Guizhou buckthorn skin wound healing ointment and preparation method thereof

Abstract

The application provides a Guizhou Rhamnus skin wound healing ointment and a preparation method thereof, and relates to the technical field of traditional Chinese medicine compositions. The ointment comprises Guizhou Rhamnus concentrated extract and base excipients. The ointment has the functions of promoting skin wound healing, preventing swelling and pain, preventing infection and scar, reducing swelling and pain, removing blood stasis and promoting tissue regeneration, dredging channels and activating collaterals, and healthy skin tissue. The active ingredients of traditional Chinese medicine have the advantages of rapid healing and repair, obvious swelling and pain relief, delicate ointment texture, easy absorption by the skin, high safety, good stability, wide source of raw materials, easy collection, simple preparation process, batch processing, no toxic side effects on the health of production personnel, and no adverse effects on the environment.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of traditional Chinese medicine composition, and particularly relates to a Guizhou Rhamnus hancei skin wound healing ointment and a preparation method thereof. BACKGROUND

[0002] Wound healing generally refers to the repair process of skin and other body tissues after being injured by external force, and the wound healing can be generally divided into four stages, namely, coagulation stage, inflammation stage, repair stage and maturation stage. The four stages are both distinguished and related to each other. The coagulation stage: after the wound is formed, the first reaction of the body is to start the coagulation mechanism to perform the self-hemostasis process. The platelets on the wound surface gather and form a blood clot, which can effectively stop the bleeding of the wound; the inflammation stage: this stage is mainly to destroy bacteria and remove necrotic tissues to lay a foundation for tissue regeneration and repair. The inflammation stage lasts about 4-6 days, and the initial stage can be accompanied by redness, swelling, heat and pain; the repair stage: mainly tissue proliferation and granulation formation. New cells appear soon after the inflammation starts, and fibroblasts, endothelial cells and new capillaries proliferate and differentiate to form granulation tissue, which fills and covers the wound to form a scar. The proliferation period after different injuries is different, and mostly lasts for 4-24 days; the maturation stage: mainly the process of scar remodeling. After the proliferation stage, the wound has been preliminarily healed. With the passage of time, the scar tissue and scab repairing the wound gradually adjust to adapt to the physiological function, and finally the appearance and function of the injured site are improved. The time required for scar remodeling is different according to the severity of the wound, and generally lasts for 21 days to 1 year.

[0003] Wound healing is a continuous process, and attention should be paid to nutrition, prevention of infection, maintenance of clean and dry wound, and less spicy and stimulating food to avoid affecting wound healing. However, the wound treatment agent in the prior art has poor treatment effect, obvious side effects, easy infection, slow healing and easy scarring. Therefore, how to provide a wound treatment agent with good treatment effect, no obvious side effects, short healing time, no easy infection and no easy scarring is a technical problem to be solved in the field. SUMMARY

[0004] To solve the above problems, the present application provides a Guizhou Rhamnus hancei skin wound healing ointment, which comprises 40 parts by weight of Guizhou Rhamnus hancei concentrated extract and 185 parts by weight of matrix adjuvant.

[0005] The matrix adjuvant comprises the following raw materials in parts by weight: 100 parts of propylene glycol, 30 parts of polyisobutylene, 20 parts of sodium dodecyl sulfate, 20 parts of lanolin, and 15 parts of gypsum powder.

[0006] Further, the preparation method of the Guizhou buckthorn concentrated extract includes the following steps: mixing Guizhou buckthorn powder with rapeseed oil and stirring uniformly, placing at 4℃ for 12h, then adding double distilled water for soaking, after soaking, heating the mixed solution in a water bath, after heating, ultrasonic treatment and filtration, repeating extraction of the filter residue twice, and reserving the combined filtrate; the filtrate is concentrated into a flow extract with a viscosity of 10-25Pa.s by rotary evaporation, and the concentrate is autoclaved for 30min to obtain the Guizhou buckthorn concentrated extract.

[0007] Further, the preparation method of the Guizhou buckthorn powder is as follows: collecting clean Guizhou buckthorn branches, drying at 55℃ until constant weight, crushing and sieving through a 200-mesh screen to obtain Guizhou buckthorn powder.

[0008] Further, the ratio of Guizhou buckthorn powder to rapeseed oil in step (1) is 1-2g / mL.

[0009] Further, the ratio of Guizhou buckthorn powder to double distilled water in step (1) is 1g / 10-20mL.

[0010] Further, the soaking time in step (1) is 3-4h, and stirring is performed every 30min during the soaking process.

[0011] Further, the water bath heating temperature in step (2) is 55-70℃, and the water bath heating time is 30-60min.

[0012] Further, the ultrasonic treatment frequency in step (2) is 25-30KHz, and the ultrasonic treatment time is 30min.

[0013] The application also provides a preparation method of the Guizhou buckthorn skin wound healing ointment patch, which specifically includes the following steps:

[0014] (1) accurately weighing lanolin, sodium dodecyl sulfate, gesso powder and polyisobutylene by weight parts, mixing and stirring the above substances uniformly, adding propylene glycol and stirring uniformly, and heating in a 75℃ water bath until the base melts under uniform stirring;

[0015] (2) adding Guizhou buckthorn concentrated extract to the melted base, mixing and stirring the raw materials uniformly, and obtaining a patch ointment solution after sterilization;

[0016] (3) placing the patch ointment solution in the circle in the middle of the patch cloth, and naturally cooling into a solid at room temperature to obtain an external use skin wound healing ointment patch.

[0017] Further, the amount of patch ointment solution in the ointment patch is 3mL / patch.

[0018] Compared with the prior art, the application has the following beneficial technical effects:

[0019] (1) The ointment of the present application has the functions of promoting skin wound healing, preventing swelling and pain, preventing infection and scar, reducing swelling and pain, removing blood stasis and promoting skin tissue healing, etc. The active ingredients of traditional Chinese medicine in the ointment have the functions of rapid healing and repair, obvious swelling and pain relief, and the ointment has fine texture and is easy to be absorbed by the skin.

[0020] (2) The effective components in the ointment have high safety and good effect on the human body, and have good stability.

[0021] (3) The raw materials of the present application are widely available, easy to collect, and the preparation process is simple, easy to mass produce, non-toxic and harmless to the health of the production personnel, and has no adverse effects on the environment. BRIEF DESCRIPTION OF DRAWINGS

[0022] The present application will be further described below in conjunction with the drawings.

[0023] Figure 1 The recovery of the wound of the rabbit at different times;

[0024] Figure 2 The histopathological changes of the granulation tissue of the New Zealand rabbits in each group at different time points (HE staining x 100);

[0025] Figure 3 The comparison of the microvessel counts of the wounds at different time points in each group;

[0026] Figure 3 Compared with the physiological saline group, extremely significant is indicated by "**" (P<0.01), significant is indicated by "*" (P<0.05), and no statistical difference is indicated by "ns" (P>0.05). DETAILED DESCRIPTION

[0027] The present application provides a Guizhou Rhamnus skin wound healing ointment, which comprises 40 parts by weight of Guizhou Rhamnus concentrated extract and 185 parts by weight of matrix adjuvant.

[0028] The matrix adjuvant comprises the following raw materials in parts by weight: propylene glycol 100 parts, polyisobutylene 30 parts, sodium dodecyl sulfate 20 parts, lanolin 20 parts, and gypsum powder 15 parts.

[0029] In one embodiment, the preparation method of the Guizhou Rhamnus concentrated extract comprises the following steps: mixing Guizhou Rhamnus powder with rapeseed oil and stirring uniformly, placing at 4℃ for 12h, then adding double distilled water for soaking, after soaking, heating the mixed solution in a water bath, after heating, ultrasonic treatment and filtration, repeating the extraction of the filtrate twice, and reserving the combined filtrate; rotary evaporation of the filtrate to concentrate into a flow extract with a viscosity of 10-25Pa.s, and high-pressure sterilization of the concentrate for 30min to obtain the Guizhou Rhamnus concentrated extract.

[0030] In one embodiment, the preparation method of the Guizhou buckthorn powder is as follows: collecting clean Guizhou buckthorn branches, drying at 55℃ until constant weight, crushing and sieving through a 200-mesh screen to obtain Guizhou buckthorn powder.

[0031] In one embodiment, the ratio of Guizhou buckthorn powder to rapeseed oil in step (1) is 1-2 g / mL.

[0032] In one embodiment, the ratio of Guizhou buckthorn powder to double distilled water in step (1) is 1 g / 10-20 mL.

[0033] In one embodiment, the soaking time in step (1) is 3-4 h, and the stirring is performed every 30 min during the soaking process.

[0034] In one embodiment, the water bath heating temperature in step (2) is 55-70℃, and the water bath heating time is 30-60 min.

[0035] In one embodiment, the ultrasonic treatment frequency in step (2) is 25-30 KHz, and the ultrasonic treatment time is 30 min.

[0036] The application also provides a preparation method of the Guizhou buckthorn skin wound healing ointment, which specifically comprises the following steps:

[0037] (1) Accurately weighing lanolin, sodium dodecyl sulfate, gesso powder and polyisobutylene by weight parts, mixing and stirring the above substances uniformly, adding propylene glycol and stirring uniformly, and heating in a 75℃ water bath until the base melts under uniform stirring conditions;

[0038] (2) Adding Guizhou buckthorn concentrated extract to the melted base, mixing and stirring the raw materials uniformly, and obtaining a plaster solution after sterilization;

[0039] (3) Placing the plaster solution in the circle in the middle of the plaster, and naturally cooling to solid at room temperature to obtain an external use skin wound healing ointment plaster.

[0040] In one embodiment, the amount of plaster solution in the ointment plaster is 3 mL / plaster.

[0041] The technical solutions provided by the application are further described below in combination with examples.

[0042] Example 1

[0043] (1) Preparation of concentrated extract:

[0044] The clean Guizhou buckthorn branches were collected and dried at 55°C until constant weight. The Guizhou buckthorn powder was obtained by crushing and sieving through a 200-mesh sieve. Then, 100 g of the Guizhou buckthorn powder was weighed into a beaker and mixed with 50 mL of rapeseed oil. The mixture was placed in a 4°C refrigerator for 12 h. Distilled water was added, and the ratio of Guizhou buckthorn powder to distilled water was 1 g:10 mL. The mixture was soaked in water for 4 h, and stirred every 30 min. The mixture was heated in a 65°C water bath for 60 min. After heating, the mixture was treated with ultrasonic waves at a frequency of 25 KHz, a power of 300 w, and a temperature of 45°C for 30 min. The mixture was filtered with sterile gauze, and the filtrate was poured out and filtered twice with a suction filter. The filtrate was concentrated with a rotary evaporator to form a flow extract (viscosity 15 Pa.s), which was the Guizhou buckthorn concentrated extract.

[0045] (2) Preparation of ointment patch

[0046] The Guizhou buckthorn concentrated extract 40 parts, propylene glycol 100 parts, polyisobutylene 30 parts, sodium lauryl sulfate 20 parts, lanolin 20 parts, and gypsum powder 15 parts were accurately weighed by weight parts, added to 100 mL of propylene glycol, and stirred until the matrix melted. The mixture was heated in a 75°C water bath and stirred at a constant speed. After the matrix melted, the heating and stirring were stopped. Then, 40 mL of the Guizhou buckthorn concentrated extract was added to the mixture and stirred until uniform. The ointment patch solution was obtained, sterilized, and ready for use. 3 mL of the ointment patch solution was taken with a 5 mL pipette and placed in the circle in the middle of the patch cloth. The mixture was naturally cooled to form a solid at room temperature, and the Guizhou buckthorn skin wound healing ointment patch was obtained.

[0047] Test Example 1: Effect verification of promoting wound healing

[0048] I. Experimental method

[0049] 1. Model preparation and grouping

[0050] 1.1 Preparation of rabbit back wound model

[0051] Twelve New Zealand white rabbits were anesthetized by injecting 35 mg / kg of 10% chloral hydrate into the marginal ear vein. The back of each rabbit was shaved with an electric shaver, and the skin was disinfected with alcohol. Then, a circular full-thickness skin defect about 1.0 cm in diameter was cut with surgical scissors to a depth of the fascia, thus creating a full-thickness wound model. Each rabbit had four wounds, which were labeled A, B, C, and D. Different substances were used to treat the different wounds, as described in 1.2. All wounds were treated with the original method after being washed with normal saline and disinfected with iodophor every day. Each rabbit was housed in a separate cage. Twelve New Zealand white rabbits were randomly divided into two groups, with six rabbits in each group. The first group of six rabbits had a total of 24 wounds on the back. The treatment effect was evaluated by the wound healing rate at 0 d, 4 d, 8 d, and 12 d after the operation. The second group of six rabbits also had a total of 24 wounds on the back. Tissue samples were taken from the skin and the newly formed granulation tissue at the wound edge at 0 d, 4 d, 8 d, and 12 d after the operation. The left wound tissue was paraffin-embedded for tissue sectioning and then stained with hematoxylin and eosin (HE) and picrosirius red. The right wound tissue was used for immunohistochemical detection of CD34 and VEGF.

[0052] 1.2 Grouping

[0053] Each rabbit had four wounds on the back, and the grouping is shown in Table 1.

[0054] Table 1 Grouping

[0055] Group Treatment Dosage A Normal saline 0.5ml B Yunnan white medicine 0.5g C Guizhou buckthorn skin wound healing ointment 1 piece D Guizhou buckthorn extract 0.5ml

[0056] After the application of the medicine, all wounds were covered with petroleum jelly gauze and then wrapped with dry gauze, which was changed every day.

[0057] The Guizhou buckthorn skin wound healing ointment of group C was prepared according to the method described in Example 1.

[0058] 2. Observation index

[0059] 2.1 General observation of the wound

[0060] The basic conditions of the New Zealand rabbits were observed by naked eye at 4 d, 8 d, and 12 d after the operation, including whether the wound was red and swollen, whether there was exudation, whether there was purulent secretion, the wound area, the contraction of the skin tissue around the wound, and the new growth at the wound edge.

[0061] 2.2 Wound healing rate

[0062] The six New Zealand rabbits were fixed at 4 d, 8 d, and 12 d after the operation. The wound was covered with transparent film and traced. The film was then applied to the grid paper for grid counting. The actual area of the wound in each group was measured, and the wound healing rate at different time points was calculated.

[0063] Wound healing rate = (original wound area - unhealed wound area) / original wound area x 100%

[0064] 2.3 Collection and processing of tissue specimens

[0065] Immediately after collection, the skin tissue was fixed in 4% paraformaldehyde for 24 hours, then dehydrated in an automated tissue dehydrator. After dehydration, it was embedded in paraffin. Paraffin sections of 4 μm thickness were continuously cut from each tissue specimen and stored at 4°C for later use. A portion of the sections underwent routine HE histological staining, and the epidermal and dermal structures, tissue necrosis, and degree of inflammation were observed under a light microscope. The remaining sections underwent SP immunohistochemical staining, CD34-labeled microvessel density (MVD) counting, and EGFR analysis using the MagePro Plus image analysis software based on the SP immunohistochemical method. The average optical density (AOD) of the images was measured.

[0066] 3. Data Processing

[0067] Statistical analysis was performed using Excel, GraphPad Prsim 8, and SPSS 23.0. Quantitative data were expressed as mean ± standard deviation (x ± s). One-way ANOVA was used for comparisons among multiple groups. P < 0.05 and P < 0.01 were considered statistically significant.

[0068] II. Experimental Results

[0069] 1. Gross observation of the wound

[0070] like Figure 1 As shown, on postoperative day 4, all four groups of skin wounds had scabbed over, with no redness, swelling, exudation, or obvious infection. Scabs began to form on each group's wounds, shrinking and causing wrinkles in the surrounding skin. On postoperative day 8, scabs had formed on all groups' wounds, darkening in color and turning brown. In the three groups treated with medication, the scabs were firmly attached to the base, with significant edge shrinkage. Yunnan Baiyao showed the best recovery, while the saline group still exhibited varying degrees of wound edge reaction. On postoperative day 12, the wounds in the Yunnan Baiyao and Guizhou Rhamnos extract groups were almost completely covered by new epithelium, with only some scabs remaining. The Guizhou Rhamnos skin wound healing ointment group recovered worse than these two groups, with some scab tissue still present. The saline group's wounds shrank significantly, with some wound tissue covered by thin and easily broken new epithelium, but wound healing was significantly worse than the other three groups.

[0071] 2. Effects of drug combination spray on wound healing rate in rabbits

[0072] The area of the wound surface of the four groups was gradually reduced with the extension of postoperative time. As shown in Table 2, the percentage of the wound healing area of the normal saline group was 33.45±8.79% on the 4th day, 50.96±8.51% on the 8th day, and 77.80±9.72% on the 12th day after the operation; the percentage of the wound healing area of the Yunnan white group was 34.21±11.42% on the 4th day, 66.07±12.68% on the 8th day, and 92.11±5.63% on the 12th day after the operation; the percentage of the wound healing area of the Guizhou buckthorn extract group was 36.86±12.5% on the 4th day, 65.61±11.60% on the 8th day, and 93.71±7.27% on the 12th day after the operation; and the percentage of the wound healing area of the Guizhou buckthorn skin ointment group was 33.11±11.47% on the 4th day, 64.11±11.46% on the 8th day, and 89.39±9.04% on the 12th day after the operation. The statistical results show that the wound healing rates of the three groups of the medicaments are higher than that of the normal saline group on the 8th day and the 12th day after the operation, and the difference is statistically significant (P<0.05, P<0.01).

[0073] Table 2 Wound healing rate

[0074]

[0075] Note: Compared with the normal saline group, extremely significant is indicated by “**” (P<0.01), and significant is indicated by “*” (P<0.05).

[0076] 3. Histopathological changes of the wound granulation tissue of the New Zealand rabbits in each group

[0077] The specimens of the wound surface of the five groups were observed under a microscope on the 4th day, the 8th day, and the 12th day after the debridement, and the results are as follows: On the 4th day after the operation, a large number of inflammatory cells were observed under the microscope in each group, and part of the collagen fibers were cracked. A small amount of epithelial cells were observed in the Yunnan white group. On the 8th day after the operation, the inflammatory cells were obviously reduced in the Yunnan white group, the hypericum group, and the gold+Chinese medicine group. Epithelial cells grew on the wound surface in each group, and the epithelial hyperplasia and the collagen fiber hyperplasia were obvious. The inflammatory cells were also reduced in the Chinese medicine group, but not as much as in the other three groups. However, a large number of inflammatory cells still existed in the normal saline group. On the 12th day after the operation, the wound tissue structure was complete, and the levels were relatively clear in the four groups of the medicaments. A large number of new epithelial cells and gland-like structures were observed in some of the groups. The collagen fibers were arranged in a woven manner, and the inflammatory cells were less. The new epithelium was thin, the collagen fibers were arranged in disorder, the gland-like structures were few, and a large number of inflammatory cells were observed in the normal saline group. The details are shown in Table 3. Figure 2 .

[0078] 4. Microvessel counting of the wound surface

[0079] The microvessel count of each group showed an upward trend at 4d, 8d, 12d after debridement. The microvessel count of Yunnan white medicine, Guizhou Rhamnus extract and Guizhou Rhamnus skin wound healing ointment group was significantly higher than that of the normal saline group at each time point. The microvessel count of Yunnan white medicine and Guizhou Rhamnus extract group was statistically different from that of the normal saline group at each time point (P<0.05, P<0.01). However, the microvessel count of Guizhou Rhamnus skin wound healing ointment group was not statistically different from that of the normal saline group at 8d (P>0.05). See Table 2 for details. Figure 3 .

[0080] 5. AOD value of EGFR

[0081] The average optical density value of EGFR in normal condition met the rule of first increasing and then decreasing. At 4d, 8d and 12d after administration, the expression of EGFR in Yunnan white medicine group and Guizhou Rhamnus skin wound healing ointment group showed a trend of first increasing and then decreasing, and reached about 0.50 at 8d, which reflected the early release and strong expression of rh-EGF induced EGFR, and possibly promoted the epithelial cell division and proliferation, leading to early epithelialization of the wound. The expression of EGFR in the normal saline group showed an upward trend, indicating that the healing tissue matured late. The expression of EGFR in Guizhou Rhamnus extract group showed a downward trend, which might be related to the early maturation of healing tissue and the early end of cell proliferation period, and involved the degree of scar hyperplasia after healing. See Table 3 for details.

[0082] Table 3 AOD of EGFR at different time points in the three groups (x±s)

[0083]

[0084] III. Conclusion

[0085] Through general observation of the wound of rabbits, wound healing rate, healing time, pathological staining, immunohistochemical marker microvessel count and average optical density value of EGFR, the following conclusions were drawn:

[0086] (1) The two kinds of traditional Chinese medicine preparations made of Guizhou Rhamnus, Guizhou Rhamnus extract and Guizhou Rhamnus skin wound healing ointment, have the effect of promoting wound healing, can eliminate inflammation, promote the formation of new blood vessels in granulation tissue of chronic skin ulcer wound, improve the pathological changes of ulcer wound, and ultimately promote wound healing.

[0087] (2) The effect of Guizhou Rhamnus extract on promoting wound healing may be better than that of Yunnan white medicine and Guizhou Rhamnus skin wound healing ointment.

[0088] The principles and implementation manners of the present application are described by using specific examples in the present application, and the above examples are only used to help understand the method of the present application and its core idea; meanwhile, for the general technical personnel in the art, the specific implementation manners and application ranges will be changed according to the idea of the present application. In conclusion, the content of the present specification should not be understood as the limitation of the present application.

Claims

1. A Guizhou buckthorn skin wound healing plaster, characterized in that, The plaster is composed of the following: 35-55 parts by weight of concentrated extract of Rhamnus ventricosa and 150-200 parts by weight of matrix excipients; The matrix auxiliary material is composed of 85-120 parts by weight of propylene glycol, 20-30 parts by weight of polyisobutylene, 15-20 parts by weight of sodium dodecyl sulfate, 20-35 parts by weight of lanolin, and 5-20 parts by weight of gypsum powder. The preparation method of the concentrated extract of Guizhou rhamnoides is as follows: Guizhou rhamnoides powder is mixed with rapeseed oil and stirred evenly. It is placed at 4°C for 12 h, and then soaked in double-distilled water. After soaking, the mixture is heated in a water bath. After heating, it is ultrasonically treated and filtered. The filter residue is extracted twice, and the filtrates are combined for later use. The filtrate is concentrated by rotary evaporation to a fluid extract with a viscosity of 10~25 Pa.s. The concentrate is sterilized by high pressure for 30 min to obtain the concentrated extract of Guizhou rhamnoides.

2. The Guizhou Rhamnus skin wound healing plaster according to claim 1, characterized in that, The preparation method of the Guizhou buckthorn powder is as follows: clean Guizhou buckthorn branches are collected, dried at 55°C to constant weight, pulverized, and passed through a 200-mesh sieve to obtain Guizhou buckthorn powder.

3. The Guizhou Rhamnus skin wound healing plaster according to claim 1, characterized in that, In the preparation method described above, the ratio of Guizhou buckthorn powder to rapeseed oil in the liquid is 1~2 g / mL.

4. The Guizhou Rhamnus skin wound healing plaster according to claim 1, characterized in that, In the preparation method described above, the ratio of Guizhou Rhamnus powder to double-distilled water is 1 g / 10~20 mL.

5. The Guizhou Rhamnus skin wound healing plaster according to claim 1, characterized in that, The soaking time in the preparation method is 3-4 hours, and the mixture is stirred once every 30 minutes during the soaking process.

6. The Guizhou Rhamnus skin wound healing plaster according to claim 1, characterized in that, In the preparation method, the water bath heating temperature is 55℃~70℃, and the water bath heating time is 30~60 min.

7. The Guizhou Rhamnus skin wound healing plaster according to claim 1, characterized in that, In the preparation method, the ultrasonic treatment frequency is 25~30 KHz and the ultrasonic treatment time is 30 min.

8. The preparation method of the Guizhou Rhamnus skin wound healing plaster according to any one of claims 1 to 7, characterized in that, Includes the following steps: (1) Weigh out lanolin, sodium dodecyl sulfate, gypsum powder and polyisobutylene by weight, mix the above substances evenly, add propylene glycol and mix evenly, and heat in a water bath at 75°C until the matrix melts under uniform stirring conditions. (2) Add concentrated extract of Guizhou buckthorn to the melted matrix, mix the raw materials evenly, and sterilize to obtain plaster solution; (3) Place the plaster solution in the circle in the middle of the plaster and let it cool naturally at room temperature to solidify, thus obtaining the external plaster that promotes skin wound healing.

9. The preparation method of the Guizhou Rhamnus skin wound healing plaster according to claim 8, characterized in that, The amount of plaster solution used in the plaster is 3 mL per plaster.

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