Method for preparing kgf2 nanoemulsion gel containing safflower seed oil

Abstract

The application discloses a preparation method of KGF2 nanoemulsion gel containing safflower seed oil, and specifically comprises the following steps: fusing KGF2 and safflower seed oil to obtain an oil phase; then preparing KGF2 nanoemulsion containing safflower seed oil by using the prepared oil phase; and finally preparing KGF2 nanoemulsion gel containing safflower seed oil by using the KGF2 nanoemulsion containing safflower seed oil.

Description

Technical Field

[0001] This patent belongs to the field of pharmaceutical technology, specifically relating to a KGF2 nanoemulsion gel containing safflower seed oil and its preparation method. Background Technology

[0002] The skin is the main barrier protecting the body's internal tissues from damage by harmful external substances. A wound is damage to normal skin (tissue) caused by external injury factors such as surgery, external force, and internal factors such as impaired local blood supply. Large skin defects, various pathological conditions, or common diseases such as infection, vascular disease, and cancer can all lead to delayed or non-healing of skin wounds. Prolonged or non-healing wounds not only seriously affect the patient's physical and mental health and quality of life but also impose a heavy economic burden on the patient's family and society. Therefore, effectively controlling the progression of the disease and improving the quality of wound healing has become one of the key issues in clinical research.

[0003] KGF2, or keratinocyte growth factor-2, is a member of the FGFs superfamily, also known as fibroblast growth factor-10 (FGF-10). It is a biologically active, naturally occurring basic protein in the human body, first isolated and purified from human lungs by Emoto et al. in 1997. KGF2 is an important regulator in skin tissue formation, effectively promoting the proliferation, metastasis, and differentiation of epithelial cells. It plays a crucial role in the repair of injuries such as acute and chronic inflammation, corneal damage, and burns, and its effects are particularly significant on epithelial tissue damage caused by chemotherapy, such as venous ulcers and ulcerative colitis. Currently, KGF2 is used in small quantities in topical formulations.

[0004] Safflower seed oil, also known as safflower oil, is an oil extracted from safflower seeds and is yellow in color. Its main components are linoleic acid, palmitic acid, oleic acid, stearic acid, and palmitic acid, in addition to natural vitamin E, flavonoids, and other active substances. Therefore, safflower seed oil has antioxidant, anti-inflammatory, anti-aging, and skin barrier repair effects. Currently, safflower seed oil is mainly used in the food and health care industries.

[0005] Currently, although products containing KGF2 are available on the market, such as Shigelos Repair Cream and Bocasi Mask, there are relatively few formulated preparations that utilize KGF2 to treat skin trauma. Furthermore, when KGF2 is applied topically for skin wound repair, its poor permeability prevents it from effectively exerting its function when applied directly. Safflower seed oil, on the other hand, often feels greasy when applied topically.

[0006] KGF2 and safflower seed oil have seen some application in wound repair, but several issues remain to be addressed. First, KGF2 has poor permeability when applied topically to wounds, hindering its effectiveness. Second, safflower seed oil has a greasy feel when applied topically, leading to lower patient compliance. Furthermore, current topical gels often prioritize good formability while neglecting to prepare them to a pH level suitable for the skin wound.

[0007] Therefore, the applicant proposed a method for preparing KGF2 nanoemulsion containing safflower seed oil. Summary of the Invention

[0008] The purpose of this invention is to overcome the problems of poor permeability of KGF2 when applied topically to wounds and the greasy feeling of safflower seed oil when applied topically. This invention provides a composite dosage form combining nanoemulsion and gel, which can increase the transdermal absorption and stability of KGF2, eliminate the greasy feeling of safflower seed oil when applied topically, ensure good formability and pH compatibility with the wound, enhance the efficiency of promoting skin wound healing, increase compliance during topical application, and enable KGF2 to exert a better effect when applied topically.

[0009] The present invention achieves the above objectives through the following technical solutions: The present invention provides a method for preparing KGF2 nanoemulsion gel containing safflower seed oil, the specific steps of which are: fusing KGF2 with safflower seed oil, preparing KGF2 nanoemulsion containing safflower seed oil, and preparing KGF2 nanoemulsion gel containing safflower seed oil.

[0010] The specific steps are as follows: A. Method for fusing KGF2 and safflower seed oil: KGF2 and safflower seed oil are added to the safflower seed oil at a ratio of 1mg:1g. The mixture is then ultrasonically fused at 700w for 10min. After stirring, the mixture is stirred thoroughly to ensure that KGF2 is uniformly fused into the safflower seed oil, thus forming an oil phase containing KGF2.

[0011] B. Preparation of KGF2 nanoemulsion containing safflower seed oil: The formulation consists of 0.3g (3%) of safflower seed oil containing KGF2 (oil phase), 0.6g (6%) of mixed emulsifiers polyoxyethylene castor oil and Tween-80 (wherein the emulsifiers polyoxyethylene castor oil and Tween-80 are mixed in a 1:1 mass ratio), 0.3g (3%) of co-emulsifier polyethylene glycol-400 (PEG-400), and 50ml (88%) of aqueous phase. During preparation, the oil phase is added to the thoroughly mixed emulsifiers and co-emulsifiers under stirring. After thorough mixing, the aqueous phase is added, and the mixture is homogenized in a high-speed homogenizer at 2500r / min for 50min to obtain the primary emulsion. The primary emulsion is then subjected to ultrasonic cell disruption at 40% rated power for 30min to obtain the KGF2 nanoemulsion containing safflower seed oil.

[0012] C. Preparation of gel matrix: The gel matrix formula is 0.8g (1.5%) carbomer-980, 10g (15%) glycerol and 50ml (83.5%) water. After adding glycerol to carbomer-980 and stirring appropriately, add water and stir thoroughly. Let it stand until it is completely swollen to obtain the gel matrix.

[0013] D. Preparation of KGF2 nanoemulsion gel containing safflower seed oil: The KGF2 nanoemulsion containing safflower seed oil prepared above was added to the swollen gel matrix under stirring. Triethanolamine was added to adjust the pH to 5.5-6.0, and then the mixture was stirred thoroughly until solidification was achieved to obtain KGF2 nanoemulsion gel containing safflower seed oil.

[0014] The beneficial effects of this invention are that it combines the pharmacological effects of KGF2 on skin and hair follicle repair with the skin barrier repair and anti-inflammatory effects of safflower seed oil, and uses a method to prepare both into a nanoemulsion. This solves the problem of poor permeability of KGF2 when used externally for skin wound repair, and the problem of greasiness associated with external application of safflower seed oil. Furthermore, by combining the nanoemulsion with a gel, the stability of the drug and the patient's compliance with medication are improved.

[0015] The nanoemulsions in this invention have small particle sizes, good stability, and excellent transdermal permeability. The resulting nanoemulsion gel has a pH that is compatible with the skin wound and good moisturizing properties, making the wound easier to absorb medication and heal. Furthermore, the resulting nanoemulsion gel has good formability, a fine and uniform appearance, is easy to spread, refreshing, non-greasy, and non-irritating when applied, resulting in good medication adherence.

[0016] The beneficial effect of this invention is that it provides a new approach for the preparation of KGF2 nanoemulsion gel containing safflower seed oil, which offers a novel method for the external application of KGF2 in skin wound repair and its combined medicinal use with safflower seed oil.

[0017] (1) By combining KGF2 with safflower seed oil, the skin barrier repair and anti-inflammatory effects of safflower seed oil and the pharmacological effects of KGF2 on skin and hair follicle repair are synergistically enhanced, thus improving the efficacy of promoting wound healing. (2) By combining nanoemulsions with gels, the problems of poor transdermal permeability of KGF2 and the greasy feeling when applying safflower seed oil can be solved, giving full play to the good transdermal permeability and drug protection of nanoemulsions and the pH wound suitability and external protection of pH-sensitive gels.

[0018] Furthermore, this invention utilizes an ultrasonic fusion method to fuse KGF2 with safflower seed oil, making it an integral part of the nanoemulsion formulation. This nanoemulsion is then converted using a homogenized primary emulsion method and an ultrasonic fine emulsification method. Subsequently, using a pH adjustment method, the nanoemulsion is added to pre-swollen carbomer-980, adjusted to a suitable pH for the wound, and then mechanically stirred to solidify it.

[0019] A method for preparing a combination of KGF2 and safflower seed oil using modern pharmaceutical techniques, and for converting it into a nanoemulsion and gel formulation, was developed. The resulting KGF2 nanoemulsion gel containing safflower seed oil was then used as a molded formulation for direct clinical applications such as promoting skin wound repair.

[0020] By utilizing modern pharmaceutical formulation technology to fuse safflower seed oil with KGF2, the stability of KGF2 is increased, preventing its denaturation during dosage form preparation, and a nanoemulsion is prepared, thus solving the problem of poor transdermal permeability of topical KGF2 and increasing overall stability.

[0021] By using a pH-sensitive gel matrix, the difficulty of preparing nanoemulsions and gels together is reduced, allowing the nanoemulsion gel to maintain a pH that is compatible with the wound, solving the greasy feeling of safflower seed oil for external use, and increasing medication compliance.

[0022] This patent, based on modern pharmaceutical formulation technology, fuses KGF2 with safflower seed oil to synergize their medicinal effects. It also combines nanoemulsion and gel, resulting in a relatively simple preparation process. The resulting KGF2 nanoemulsion gel containing safflower seed oil, when used topically to repair skin wounds, not only exerts the skin barrier repair and anti-inflammatory effects of safflower seed oil but also the pharmacological effects of KGF2 on skin and hair follicle repair. The combined use of nanoemulsion and gel improves the stability and transdermal absorption of KGF2, reduces the greasy feeling of safflower seed oil during topical application, prevents bacterial infection, increases compliance during application, maintains relative humidity, and promotes rapid wound healing.

[0023] KGF2 was incorporated into seed oil using modern pharmaceutical formulation technology to prepare an oil-in-water KGF2 nanoemulsion containing safflower seed oil; and KGF2 nanoemulsion gel containing safflower seed oil was prepared by combining two different dosage forms, nanoemulsion and gel, for clinical treatment such as skin wound repair. Attached Figure Description

[0024] Figure 1 Photograph of the appearance of KGF2 nanoemulsion containing safflower seed oil.

[0025] Figure 2 The particle size distribution of KGF2 nanoemulsion containing safflower seed oil is shown.

[0026] Figure 3 This is a potential diagram of KGF2 nanoemulsion containing safflower seed oil.

[0027] Figure 4 Photographs showing the physical stability of KGF2 nanoemulsion containing safflower seed oil.

[0028] Figure 5 Photograph of the appearance of KGF2 nanoemulsion gel containing safflower seed oil.

[0029] Figure 6 Photographs showing the physical stability of KGF2 nanoemulsion gel containing safflower seed oil.

[0030] Figure 7 The changes in the healing area of ​​mechanical trauma wounds in each group of SD rats.

[0031] Figure 8 A statistical chart showing the healing area of ​​wounds in each group. Detailed Implementation

[0032] The following embodiments further illustrate the above-described content of the present invention in detail. Obviously, the described embodiments are merely a part of the embodiments of the present invention, and not all of them. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative effort are within the scope of protection of the present invention.

[0033] Example 1: This example discloses a method for preparing KGF2 nanoemulsion gel containing safflower seed oil, specifically including the following steps: Step A: Preparation of the oil phase of the mixture of KGF2 and safflower seed oil: KGF2 and safflower seed oil were added to the safflower seed oil at a ratio of 1 mg: 1 g while stirring. After ultrasonic fusion at 700 W for 10 min, the mixture was stirred thoroughly to ensure that KGF2 was uniformly fused into the safflower seed oil, thus forming safflower seed oil containing KGF2 as the oil phase. Step B: Preparation of KGF2 nanoemulsion containing safflower seed oil. The preparation process is as follows: Weigh 0.3g of polyoxyethylene castor oil and place it in a 50℃ oven for 2-3 minutes to melt it into a liquid. Then, while stirring, add 0.3g of Tween-80, 0.3g of PEG-400, 0.3g of KGF2-infused seed oil, and 50ml of distilled water in sequence. Stir thoroughly and homogenize using a high-speed homogenizer at 2500r / min for 50 minutes to obtain the colostrum. Use an ultrasonic cell disruptor to sonicate the colostrum at 40% of its rated power for 30 minutes to obtain KGF2 nanoemulsion A containing safflower seed oil.

[0034] The appearance of the prepared KGF2 nanoemulsion containing safflower seed oil was observed, referring to... Figure 1 (a) It can be seen that the KGF2 nanoemulsion containing safflower seed oil is light milky white and slightly transparent, according to reference. Figure 1 (b) It can be seen that there is a clear Tyndall effect.

[0035] Physical particle size determination of KGF2 nanoemulsion containing safflower seed oil: The experimental method is as follows: 1 mL of nanoemulsion was taken into a Malvern quartz cuvette, and the cuvette was placed in a laser particle size Eeta potential and absolute molecular weight analyzer for particle size detection.

[0036] The experimental results are as follows: The results of the laser particle size Eeta potential and absolute molecular weight analyzer for measuring the nanoemulsion particle size are as follows: Figure 2 As shown in the figure, the size range of the nanoemulsion is 20–100 nm, with an average size of approximately 33.07 nm and a PDI of 0.196.

[0037] Potential detection of KGF2 nanoemulsion containing safflower seed oil: The experimental method is as follows: 1 mL of nanoemulsion was pipetted into a Malvern potential sample cell, and the Malvern potential sample cell was placed in a laser particle size Eeta potential and absolute molecular weight analyzer for potential program detection.

[0038] The experimental results are as follows: Laser particle size Eeta potential and absolute molecular weight analyzer results for nanoemulsion potential testing are as follows: Figure 3 As shown in the figure, the potential range of the nanoemulsion is -30 to 0 mV, with an average potential of approximately -16.7 mV.

[0039] Physical stability of KGF2 nanoemulsion containing safflower seed oil: The experimental method is as follows: The physical stability was investigated by high-speed centrifugation. Three nanoemulsions were prepared and placed at room temperature and pressure for 24h, 48h and 72h respectively. After centrifugation at 3000r / min for 10min, the nanoemulsions were observed to see if they showed stratification, precipitation or turbidity.

[0040] The experimental results are as follows: Figure 4 It can be seen that after standing for 24h, 48h, and 72h, no layering, precipitation, or turbidity was observed, indicating that the KGF2 nanoemulsion containing safflower seed oil has good physical stability.

[0041] Step C: Preparation of KGF2 nanoemulsion gel containing safflower seed oil. The preparation process is as follows: Add 10g of glycerol to 0.8g of carbomer-980 and stir appropriately. Then add 50ml of distilled water and stir thoroughly. Let it stand for 24 hours until fully swollen. Add the KGF2 nanoemulsion containing safflower seed oil prepared in step B to the swollen gel matrix while stirring. Add triethanolamine to adjust the pH to 5.5, stir thoroughly, and centrifuge at 3000r / min for 10min to remove air bubbles to obtain the KGF2 nanoemulsion gel containing safflower seed oil.

[0042] Observation of the morphology of the prepared KGF2 nanoemulsion gel containing safflower seed oil: Depend on Figure 5 (b) KGF2 nanoemulsion gel containing safflower seed oil is transparent, slightly milky white, uniform and delicate, has good formability, moderate viscosity, and is non-irritating, refreshing and has good spreadability when applied.

[0043] Physical stability of KGF2 nanoemulsion gel containing safflower seed oil was investigated. The experimental method is as follows: The physical stability was investigated by high-speed centrifugation. Three KGF2 nanoemulsion gels containing safflower seed oil were prepared and placed at room temperature and pressure for 24h, 48h and 72h respectively. After centrifugation at 3000r / min for 10min, the KGF2 nanoemulsion gels containing safflower seed oil were observed to see whether there was layering, precipitation or turbidity.

[0044] The experimental results are as follows: Depend on Figure 6 It can be seen that after standing for 24h, 48h, and 72h, no layering, precipitation, or turbidity was observed, indicating that the KGF2 nanoemulsion containing safflower seed oil has good physical stability.

[0045] Effects of KGF2 nanoemulsion gel containing safflower seed oil on the repair of mechanical wounds in SD rats: The experimental method is as follows: (1) Modeling of mechanical wounds on the skin After fasting for 12 hours, SD rats were anesthetized by intraperitoneal injection of 10% chloral hydrate at a dose of 0.3 ml / 100 g. Then, a circular piece of skin with a diameter of 1 cm was cut from the back of the rat using a punch and scissors, with the standard depth reaching the deep fascia of the muscle layer, to create a skin wound model for SD rats.

[0046] (2) Applying medicine to the wound The wounds in the model group were treated with physiological saline. The positive drug group was divided into two groups and treated with safflower seed oil and KGF2 solution respectively. The gel group was treated with nanoemulsion gel prepared in Example 7, which was divided into high, medium and low groups.

[0047] The experimental results are as follows: The wound area healing effect of the KGF2 nanoemulsion gel containing safflower seed oil prepared in this invention on wound repair in SD rats, and the statistical results of the wound area data are shown in the figure below. Figure 7 and Figure 8 As shown.

[0048] Figure 7 The results showed that KGF2 nanoemulsion gel containing safflower seed oil could effectively accelerate wound healing time. 14 days after the operation, the skin wounds of SD rats in the gel group were basically completely healed, but there were still obvious wounds in the model group, and the healing was better than that of the two positive drug groups. Figure 8 Compared to the model group, the gel group showed more significant changes in wound healing. In addition, compared with the two positive drug groups, the gel group had a more stable high healing rate, indicating that the nanoemulsion gel has a good ability to promote wound healing.

[0049] Those skilled in the art can make various modifications and variations to this invention without departing from its spirit and scope. Therefore, if these modifications and variations fall within the scope of the claims and their equivalents, this invention is also intended to include these modifications and variations.

Claims

1. A method for preparing KGF2 nanoemulsion gel containing safflower seed oil, characterized in that... Includes the following steps: A. Preparation of the oil phase of KGF2 and safflower seed oil mixture: KGF2 and safflower seed oil were added to the safflower seed oil at a ratio of 1 mg: 1 g while stirring. After ultrasonic fusion at 700 W for 10 min, the mixture was stirred thoroughly to ensure that KGF2 was uniformly fused into the safflower seed oil, thus forming safflower seed oil containing KGF2 as the oil phase. B. Preparation of KGF2 nanoemulsion containing safflower seed oil: Raw material selection: 0.3g of the oil phase of safflower seed oil containing KGF2 obtained in step A, 0.6g of mixed emulsifiers polyoxyethylene castor oil and Tween-80, 0.3g of co-emulsifier polyethylene glycol-400 (PEG-400) and 50ml of water, wherein the emulsifiers polyoxyethylene castor oil and Tween-80 are mixed at a mass ratio of 1:1; The preparation steps are as follows: The safflower seed oil phase containing KGF2 is added to the well-mixed emulsifier and co-emulsifier under stirring. After being stirred evenly, the aqueous phase is added and homogenized in a high-speed homogenizer at 2500 r / min for 50 min to obtain the primary emulsion. The primary emulsion is then subjected to ultrasonic cell disruption at 40% rated power for 30 min to obtain KGF2 nanoemulsion containing safflower seed oil. C. Preparation of gel matrix: The raw materials selected are 0.8g of carbomer-980, 10g of glycerol and 50ml of water; after adding glycerol to carbomer-980 and stirring appropriately, water is added and stirred thoroughly, and left to swell completely to obtain the gel matrix; D. Preparation of KGF2 nanoemulsion gel containing safflower seed oil: The KGF2 nanoemulsion containing safflower seed oil prepared in step B is added to the gel matrix prepared in step C under stirring. Triethanolamine is added to adjust the pH to 5.5-6.0, and then the mixture is stirred thoroughly until solidification is achieved to obtain KGF2 nanoemulsion gel containing safflower seed oil.

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