1, the solubility was 17.4 mg / g, 23.5 mg / g, and 32.75 mg / g, respectively. When the ratio of polyethylene glycol 400 to oil was 1:2, 1:1, and 2:1, the solubility was 2.25 mg / g, 6.7 mg / g, and 10.7 mg / g, respectively. Therefore, in the experiment with maximum solubility, the ratio of anhydrous ethanol, polyethylene glycol 400, and oil was 2:1. Based on the analysis, the ratio of isopropyl myristate to anhydrous ethanol in the EL-35 / ethanol / isopropyl myristate / water system was determined to be 1:2.

[0049] 4. Temperature Selection

[0050] The prepared O / W microemulsion was mixed with an excess of quercetin and placed in a water bath at temperatures of 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, and 65°C. After magnetic stirring for one hour, the mixture was centrifuged and filtered to obtain the quercetin microemulsion. Quantitative analysis was performed using ultraviolet spectrophotometry to investigate the effect of the microemulsion on the solubilization of quercetin at different temperatures. Figure 4It can be seen that when the temperature increases from 10℃ to 25℃, the solubility of quercetin increases from 0.4 mg / g to 0.8 mg / g; when the temperature increases to 55℃, the solubility of quercetin increases to 0.151 mg / g; and as the temperature continues to rise, the solubility of quercetin decreases. Therefore, the solubility of quercetin is highest at 55℃, and thus 55℃ is the optimal treatment temperature.

[0051] 5. pH selection

[0052] O / W microemulsions were prepared with phosphate buffer and excess quercetin was added at 55°C. The effect of different pH values ​​on the solubilization of quercetin was observed.

[0053] according to Figure 5 It can be seen that below pH 7.4, as the pH increases from 4 to 7.4, the solubility of quercetin increases from 0.4 mg / g to 1.51 mg / g; when the pH increases from 7.4 to 10, the solubility of quercetin decreases from 1.51 mg / g to 0.87 mg / g. Therefore, the solubility of quercetin is highest at pH 7, so pH 7.4 is chosen as the optimal pH.

[0054] Example 1: An O / W Microemulsion

[0055] Raw material formula: (unit: g)

[0056] EL-35 30

[0057] Ethanol 13.5

[0058] Isopropyl myristate 6.5

[0059] Water 50

[0060] Prepared by the following method:

[0061] First, ethanol and isopropyl myristate are mixed to obtain an oil phase solution. Then, EL-35 is added to the mixing vessel while the magnetic stirrer is continuously stirring at 250 rpm. Finally, water is added until the mixture is clear and transparent, thus obtaining a microemulsion with an average particle size of 215 nm.

[0062] Example 2: An O / W Microemulsion

[0063] Raw material formula: (unit: kg)

[0064] EL-35 10

[0065] 1.8g of ethanol

[0066] Isopropyl myristate 0.2

[0067] Water 88

[0068] Prepared by the following method:

[0069] First, ethanol and isopropyl myristate are mixed to obtain an oil phase solution. Then, EL-35 is added to the mixing vessel while the magnetic stirrer is continuously stirring at 100 rpm. Finally, water is added until the mixture is clear and transparent, thus obtaining a microemulsion with an average particle size of 227 nm.

[0070] Example 3: An O / W Microemulsion

[0071] Raw material formula: (unit: g)

[0072] EL-35 42

[0073] 16% ethanol

[0074] Isopropyl myristate 16

[0075] Water 26

[0076] Prepared by the following method:

[0077] First, ethanol and isopropyl myristate are mixed to obtain an oil phase solution. Then, EL-35 is added to the mixing vessel while the magnetic stirrer is continuously stirring at 500 rpm. Finally, water is added until the mixture is clear and transparent, thus obtaining a microemulsion with an average particle size of 263 nm.

[0078] Example 4: A Quercetin O / W Microemulsion

[0079] Raw material formula: (unit: g)

[0080] Quercetin 0.151

[0081] EL-35 30

[0082] Ethanol 13.5

[0083] Isopropyl myristate 6.5

[0084] Water 50

[0085] Prepared by the following method:

[0086] First, ethanol and isopropyl myristate are mixed to obtain an oil phase solution. With continuous stirring on a magnetic stirrer at 250 rpm, EL-35 is added to a mixing vessel placed in a 55°C water bath, followed by the oil phase solution. Water is added to adjust the pH of the solution to 7.4, then excess quercetin is added, stirred until dissolved, and then centrifuged and filtered to obtain a clear, transparent liquid, which is the final product.

[0087] The obtained quercetin microemulsion has the following characteristics:

[0088] (1) Appearance: Transparent or translucent, colorless or pale yellow liquid;

[0089] (2) Average particle size: 232 nm;

[0090] (3) Drug release: such as Figure 6 As shown, the cumulative release rate of quercetin microemulsion within 24 hours was 47%, indicating that quercetin microemulsion has a good sustained-release effect.

[0091] Example 5: A Quercetin O / W Microemulsion

[0092] Raw material formula: (unit: g)

[0093] Quercetin 0.04

[0094] EL-35 10

[0095] 1.8g of ethanol

[0096] Isopropyl myristate 0.2

[0097] Water 88

[0098] Prepared by the following method:

[0099] First, ethanol and isopropyl myristate are mixed to obtain an oil phase solution. With continuous stirring at 100 rpm using a magnetic stirrer, EL-35 is added to a mixing vessel placed in a 10°C water bath, followed by the oil phase solution. Water is added to adjust the pH of the solution to 4, and then excess quercetin is added. The mixture is stirred until dissolved, then centrifuged and filtered to obtain a clear, transparent liquid, which is the final product.

[0100] The obtained quercetin microemulsion had an average particle size of 270 nm and a cumulative release rate of 25% over 24 hours.

[0101] Example 6: A Quercetin O / W Microemulsion

[0102] Raw material formula: (unit: g)

[0103] Quercetin 0.042

[0104] EL-35 42

[0105] 16% ethanol

[0106] Isopropyl myristate 16

[0107] Water 26

[0108] Prepared by the following method:

[0109] Ethanol and isopropyl myristate were first mixed to obtain an oil phase solution. Under continuous magnetic stirring at 250 rpm, EL-35 was added to a mixing vessel placed in a 65°C water bath, followed by the oil phase solution. Water was added to adjust the pH of the solution to 11, and then excess quercetin was added. The mixture was stirred until dissolved, then centrifuged and filtered to obtain a clear, transparent liquid. The average particle size of the obtained quercetin microemulsion was 294 nm. The cumulative release rate of the quercetin microemulsion within 24 hours was 32%, indicating that the quercetin microemulsion has a good sustained-release effect.

[0110] Experimental Example 1: Investigation of Microemulsion Performance

[0111] 1. Determination of microemulsion particle size

[0112] The particle size of the microemulsion samples prepared in Examples 1-6 was determined using a Nano-Zs90 Malvern laser scattering instrument at 25 °C, and the average particle size of the microemulsions was calculated. The results are shown in Table 1.

[0113] Table 1. Results of microemulsion particle size determination

[0114]

[0115] Table 1 shows that the O / W microemulsions prepared in Examples 1-6 all had particle sizes less than 300 nm. Moreover, compared with the corresponding blank microemulsions (microemulsions prepared in Examples 1-3), the particle size of the quercetin O / W microemulsions was slightly increased, while the change in dispersibility was not significantly different.

[0116] Test Example 2: Release test of the quercetin O / W microemulsion described in this invention.

[0117] Experimental method: The in vitro release effect of the quercetin microemulsion system was investigated by using a dialysis bag to simulate in vitro release.

[0118] A quercetin 1.51 mg / g oil phase solution was used as a reference. 5 mL of the quercetin microemulsion and the quercetin 1.51 mg / g oil phase solution prepared in Examples 4-6 were respectively placed in dialysis bags. The dialysis bags were then placed in Erlenmeyer flasks containing 50 mL of PBS and magnetically stirred at room temperature. At different time points, 5 mL of the dialysis solution was taken from each Erlenmeyer flask and replenished with the same volume of PBS. The quercetin content was determined by ultraviolet spectrophotometry.

[0119] The results of Example 4 are shown in Figure 6 .

[0120] By calculating the cumulative release rate, it can be seen that the release rates of quercetin oil phase solution at 3 h and 24 h are 72.35% and 100%, respectively, with complete release at 24 h. In contrast, the release rates of quercetin O / W microemulsion prepared in Example 4 are only 35.6% and 47% at 3 h and 24 h, respectively; the release rate then slows down, reaching only 58% at 60 h. Therefore, the above O / W microemulsion has a significant sustained-release effect on quercetin.

[0121] Test Example 3: Photostability Test of the Quercetin O / W Microemulsion of the Present Invention

[0122] Test method:

[0123] At 25°C, the quercetin O / W microemulsion and quercetin oil phase solution prepared in Examples 4-6 were placed in colorless open glass containers and exposed to sunlight outdoors for 110 days. The quercetin content was determined on days 1, 2, 3, and 110 using the method described above. Taking the quercetin content on day 0 as 100%, the relative percentage content of quercetin at other measurement points was calculated, and the results are shown in Table 2.

[0124] Table 2. Results of the stability study of quercetin O / W microemulsion

[0125]

[0126] As can be seen from Table 2, the quercetin O / W microemulsion described in this invention can significantly improve the stability of quercetin. Furthermore, the relative quercetin content in the quercetin O / W microemulsion prepared in Example 4 is consistently higher than that in the quercetin O / W microemulsions prepared in Examples 5 and 6; indicating that the O / W microemulsion in Example 4 has a better encapsulation effect on quercetin.

[0127] This invention provides a food-grade O / W quercetin microemulsion, its preparation method, and its application. Many methods and approaches exist for implementing this technical solution; the above description is merely a preferred embodiment of the invention. It should be noted that those skilled in the art can make various improvements and modifications without departing from the principles of this invention, and these improvements and modifications should also be considered within the scope of protection of this invention. All components not explicitly stated in this embodiment can be implemented using existing technologies.

Claims

1. A food-grade O / W quercetin microemulsion, characterized in that, The raw material formula is as follows: Quercetin 0.151g, EL-35 30g 13.5g of ethanol Isopropyl myristate 6.5g 50g of water Prepared by the following method: First, ethanol and isopropyl myristate are mixed to obtain an oil phase solution. Under continuous stirring with a magnetic stirrer at a speed of 250 rpm, EL-35 is added to a mixing vessel placed in a 55°C water bath, followed by the oil phase solution. Water is added to adjust the pH of the solution to 7.

4. Then, excess quercetin is added, stirred to dissolve, and then centrifuged and filtered to obtain a clear and transparent liquid, which is the product. The obtained quercetin microemulsion has the following characteristics: (1) Appearance: Transparent or translucent, colorless or pale yellow liquid; (2) Average particle size: 232 nm; (3) Drug release: The cumulative release rate of quercetin microemulsion within 24 hours was 47%.

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