Carboxyamidotriazole water-soluble ointment and preparation method thereof

Abstract

The invention provides a carboxamide triazole water-soluble ointment and a preparation method thereof, and relates to the technical field of pharmaceutical preparations. The water-soluble carboxamide triazole ointment comprises carboxamide triazole and polyethylene glycol, and further comprises an antioxidant, and / or a pH regulator, and / or a penetration enhancer. Wherein the polyethylene glycol is a mixture of low-molecular-weight polyethylene glycol and high-molecular-weight polyethylene glycol, the low-molecular-weight polyethylene glycol is PEG400, and the high-molecular-weight polyethylene glycol is selected from at least one of PEG6000 and PEG8000. According to the water-soluble carboxamidotriazole ointment, polyethylene glycol with different molecular weights is used as a matrix and is matched with other auxiliary material components, so that the water-soluble carboxamidotriazole ointment is moderate in viscosity, convenient to smear and good in chemical stability, the medicine can be quickly released from the preparation, the in-vitro release capacity and the in-vitro skin penetration amount are improved, and the treatment effect on psoriasis is improved.

Description

Technical Field

[0001] The invention belongs to the technical field of pharmaceutical preparations, and particularly relates to a carboxyamide triazole water-soluble ointment and a preparation method thereof. Background Art

[0002] Carboxyamidotriazole (CAI) is a synthetic small molecule compound. Many studies have confirmed that carboxyamidotriazole is a non-cytotoxic calcium ion antagonist. It has shown the effect of inhibiting the proliferation and metastasis of various tumors in both in vivo and in vitro models, and can inhibit angiogenesis. The results of previous studies have shown that carboxyamidotriazole also has significant anti-acute and chronic inflammatory effects. Patent CN101553225B experimentally confirmed that carboxyamidotriazole compounds can be used as inhibitors of inflammatory cytokines TNF-α and IL-1, and proposed the use of carboxyamidotriazole compounds in the treatment of pain caused by any reason in mammals and inflammation involving inflammatory cytokines, as well as other immunological-based related diseases. Patent CN111249277A discloses the role of carboxyamidotriazole compounds in treating a series of autoinflammatory diseases, including common skin diseases such as psoriasis.

[0003] Psoriasis is a chronic inflammatory skin disease with a high incidence, stubborn condition and easy recurrence. It is actually an immune disease, and the course of the disease mostly lasts a lifetime, which is extremely harmful to people's physical and mental health. Psoriasis is characterized by erythema, papules and scales. Its main histopathological changes are: excessive proliferation of keratinocytes (KC), infiltration of inflammatory cells in the dermis, and dilation of blood vessels in the dermal papilla; among them, excessive proliferation of KC is its most important feature, which is manifested as accelerated proliferation and metabolism of keratinocytes, shortened epidermal cell conversion cycle, thickening of the epidermis accompanied by abnormal keratinization such as hyperkeratosis and parakeratosis, and disappearance of the granular cell layer.

[0004] The etiology and pathogenesis of psoriasis are extremely complex and have not yet been fully understood. In recent years, it is believed that psoriasis is an immune disease caused by the complex interaction between highly proliferating keratinocytes and immune cells such as T cells, neutrophils, dendritic cells and macrophages, which is caused by the combined effects of genetic gene regulation and environmental factors, resulting in dysfunction of the patient's nervous system, biochemical substances, and immune system. The interaction between KC and these immune cells ultimately leads to excessive proliferation of KC and dermal inflammation, showing the unique clinical and pathological characteristics of psoriasis. A large number of cytokines are involved in its pathogenesis, among which cytokines such as TNF-α, IL-17, IL-12 and IL-23 are very important immune mediators that have been identified. At present, there is no specific treatment for psoriasis, and the main treatment methods are symptomatic treatment of topical and oral drugs. Most of these drugs are hormones or antibiotics. Appropriate symptomatic treatment can temporarily control symptoms, but it only treats the symptoms and not the root cause. Long-term use will also cause repeated illness, incurable, and cause many adverse reactions.

[0005] As inhibitors of inflammatory cytokines such as TNF-α, carboxyamide triazole compounds provide new ideas for the treatment of psoriasis. Currently, only carboxyamide triazole soft capsules have completed phase III clinical trials for the treatment of non-small cell lung cancer. Patent CN112353778B discloses a carboxyamide triazole soft capsule and a preparation method thereof. The soft capsule has good stability, and the drug content does not decrease during long-term storage, and the increase of related substances is not obvious. However, the amount of drug absorbed by the soft capsule after oral administration through the blood circulation to the skin is limited, so it is particularly important to develop local external preparations. External preparations are administered to the affected skin, requiring the drug to be released from the preparation and penetrate the skin stratum corneum to enter the skin, and to be able to stay in the skin for a long time to play a role.

[0006] Topical preparations are generally ointments, which can be evenly applied when used, and the drug adheres well to the skin surface. According to the type of matrix, it can be divided into oil-soluble matrix, water-soluble matrix and emulsion matrix. The oil-soluble matrix is ​​lubricating and non-irritating. It can form a closed oil film when applied to the skin, promote skin hydration, and have a protective and softening effect on the skin. The disadvantage is that it is greasy and has poor water absorption. The emulsion matrix is ​​composed of an aqueous phase, an oil phase and an emulsifier. Its consistency is suitable and easy to apply, but due to the presence of a large amount of aqueous phase, the drug is required to have a certain solubility in water, and other additives such as preservatives and moisturizers need to be added. The water-soluble matrix is ​​a semi-solid ointment matrix made of natural or synthetic water-soluble polymer materials. This type of matrix is ​​soluble in water, easy to wash off, and the drug release is faster. The water-soluble matrix is ​​mainly polyethylene glycol and cellulose. Patent CN108969531 reports that polyethylene glycol can fix water molecules through a large number of hydrogen bonds to retain water in the stratum corneum or dermis of the skin, increase the amount of water retained in the skin, and enhance the permeability of the skin.

[0007] At present, there is very little research on carboxyamide triazole ointment, and carboxyamide triazole has poor water solubility, but can be dissolved in polyethylene glycol adjuvants. In view of this, the present invention is mixed with polyethylene glycol adjuvants of different molecular weights by carboxyamide triazole in the same preparation, and a carboxyamide triazole water-soluble ointment and preparation method thereof are provided, with polyethylene glycol adjuvants of different molecular weights as matrix, and carboxyamide triazole and other additives are dissolved in polyethylene glycol matrix. Other auxiliary material components are used in conjunction with, and the viscosity of the ointment provided is moderate, convenient to apply, and chemical stability is good, and the drug can be quickly released from the preparation, through the skin stratum corneum, penetrate and be retained in the skin, and each component, especially the polyethylene glycol component, is combined to play the effect of treating psoriasis in coordination. Summary of the invention

[0008] The invention aims at the problems existing in the prior art and provides a carboxyamide triazole water-soluble ointment and a preparation method thereof, which overcomes the problems of poor chemical stability of active ingredients in the carboxyamide triazole ointment, migration of polyethylene glycol molecules during storage, and hardening of the ointment; the ointment prepared by the invention has moderate viscosity, stable chemical components during long-term storage, strong skin penetration ability, a certain skin stratum corneum penetration amount and skin retention amount, and fully exerts the drug effect.

[0009] To achieve the above purpose, the technical solution adopted by the present invention is as follows:

[0010] Firstly, the invention provides a carboxyamide triazole water-soluble ointment, which comprises the following components: carboxyamide triazole and polyethylene glycol.

[0011] Preferably, the carboxyamide triazole water-soluble ointment comprises the following ingredients in parts by weight: 0.5-8 parts of carboxyamide triazole and 92-99.5 parts of polyethylene glycol.

[0012] Further preferably, the polyethylene glycol is a mixture of low molecular weight polyethylene glycol and high molecular weight polyethylene glycol, the low molecular weight polyethylene glycol is polyethylene glycol 400 (PEG400), and the high molecular weight polyethylene glycol is at least one selected from polyethylene glycol 6000 (PEG6000) and polyethylene glycol 8000 (PEG8000).

[0013] Further preferably, in the polyethylene glycol, the weight ratio of the low molecular weight polyethylene glycol to the high molecular weight polyethylene glycol is 59-68:30-39.

[0014] Preferably, the carboxyamide triazole water-soluble ointment further comprises an antioxidant as an ingredient.

[0015] Further preferably, the carboxyamide triazole water-soluble ointment comprises the following ingredients in parts by weight: 0.5-8 parts of carboxyamide triazole, 91.9-99.499 parts of polyethylene glycol and 0.001-0.5 parts of antioxidant.

[0016] Further preferably, the antioxidant is selected from at least one of tocopherol, BHA (butylated hydroxyanisole) and BHT (butylated hydroxytoluene).

[0017] Preferably, the carboxyamide triazole water-soluble ointment further comprises a pH regulator to adjust the pH value of the carboxyamide triazole water-soluble ointment to 4-9.

[0018] Further preferably, the pH adjuster is selected from at least one of phosphate buffered saline (PBS), citric acid solution, sodium hydroxide solution and hydrochloric acid solution.

[0019] Preferably, the carboxyamide triazole water-soluble ointment comprises the following ingredients: carboxyamide triazole, polyethylene glycol, an antioxidant, a pH regulator and a penetration enhancer.

[0020] Further preferably, the carboxyamide triazole water-soluble ointment comprises the following ingredients in parts by weight: 0.5-3 parts of carboxyamide triazole, 90-98 parts of polyethylene glycol, 2-10 parts of a penetration enhancer and 0.001-0.5 parts of an antioxidant, and the pH adjuster adjusts the pH value of the carboxyamide triazole water-soluble ointment to 4-9.

[0021] More preferably, the carboxyamide triazole water-soluble ointment comprises the following ingredients in parts by weight: 0.5-3 parts of carboxyamide triazole, 90-98 parts of polyethylene glycol, 2-10 parts of a penetration enhancer and 0.001-0.3 parts of an antioxidant, and the pH adjuster adjusts the pH value of the carboxyamide triazole water-soluble ointment to 4-9.

[0022] More preferably, the carboxyamide triazole water-soluble ointment comprises the following ingredients in parts by weight: 1-2.5 parts of carboxyamide triazole, 92-93 parts of polyethylene glycol, 2-10 parts of a penetration enhancer, 0.001-0.1 parts of an antioxidant, and a pH adjuster to adjust the pH value of the carboxyamide triazole water-soluble ointment to 5-7.

[0023] Preferably, the polyethylene glycol is a mixture of low molecular weight polyethylene glycol and high molecular weight polyethylene glycol; the low molecular weight polyethylene glycol is polyethylene glycol 400 (PEG400), and the high molecular weight polyethylene glycol is selected from at least one of polyethylene glycol 6000 (PEG6000) and polyethylene glycol 8000 (PEG8000).

[0024] Further preferably, the polyethylene glycol is a mixture of polyethylene glycol 400 and polyethylene glycol 6000; or a mixture of polyethylene glycol 400 and polyethylene glycol 8000; or a mixture of polyethylene glycol 400, polyethylene glycol 6000 and polyethylene glycol 8000.

[0025] Further preferably, in the polyethylene glycol, the mass ratio of the low molecular weight polyethylene glycol to the high molecular weight polyethylene glycol is 54-68:30-39.

[0026] Preferably, the penetration enhancer is selected from at least one of dimethyl sulfoxide, oleic acid, azone, and diethylene glycol monoethyl ether.

[0027] More preferably, the penetration enhancer is dimethyl sulfoxide.

[0028] Preferably, the antioxidant is selected from at least one of tocopherol, BHA and BHT.

[0029] More preferably, the antioxidant is selected from at least one of tocopherol and BHT.

[0030] Preferably, the antioxidant accounts for 0.001-0.5% by weight in the carboxyamide triazole water-soluble ointment.

[0031] Preferably, the pH adjuster is selected from at least one of phosphate buffered saline (PBS), citric acid solution, sodium hydroxide solution and hydrochloric acid solution.

[0032] Then, the present invention provides a method for preparing the above-mentioned carboxyamide triazole water-soluble ointment, comprising the steps of:

[0033] (1) mixing part of the low molecular weight polyethylene glycol with all of the high molecular weight polyethylene glycol, heating to melt, stirring, and cooling to obtain a PEG matrix;

[0034] (2) mixing the carboxyamide triazole with the remaining weight portion of low molecular weight polyethylene glycol and the PEG matrix under heat preservation conditions, and stirring to homogenize and disperse;

[0035] (3) Stir, cool and bottle.

[0036] Preferably, in step (1), the part of low molecular weight polyethylene glycol accounts for 1 / 3-2 / 3 of the weight of all low molecular weight polyethylene glycol, g / g.

[0037] Preferably, in step (1), the heating and melting temperature is 65-80°C, and the cooling temperature is 50-65°C.

[0038] Preferably, in step (2), the stirring and homogenizing dispersion time is 10-15 minutes, and the insulation condition is 50-65°C.

[0039] Preferably, in step (3), the cooling temperature is 35-40°C.

[0040] Preferably, when the carboxyamide triazole water-soluble ointment further comprises ingredients: an antioxidant and / or a pH regulator and / or a penetration enhancer, the preparation method comprises the steps of:

[0041] (1) mixing part of the low molecular weight polyethylene glycol with all the high molecular weight polyethylene glycol, adding an antioxidant and / or a penetration enhancer, heating to melt, stirring, and cooling to obtain a PEG matrix;

[0042] (2) mixing the PEG matrix and the pH adjuster, and homogenizing for 10-15 minutes under heat preservation conditions to obtain a PEG mixed matrix;

[0043] (3) mixing the carboxyamide triazole with the remaining weight portion of low molecular weight polyethylene glycol and the PEG matrix under heat preservation conditions, and stirring to homogenize and disperse;

[0044] (4) under heat preservation conditions, mixing the carboxyamide triazole solution with the PEG mixed matrix of step (2), and stirring and homogenizing until the dispersion is uniform;

[0045] (5) Stir, cool and bottle.

[0046] Further preferably, in step (1), the part of low molecular weight polyethylene glycol accounts for 1 / 3-2 / 3 of the weight of all low molecular weight polyethylene glycol; the heating and melting temperature is 65-80°C, and the cooling temperature is 50-65°C.

[0047] Further preferably, in step (2), the insulation condition is 50-65° C., and the homogenization time is 10-15 min.

[0048] Further preferably, in steps (3)-(4), the insulation condition is 50-65°C.

[0049] More preferably, in steps (3)-(4), the temperature under the insulation conditions is 50-60°C.

[0050] Further preferably, in step (5), the cooling temperature is 35-40°C.

[0051] Finally, the present invention provides the use of polyethylene glycol 400 and / or polyethylene glycol 6000 and / or polyethylene glycol 8000 in preparing a highly stable carboxyamide triazole water-soluble ointment.

[0052] Compared with the prior art, the present invention has the following beneficial effects:

[0053] 1. The carboxyamide triazole water-soluble ointment provided by the present invention has good chemical stability and stable content under the specific ratio of polyethylene glycol with different molecular weights, and the related substances increase slowly, and the ointment does not harden after long-term storage; carboxyamide triazole is combined with a specific polyethylene glycol component to synergistically play a role in treating psoriasis, and the drug effect takes effect quickly and lasts for a long time; at a certain dosage, the effect of treating psoriasis is significant, and even better than the conventionally used drugs on the market.

[0054] 2. The carboxyamide triazole water-soluble ointment provided by the present invention further improves the chemical stability of the ointment under high temperature conditions by adding a pH adjuster with a specific pH value, and has less degradable impurities.

[0055] 3. The carboxyamide triazole water-soluble ointment provided by the present invention, by adding specific types and contents of other excipients and selecting appropriate penetration enhancers and antioxidants, can rapidly release the drug from the preparation while maintaining chemical stability, thereby increasing the amount of drug permeation through the skin stratum corneum and the amount of drug retained in the skin.

[0056] 4. The carboxyamide triazole water-soluble ointment prepared by a specific mixing, stirring and homogenizing preparation method has uniform and stable properties, moderate viscosity and is easy to apply. BRIEF DESCRIPTION OF THE DRAWINGS

[0057] Figure 1 It is an in vitro permeation curve of the carboxyamide triazole water-soluble ointment of Example 4 of the present invention.

[0058] Figure 2 The microscope structure diagram of the carboxyamide triazole water-soluble ointment of the present invention is shown in the figure below, wherein the magnification of the left diagram is 10×10, and the magnification of the right diagram is 40×10.

[0059] Figure 3 The invention discloses an in vitro release curve of the carboxyamide triazole water-soluble ointment.

[0060] Figure 4 It is a linear fitting diagram of the release curve of the in vitro release behavior of the carboxyamide triazole water-soluble ointment of the present invention.

[0061] Figure 5 The figure is a cumulative permeation curve diagram of the in vitro skin permeation test of the carboxyamide triazole water-soluble ointment of the present invention.

[0062] Figure 6 The present invention is a permeation rate curve of the in vitro skin permeation test of the carboxyamide triazole water-soluble ointment.

[0063] Figure 7 The present invention is a graph showing the changes in the body weight of mice at different time points in an anti-psoriasis test using the carboxyamide triazole water-soluble ointment.

[0064] Figure 8 The present invention is a graph showing changes in mouse erythema scores at different time points in an anti-psoriasis test of the carboxyamide triazole water-soluble ointment.

[0065] Fig. 9 The present invention is a graph showing the change in mouse scale scores at different time points in an anti-psoriasis test of the carboxyamide triazole water-soluble ointment.

[0066] Fig.10The present invention is a graph showing the change in mouse skin thickness scores at different time points in an anti-psoriasis test of the carboxyamide triazole water-soluble ointment.

[0067] Fig.11 The diagram is a graph showing changes in the PASI cumulative scores of mice at different time points in an anti-psoriasis test of the carboxyamide triazole water-soluble ointment of the present invention.

[0068] Fig.12 A diagram showing changes in skin lesions at the mouse modeling site at different time points in an anti-psoriasis test of the carboxyamide triazole water-soluble ointment of the present invention.

[0069] Fig.13 It is a bar graph of the IL-12 concentration results of the biochemical index test mice of the present invention.

[0070] Fig.14 It is a bar graph of the IL-17 concentration results of the biochemical index test mice of the present invention.

[0071] Fig.15 It is a bar graph of the IL-23 concentration results of the biochemical index test mice of the present invention.

[0072] Fig.16 It is a bar graph of the TNF-α concentration results of the biochemical index test mice of the present invention.

[0073] Fig.17a This is a picture of skin tissue pathology detection of mice in the model group of the present invention.

[0074] Fig.17b This is a skin tissue pathology detection diagram of mice in the positive control group 1 of the present invention.

[0075] Fig.17c This is a skin tissue pathology detection diagram of mice in the positive control group 2 of the present invention.

[0076] Fig.17d This is a picture of skin tissue pathology detection of mice in group 1 of the CAI ointment dosage of the present invention.

[0077] Fig.17e It is the skin tissue pathological detection picture of the mice in two groups of CAI ointment dosage of the present invention.

[0078] Fig.18 It is a statistical diagram of case scoring results of the present invention. DETAILED DESCRIPTION

[0079] The following non-limiting examples can enable those skilled in the art to more fully understand the present invention, but do not limit the present invention in any way. The following content is merely an exemplary description of the scope of the present invention, and those skilled in the art can make various changes and modifications to the present invention based on the disclosed content, which should also fall within the scope of the present invention.

[0080] When the embodiments give numerical ranges, it should be understood that, unless otherwise specified in the present invention, both endpoints of each numerical range and any numerical value between the two endpoints can be selected. Unless otherwise defined, all technical and scientific terms used in the present invention have the same meanings as those generally understood by those of ordinary skill in the art to which the present invention belongs.

[0081] The present invention is further described below by way of specific examples. The various chemical reagents used in the examples of the present invention are obtained through conventional commercial channels unless otherwise specified. The reagents and components described in the present invention have no significant effect on the effects of products from different manufacturers.

[0082] Example 1 Carboxyamide triazole water-soluble ointment polyethylene glycol matrix composition

[0083] The ingredients and weight portions of carboxyamide triazole water-soluble ointment are specifically shown in Table 1:

[0084] Table 1

[0085]

[0086] The stability of the carboxyamide triazole water-soluble ointment obtained by the above prescription was investigated, and the test results are shown in Table 2.

[0087] The viscosity detection method is as follows: using a Brookfield DVNext rheometer, CPA-51Z conical rotor, setting the speed to 0.4rpm, the detection temperature to 25°C, and the running time to 4min, placing an appropriate amount of sample between the rotor and the sample cup, and detecting the viscosity after the temperature is balanced.

[0088] The content detection method is: adopt HPLC method, use octylsilane bonded silica gel as the filling agent of the chromatographic column, the mobile phase is potassium dihydrogen phosphate buffer (pH3.2)-acetonitrile (64:36) and acetonitrile mixed in a volume ratio of 94:6, and detect at a detection wavelength of 264nm.

[0089] The detection method for the relevant substances is: adopting HPLC method, using octylsilane bonded silica gel as a chromatographic column filler, the detection wavelength is 264nm, and potassium dihydrogen phosphate buffer (pH3.2)-acetonitrile (64:36) and acetonitrile are used as the mobile phase for gradient elution detection.

[0090] Table 2

[0091]

[0092]

[0093] At 60°C, the growth rates of related substances in ointments based on polyethylene glycol 3350, polyethylene glycol 4000, polyethylene glycol 6000, and polyethylene glycol 8000 were similar, with no significant differences; however, the properties and viscosity stability of the ointments were different: the viscosity of the ointments based on polyethylene glycol 6000 and polyethylene glycol 8000 did not change significantly, and the spreadability was good. In addition, under low temperature conditions, the ointments based on polyethylene glycol 6000 and polyethylene glycol 8000 did not crystallize, while the ointments based on polyethylene glycol 3350 and polyethylene glycol 4000 did crystallize. The results showed that the ointments based on polyethylene glycol 6000 and polyethylene glycol 8000 had better stability.

[0094] Example 2 Type and dosage of antioxidant in carboxyamide triazole water-soluble ointment

[0095] The composition and weight of carboxyamide triazole water-soluble ointment are shown in Table 3, and the comparison is made between ointments containing different types of antioxidants:

[0096] Table 3

[0097]

[0098] The carboxyamide triazole water-soluble ointment obtained by the above prescription was subjected to a stability test using the same method as in Example 1. The test results are shown in Table 4.

[0099] Table 4

[0100]

[0101] The dosage of the antioxidant was investigated, and the specific components and weight portions of the carboxyamide triazole water-soluble ointment are shown in Table 5:

[0102] Table 5

[0103]

[0104] The carboxyamide triazole water-soluble ointment obtained by the above prescription was subjected to a stability test using the same method as in Example 1. The test results are shown in Table 6.

[0105] Table 6

[0106]

[0107] Since carboxylic acid amine triazole chemistry is sensitive to oxygen, antioxidants are added to the ointment. The effects of different antioxidants on the stability of the ointment were compared. The results showed that the results of related substances in prescriptions 5-7 at 60°C were similar, which were lower than those in prescriptions 2 without antioxidants and prescriptions 8 and 9, indicating that the addition of antioxidants dl-α-tocopherol, BHA, and BHT was beneficial to improve the stability of the ointment. Increasing the amount of antioxidants dl-α-tocopherol, BHA, and BHT to 0.5 parts did not significantly affect the stability.

[0108] Example 3 Type and dosage of pH regulator in carboxyamide triazole water-soluble ointment

[0109] The composition and weight of carboxyamide triazole water-soluble ointment are shown in Table 7, and the comparison is made between ointments containing different types of pH regulators and different pH values:

[0110] Table 7

[0111]

[0112] The carboxyamide triazole water-soluble ointment obtained by the above prescription was subjected to a stability test using the same method as in Example 1. The test results are shown in Table 8.

[0113] Table 8

[0114]

[0115]

[0116] Since carboxyamide triazole chemistry is sensitive to pH, the pH is adjusted in the formulation. pH adjusters generally include salt substances such as phosphates, sodium hydroxide, hydrochloric acid, and triethanolamine. The effects of different pH adjusters on stability were investigated, and the results are shown in Table 8. According to the investigation results, when the pH of the ointment is adjusted to 4-9, the maximum degradation impurity growth under high temperature conditions is smaller, and the stability is better.

[0117] Example 4 Types of penetration enhancers in carboxyamide triazole water-soluble ointment

[0118] The composition and weight of carboxyamide triazole water-soluble ointment are shown in Table 9, which compares ointments containing different types of penetration enhancers:

[0119] Table 9

[0120]

[0121] The carboxyamide triazole water-soluble ointment obtained by the above prescription was subjected to a stability test using the same method as in Example 1. The test results are shown in Table 10.

[0122] Table 10

[0123]

[0124] The carboxyamide triazole water-soluble ointment obtained by the above prescriptions 21-26 was subjected to an in vitro skin permeation experiment. The experimental method was: using a vertical diffusion cell, miniature pig skin as the experimental skin, isopropanol: 0.01M pH7.4 phosphate buffer (50:50, v / v) as the receiving medium, the ointment dosage was 0.3 g, the experiment was carried out at a temperature of 32°C and a rotation speed of 600 rpm, sampling was performed at different time points, each sampling volume was 12 mL, and the same volume of blank receiving medium was immediately supplemented after sampling, and the drug content in the receiving medium was determined.

[0125] The in vitro permeation curves of different formulation samples are shown in Figure 1 .

[0126] Polyethylene glycol excipients have poor skin permeability. In order to ensure that the raw materials in the preparations can be absorbed through the skin, the effect of the prescription penetration enhancer on in vitro permeation was investigated. Pig skin was used to compare the effects of different penetration enhancers on the in vitro permeation of the preparations, and their stability was investigated. The results show that the in vitro permeation results of prescriptions 21-25 are better than those of prescription 18 without a penetration enhancer, while prescription 26 is similar to prescription 18. Dimethyl sulfoxide has the greatest ability to promote the penetration of ointment drugs; oleic acid, azone, and diethylene glycol monoethyl ether are second. The stability of the ointment with dimethyl sulfoxide as a penetration enhancer is better than that of other prescriptions.

[0127] Test 1: Ointment appearance and properties

[0128] The appearance of the carboxyamide triazole water-soluble ointment was evaluated. The ointments of all the above prescriptions were white to off-white.

[0129] Among them, the ointments of prescriptions 2-4, 5-7, 10-12, 14-15, 18-19, 21, and 23-25 ​​have moderate viscosity. After being squeezed out, they can be evenly spread and adhered to the skin by applying them with a little force. No crystals are found under microscopic observation, and high molecular weight polyethylene glycol is evenly distributed in clusters. The microscopic structure of the ointment of prescription 21 is shown in the figure below: Figure 2 As shown, the magnification of the left image is 10×10, and the magnification of the right image is 40×10.

[0130] Test 2 Chemical stability test

[0131] Chemical stability tests were performed on prescriptions 21, 8, 16, 22, and 28. The results of the accelerated stability tests are shown in Tables 11-12. Table 11 shows the stability results of the ointment at 30±2°C (65%RH±5%RH), and Table 12 shows the accelerated stability results of the ointment at 40±2°C (65%RH±5%RH).

[0132] The specific detection method is: stability testing is carried out at 30±2°C (65%RH±5%RH) and 40±2°C (65%RH±5%RH) respectively.

[0133] Table 11

[0134]

[0135]

[0136] Table 12

[0137]

[0138]

[0139] Long-term chemical stability test was conducted on prescription 21. The results of long-term stability test are shown in Tables 13-14. Table 13 is the long-term stability result of the ointment at 20±2℃ (60%RH±5%RH), and Table 14 is the long-term stability result of the ointment at 25±2℃ (65%RH±5%RH). The accelerated chemical stability results of the remaining prescriptions showed poor stability at 6 months, and no long-term stability test was required.

[0140] The specific testing method is: testing at 20±2℃ (60%RH±5%RH) and 25±2℃ (65%RH±5%RH) respectively. The results showed that the stability of prescription 21 was good for 18 months and met the medicinal requirements of ointment preparations.

[0141] Table 13

[0142]

[0143] Table 14

[0144]

[0145] The prescription ointment of the present invention has good chemical stability under accelerated test conditions, stable content, slow increase of related substances, no increase in viscosity, and no tendency of the ointment to harden.

[0146] The prescription ointment of the present invention has good chemical stability under long-term test conditions, stable content, slow increase of related substances, no increase in viscosity, and no tendency of the ointment to harden.

[0147] Test 3: In vitro release and in vitro skin permeation test

[0148] Prescriptions 21, 8, 16, 22, and 28 were tested for in vitro release and in vitro skin permeation.

[0149] The in vitro release results of each prescription ointment are as follows Figure 3-4 As shown, Figure 3 is the in vitro release curve, Figure 4 The linear fitting diagram of the release curve of in vitro release behavior.

[0150] The in vitro skin permeation test results of each prescription ointment are as follows: Figure 5-6 As shown, Figure 5 This is the cumulative permeation curve of the in vitro skin permeation test. Figure 6 It is the permeation rate curve of in vitro skin permeation test.

[0151] Test 4 Anti-psoriasis test

[0152] The therapeutic effect of the test substance on psoriasis was investigated using a psoriasis mouse model induced by imiquimod cream.

[0153] 1. Experimental method: After the quarantine period, 40 quarantine qualified mice were selected and randomly divided into model group (n=10), positive control group 1 (n=5), positive control group 2 (n=5), CAI ointment dose group 1 (n=10), and CAI ointment dose group 2 (n=10) according to body weight. Ensure that the weight level of animals between groups is balanced. Modeling was carried out immediately after grouping. The area of ​​about 5cm×3cm on the back of all mice was depilated and skin was prepared. Imiquimod cream was applied to the back of the mice twice a day for 5 consecutive days. The drug was administered on the same day after the modeling was completed. The drug administration period was 3 days, and the corresponding test substance was applied to the modeling site twice a day. During the experimental period, the mice were photographed, weighed, and PASI scored every day. One day after the last administration, the mice were euthanized, and the full-thickness skin of the modeling site was separated for ELISA detection and pathological detection. The CAI ointment described in this experiment was homemade from the ointment of prescription 21. Table 15 shows the information of the test substance, reference substance, and modeling agent used in the experiment.

[0154] Table 15

[0155]

[0156] 2. Modeling method: 40 SPF-grade BALB / c male mice were adaptively raised in a barrier facility for 7 days and observed daily to confirm that the mice were healthy and normal before modeling. All mice were depilated and skinned on an area of ​​about 5cm×3cm on the back. Imiquimod cream was applied to the back of the animals twice a day for 7 consecutive days. After confirming that the animals were in normal condition, they were returned to the cage and raised normally. After modeling, the skin lesions at the modeling site were closely monitored.

[0157] The BALB / c mice were SPF grade, male, 7 weeks old, 40 mice, with a planned weight range of 20.9-26.4 g when purchased and a weight range of 20.6-26.0 g during the formal test. Source: Guangdong Yaokang Biotechnology Co., Ltd., Laboratory Animal Production License: SCXK (Guangdong) 2020-0054; Laboratory Animal Quality Certificate: No. 44824700017889.

[0158] 3. Grouping and administration: After the quarantine period, 40 quarantine-qualified mice were selected and randomly divided into 5 groups according to their weight, with 5 or 10 mice in each group. Ensure that the weight level of mice between groups is balanced. Modeling was carried out immediately after grouping, and administration was carried out on the same day after modeling. Details of grouping and administration are shown in Table 16. Among them, the model group was smeared with water; the positive control group 1 was administered with calcipotriol ointment; the positive control group 2 was administered with retinoic acid cream; CAI ointment dose group 1 and group 2 were administered with the carboxyamide triazole water-soluble ointment of the present invention, prescription 21.

[0159] Table 16

[0160] Serial number Group Dosage Dosage Number of animals 1 Model Group / / 10 2 Positive control group 1 0.075g Transdermal administration, bid*3 5 3 Positive control group 2 0.075g Transdermal administration, bid*3 5 4 CAI ointment dosage 1 group 0.075g Transdermal administration, bid*3 10 5 CAI ointment dosage group 2 0.150g Transdermal administration, bid*3 10

[0161] Note: bid*3 in the above table means that the drug is administered twice a day for 3 consecutive days.

[0162] 4. Observation and indicator monitoring

[0163] General observation modeling period: After the animal modeling, the animals are observed and recorded every day. The observation contents include: whether they are dead, dying, feed and water intake, trauma, feces, appearance and coat, mental state, activity status, etc.

[0164] Dosing period: The animals in this group were observed and recorded within 15 minutes to 1 hour after daily dosing. The observation contents were the same as above.

[0165] Body weight, quarantine period: Animals are weighed upon receipt and after the 7-day quarantine period.

[0166] Modeling and drug administration period: The animals were weighed and recorded daily after the modeling began, and were weighed and recorded daily during the drug administration period.

[0167] PASI score: After modeling, the animals were scored and recorded daily according to Table 17. Table 17 is the PASI scoring standard for psoriasis models.

[0168] Table 17

[0169] score erythema Scaly thickness 0 No erythema No scale on the surface The lesions are flush with normal skin 1 Reddish red erythema Some lesions have scales on the surface The lesions are slightly higher than normal skin 2 Red erythema Most lesions have scaling Moderately raised lesions 3 Dark red erythema The lesion is almost covered with scales Skin lesions are thickened and raised 4 Very deep erythema The lesions are completely covered with scales The lesions are highly thickened and prominent

[0170] 5. Animal handling

[0171] After the last administration, the mice were killed by cervical dislocation, and the full-thickness skin of the modeling site of each animal was separated, fixed with 4% paraformaldehyde and frozen at -80°C.

[0172] Pathological examination and cytokine level detection: The sample tissue fixed with 4% paraformaldehyde was dehydrated, paraffin-embedded and sectioned, and HE staining was performed to observe the pathological changes of the skin at each animal modeling site under a microscope. The sample tissue homogenate frozen at -80℃ was tested by Elisa for changes in IL-12, IL-23, IL-17, and TNF-α.

[0173] 6. Data analysis and processing

[0174] All experimental data are expressed as mean ± standard deviation (Mean ± SD). Before comparing the experimental groups, the inter-group variance analysis (F-test) was performed. When the inter-group variances were equal, the inter-group comparison was performed using the Student-T test (unpaired T test). When the inter-group variances were unequal, the corrected Student-T test was used for statistical analysis. The data were statistically analyzed and statistical graphs were drawn using Graphpad prism 8 software.

[0175] 7. Test results:

[0176] 7.1 Gross Observation: After administration, the mental state and activity of the mice in each group were normal, and no abnormalities were observed.

[0177] 7.2 Body weight: The body weight changes of mice in each group at different time points are as follows: Figure 7 As shown in Table 18. The weight levels of mice in each drug administration group changed consistently, all of which decreased to a certain extent during the modeling period (d0-d5), and the weight levels stabilized from the end of the late modeling period to after drug administration (d5-d10). There was no significant difference in the weight levels of mice in each group (P>0.05).

[0178] Table 18

[0179]

[0180] 7.3PASI score Erythema score: The erythema scores of the modeling sites of each group of mice at different time points are as follows Figure 8 , as shown in Table 19 (n = 5 or 10). Compared with the model group, the erythema score of mice in CAI ointment dose group 1 was significantly reduced on d8 (1 day after administration) (P < 0.05), and the erythema score was significantly reduced on d9-d10 (2-3 days after administration) (P < 0.01); the erythema score of mice in CAI ointment dose group 2 was significantly reduced on d10 (3 days after administration) (P < 0.01); the erythema score of mice in the positive control group 1 was significantly reduced on d9-d10 (2-3 days after administration) (P < 0.01). There was no statistical difference in the erythema score of mice in the positive control group 2 compared with the control group (P > 0.05).

[0181] Table 19

[0182]

[0183]

[0184] In Table 19, compared with the model group, * indicates P < 0.05, and ** indicates P < 0.01.

[0185] 7.4 Scale score

[0186] The scale scores of the modeling sites of each group of mice at different time points are as follows Fig. 9 As shown in Table 20 (n=5 or 10), compared with the model group, the squama score of mice in CAI ointment dose group 1 was significantly reduced on d9-d10 (2-3 days after administration) (P<0.01); the squama score of mice in CAI ointment dose group 2 was significantly reduced on d10 (3 days after administration) (P<0.01); the squama score of mice in the positive control group 1 was significantly reduced on d8-d10 (1-3 days after administration) (P<0.05 or P<0.01). The squama score of mice in the positive control group 2 was significantly reduced on d10 (3 days after administration) (P<0.01).

[0187] Table 20

[0188]

[0189] In Table 20, compared with the model group, * indicates P < 0.05, and ** indicates P < 0.01.

[0190] 7.5 Skin Thickness Score

[0191] The skin thickness scores of the modeling sites of each group of mice at different time points are as follows Fig.10 As shown in Table 21 (n=5 or 10), compared with the model, the skin thickness scores of mice in CAI ointment dose group 1 were significantly reduced on d9-d10 (2-3 days after administration) (P<0.05 or P<0.01); the skin thickness scores of mice in CAI ointment dose group 2 were significantly reduced on d10 (3 days after administration) (P<0.01); the skin thickness scores of mice in the positive control group 1 were significantly reduced on d8-d10 (1-3 days after administration) (P<0.05). The skin thickness scores of mice in the positive control group 2 were significantly reduced on d10 (3 days after administration) (P<0.05).

[0192] Table 21

[0193]

[0194] In Table 21, compared with the model group, * indicates P < 0.05, and ** indicates P < 0.01.

[0195] 7.6 cumulative score

[0196] The PASI cumulative scores of the modeling sites of each group of animals at different time points are as follows Figure 5As shown in Table 22 (n=5 or 10), compared with the model group, the PASI cumulative score of mice in CAI ointment dose group 1 was significantly reduced on d9-d10 (2-3 days after administration) (P<0.01); the PASI cumulative score of mice in CAI ointment dose group 2 was significantly reduced on d10 (3 days after administration) (P<0.01); the PASI cumulative score of mice in the positive control group 1 was significantly reduced on d8-d10 (1-3 days after administration) (P<0.01); the PASI cumulative score of mice in the positive control group 2 was significantly reduced on d10 (3 days after administration) (P<0.01).

[0197] Table 22

[0198]

[0199] In Table 22, ** indicates P < 0.01 compared with the model group.

[0200] 7.7 Psoriasis Lesions in Mice

[0201] Skin lesions of mice in each group at different time points Fig.12 As shown, it can be clearly seen that compared with the model group, the skin lesions at the modeling site of mice in CAI ointment dose group 1 were significantly improved on d8 (1 day after administration) and significantly improved on d9-d10 (2-3 days after administration); the skin lesions at the modeling site of mice in CAI ointment dose group 2 were significantly improved on d9 (2 days after administration) and significantly improved on d10 (3 days after administration).

[0202] 7.8 Biochemical index test

[0203] IL-12 concentration detection: The ELISA method was used to detect the IL-12 concentrations of the mouse skin tissue model group, positive control group 1, positive control group 2, CAI ointment dose group 1 and CAI ointment dose group 2.

[0204] Test operation steps:

[0205] (1) After centrifugation of mouse skin tissue samples, the supernatant was taken for ELISA experiment;

[0206] (2) ELISA experiment:

[0207] (2.1) Standard sample addition: Set up standard sample wells and sample wells, and add 50 μL of standard samples of different concentrations to each standard sample well;

[0208] (2.2) Sample addition: Add 50 μL of the sample to be tested to the sample well; do not add to the blank well;

[0209] (2.3) Add enzyme: Add 100 μL of enzyme-labeled reagent to each well except the blank well;

[0210] (2.4) Incubation: Seal the reaction wells with a sealing film and incubate in a 37°C water bath or incubator for 60 min;

[0211] (2.5) Washing: Tear off the sealing film, discard the liquid, pat dry on absorbent paper, fill each well with washing solution (350 μL), let stand for 30 seconds, shake off the washing solution, pat dry on absorbent paper, and repeat this washing process 5 times;

[0212] (2.6) Color development: Add 50 μL of substrate A and B to each well and incubate at 37°C in the dark for 15 min;

[0213] (2.7) Stop: Add 50 μL of stop solution to each well;

[0214] (2.8) Determination: Within 15 min, use an ELISA reader to measure the OD value of each well at a wavelength of 450 nm.

[0215] The test results are shown in Tables 23-26 and Figure 13-Figure 16 Among them, Table 23 is the IL-2 concentration results, Table 24 is the IL-17 concentration results, Table 25 is the IL-23 concentration results, and Table 26 is the TNF-a concentration results. Figure 13-Figure 16 They are bar graphs of IL-12, IL-17, IL-23 and TNF-α concentration results of biochemical index test mice after smearing various drugs of the present invention.

[0216] Table 23

[0217]

[0218]

[0219] Table 24

[0220]

[0221] Table 25

[0222]

[0223]

[0224] Table 26

[0225]

[0226] From the above results, it can be seen that compared with the model group, the carboxyamide triazole ointment treatment group can reduce the concentrations of IL-12 and IL-23 to a certain extent. Compared with the model group, the IL-17 level in the CAI ointment dose 1 group was significantly reduced (P<0.05). Compared with the model group, the TNF-a level in the skin of the CAI ointment dose 1 group was significantly reduced (P<0.05).

[0227] in conclusion:

[0228] (1) General observations: The mental state and activity of animals in each group were normal after administration, and no abnormalities were observed.

[0229] (2) Body weight: The body weight levels of animals in each drug-administered group changed consistently, with a certain degree of decrease during the modeling period (d0-d5). The body weight levels stabilized from the end of the late modeling period to the start of drug administration (d5-d10). There was no significant difference in the body weight levels of animals in each group (P>0.05).

[0230] (3) Erythema score: Compared with the model group, the erythema score of mice in CAI ointment dose group 1 was significantly reduced on d8 (1 day after administration) (P < 0.05), and the erythema score was significantly reduced on d9-d10 (2-3 days after administration) (P < 0.01); the erythema score of mice in CAI ointment dose group 2 was significantly reduced on d10 (3 days after administration) (P < 0.01); the erythema score of mice in the positive control group 1 was significantly reduced on d9-d10 (2-3 days after administration) (P < 0.01). There was no statistical difference in the erythema score of mice in the positive control group 2 compared with the control group (P > 0.05).

[0231] (4) Squamous scale score: Compared with the model group, the squamous scale score of mice in CAI ointment dose group 1 was significantly reduced on d9-d10 (2-3 days after administration) (P < 0.01); the squamous scale score of mice in CAI ointment dose group 2 was significantly reduced on d10 (3 days after administration) (P < 0.01); the squamous scale score of mice in positive control group 1 was significantly reduced on d8-d10 (1-3 days after administration) (P < 0.05 or P < 0.01). The squamous scale score of mice in positive control group 2 was significantly reduced on d10 (3 days after administration) (P < 0.01).

[0232] (5) Skin thickness score: The skin thickness score of mice in CAI ointment dose group 1 was significantly reduced on d9-d10 (2-3 days after administration) (P < 0.05 or P < 0.01); the skin thickness score of mice in CAI ointment dose group 2 was significantly reduced on d10 (3 days after administration) (P < 0.01); the skin thickness score of mice in positive control group 1 was significantly reduced on d8-d10 (1-3 days after administration) (P < 0.05). The skin thickness score of mice in positive control group 2 was significantly reduced on d10 (3 days after administration) (P < 0.05).

[0233] (6) PASI cumulative score: Compared with the model group, the PASI cumulative score of mice in CAI ointment dose group 1 was significantly reduced on d9-d10 (2-3 days after administration) (P < 0.01); the PASI cumulative score of mice in CAI ointment dose group 2 was significantly reduced on d10 (3 days after administration) (P < 0.01); the PASI cumulative score of mice in the positive control group 1 was significantly reduced on d8-d10 (1-3 days after administration) (P < 0.01); the PASI cumulative score of mice in the positive control group 2 was significantly reduced on d10 (3 days after administration) (P < 0.01).

[0234] (7) Animal psoriasis skin lesions: It can be clearly seen that compared with the model group, the skin lesions at the modeling site of mice in CAI ointment dose group 1 were significantly improved on d8 (1 day after administration) and significantly improved on d9-d10 (2-3 days after administration); the skin lesions at the modeling site of mice in CAI ointment dose group 2 were significantly improved on d9 (2 days after administration) and significantly improved on d10 (3 days after administration).

[0235] Test 5 Pathological Detection

[0236] 1. Materials and methods

[0237] 1.1 Samples and Film Reading

[0238] The samples submitted for examination were fixed with 4% paraformaldehyde. After being fixed in good condition, they were trimmed, dehydrated, embedded, sliced, stained, and sealed in strict accordance with the SOP procedures of the unit's pathological laboratory testing, and finally qualified samples were examined under the microscope.

[0239] Browse the slices under a microscope or browse digital slices, observe the tissue slices in detail at different magnifications, describe the basic pathological changes in the slices, such as congestion, congestion, hemorrhage, edema, degeneration, necrosis, hyperplasia, fibrosis, organization, granulation tissue, inflammatory changes, etc., and reflect the differences between slices. Use arrows to mark the imaging of typical lesions.

[0240] 1.2 Image acquisition instrument

[0241] The upright white light photography microscope was purchased from Nikon (Japan), model Eclipse Ci-L.

[0242] 1.3 Evaluation Criteria

[0243] Evaluation criteria are based on: Peter. Mann et al. International Standards for Terminology and Diagnostic Criteria for Pathological Changes in Rats and Mice (INHAND) [M]. Translated by Yang Lifeng, Zhou Xiangmei, and Zhao Deming. Beijing: China Agriculture Press, 2019. Specific evaluation criteria are shown in Table 27 below.

[0244] Table 27

[0245]

[0246] 2. Check the results

[0247] 2.1 Lesion description

[0248] Fig.17a This is a picture of skin tissue pathology of mice in the model group. For the comparison of fixed #22-40 and 200 in the skin model group, small focal necrosis and nuclear fragmentation were occasionally seen in the superficial layer of the epidermis and dermis of the skin tissue, accompanied by a small amount of basophilic inflammatory exudate, thick local epidermis, irregular arrangement of local collagen fibers, and scattered infiltration of many lymphocytes and granulocytes in the dermis. For the comparison of fixed #24-40 and 200 in the skin model group, the dermis of the skin tissue was thin, and the number of accessory organs such as hair follicles and sebaceous glands was small; more epidermal necrosis and shedding, nuclear fragmentation, eosinophilic homogeneity, accompanied by more inflammatory exudate, irregular arrangement of local dermal collagen fibers, and scattered infiltration of many lymphocytes and granulocytes were seen. For the comparison of the fixed #28-40 and 200 in the skin model group, a small amount of epidermal necrosis and shedding, nuclear fragmentation, eosinophilic homogeneity, accompanied by a small amount of inflammatory exudate, dermal collagen fibers are arranged regularly, hair follicles, sebaceous glands and other accessory organs are abundant, and a small amount of lymphocytes are scattered infiltration. For the comparison of the fixed #60-40 and 200 in the skin model group, small focal necrosis and shedding of the epidermis, nuclear fragmentation, eosinophilic homogeneity, accompanied by a small amount of inflammatory exudate, a small amount of prickle cells with loose cytoplasm, irregular cell arrangement, local dermal collagen fibers are arranged irregularly, and a small amount of lymphocytes are scattered infiltration.

[0249] Fig.17b This is a picture of skin tissue pathology of mice in the positive control group 1. For the comparison of fixed #7-40 and 200 of the skin positive control group 1, small focal necrosis of the epidermis, nuclear fragmentation, local thick epidermis, irregular arrangement of local dermal collagen fibers, and scattered infiltration of a small number of lymphocytes and granulocytes can be seen in the skin tissue. For the comparison of fixed #18-40 and 200 of the skin positive control group 1, the epidermis of the skin tissue is intact, the structure is clear, and the keratinized layer can be seen; the dermis has rich collagen fibers and is arranged regularly, and accessory organs such as hair follicles and sebaceous glands can be seen, and a small number of scattered lymphocytes can be seen in the skin tissue. For the comparison of fixed #54-40 and 200 of the skin positive control group 1, small focal necrosis of the epidermis, nuclear fragmentation, local thick epidermis, regular arrangement of dermal collagen fibers, and a small number of scattered lymphocytes can be seen in the skin tissue.

[0250] Fig.17cThis is a picture of skin tissue pathology of mice in the positive control group 2. For the comparison of fixed #38-40 and 200 of the skin positive control group 2, small focal necrosis is occasionally seen in the epidermis and superficial dermis of the skin tissue, the nuclear fragmentation is eosinophilic and homogeneous, accompanied by a small amount of inflammatory exudate, the dermal collagen fibers are arranged regularly, and a small amount of lymphocytes are occasionally scattered infiltration. For the comparison of fixed #44-40 and 200 of the skin positive control group 2, the epidermis of the skin tissue is intact, the structure is clear, and the keratinized layer is visible, and the local epidermis is thick; the dermis is rich in collagen fibers and arranged regularly, and accessory organs such as hair follicles and sebaceous glands can be seen, and a small amount of lymphocytes are occasionally scattered infiltration. For the comparison of the skin positive control group 2 fixed #52-40 and 200, small focal necrosis of the epidermis was occasionally seen in the skin tissue, with nuclear fragmentation, eosinophilic homogeneity, accompanied by a small amount of inflammatory exudate, local epidermis was thick, dermal collagen fibers were arranged more regularly, and a small amount of lymphocytes and granulocytes were occasionally scattered infiltration.

[0251] Fig.17d This is a picture of skin tissue pathology of mice in CAI ointment dose group 1. For the comparison of CAI ointment dose group 1 fixed #31-40 and 200, small focal necrosis and shedding of the epidermis, nuclear fragmentation, eosinophilic homogeneity, accompanied by a small amount of inflammatory exudate, and regular arrangement of dermal collagen fibers. A small amount of lymphocytes and granulocytes can be seen in the skin tissue. For the comparison of CAI ointment dose group 1 fixed #40-40 and 200, a small amount of small focal necrosis and shedding of the epidermis, nuclear fragmentation, eosinophilic homogeneity, accompanied by a small amount of inflammatory exudate, low content of dermal collagen fibers, loose and irregular arrangement, and a small amount of lymphocytes can be seen in the skin tissue. For the comparison of CAI ointment dose group 1 fixed #53-40 and 200, small focal necrosis and shedding of the epidermis, nuclear fragmentation, regular arrangement of dermal collagen fibers, and a small amount of lymphocytes can be seen in the skin tissue. For the comparison of CAI ointment dose group 1 fixed #59-40 and 200, small focal necrosis of the epidermis was occasionally seen in the skin tissue, with nuclear fragmentation, eosinophilic homogeneity, accompanied by a small amount of inflammatory exudate, low content of dermal collagen fibers, loose and irregular arrangement, and a large number of scattered lymphocytes and granulocytes infiltration.

[0252] Fig.17eThis is a picture of skin tissue pathology of mice in two groups of CAI ointment doses. For the comparison of CAI ointment doses #1-40 and 200 in two groups, small focal necrosis, nuclear fragmentation, eosinophilic homogeneity, and a small amount of inflammatory exudate were occasionally seen in the epidermis and superficial dermis of the skin tissue, and the local dermal collagen fibers were irregularly arranged, and the number of accessory organs such as hair follicles and sebaceous glands was abundant, and a small amount of lymphocytes were scattered infiltrates. For the comparison of CAI ointment doses #13-40 and 200 in two groups, small focal necrosis, nuclear fragmentation, eosinophilic homogeneity, and a small amount of inflammatory exudate were occasionally seen in the skin tissue. The local dermal collagen fibers were irregularly arranged, and a small amount of lymphocytes were scattered infiltrates. For the comparison of CAI ointment dose 2 groups #39-40 and 200, the skin tissue occasionally showed small focal necrosis and shedding of the epidermis, nuclear fragmentation, eosinophilic homogeneity, accompanied by a small amount of inflammatory exudate, and the dermal collagen fibers were arranged more regularly, with a small amount of scattered lymphocytes occasionally infiltrating. For the comparison of CAI ointment dose 2 groups #56-40 and 200, the epidermis of the skin tissue was intact, with a clear structure, and the keratinized layer was visible; the dermis had rich collagen fibers and regular arrangement, and accessory organs such as hair follicles and sebaceous glands were visible, with a small amount of scattered lymphocytes occasionally infiltrating. The # in Figure 17 represents the animal number symbol; Figure 17a-Figure 17e The numbers 40 and 200 in the table represent observations at 40 times and 200 times, respectively; Figure 17a-Figure 17e In the figure, at a specific magnification, the unit of the scale bar length in the left figure is 500 μm, and the unit of the scale bar length in the right figure is 100 μm.

[0253] 2.2 Pathological scoring

[0254] The pathological scores are shown in Table 28. Fig.18 As shown, compared with the model group, the pathological scores of animals in CAI ointment dose 1 and 2 groups were significantly decreased (P < 0.05), and the pathological scores of animals in the other groups were not statistically significant (P > 0.05). The # in Table 28 is the experimental animal number and has no special meaning.

[0255] Table 28

[0256]

[0257] Conclusion: Under the experimental conditions, the test substance has a significant therapeutic effect on imiquimod-induced psoriasis mice. It has a significant effect on the changes in erythema, scaling, thickness of the skin lesions in the model mice, and the onset time of the drug effect. The test substance was set up in two dose groups, and both showed that the low dose was more effective.

[0258] Finally, it should be noted that the above content is only used to illustrate the technical solution of the present invention, rather than to limit the scope of protection of the present invention. Simple modifications or equivalent substitutions of the technical solution of the present invention by ordinary technicians in this field do not deviate from the essence and scope of the technical solution of the present invention.

Claims

1. A carboxyamide triazole water-soluble ointment, characterized in that: The invention comprises the following components: carboxyamide triazole and polyethylene glycol; the polyethylene glycol is a mixture of low molecular weight polyethylene glycol and high molecular weight polyethylene glycol, the low molecular weight polyethylene glycol is polyethylene glycol 400, and the high molecular weight polyethylene glycol is selected from at least one of polyethylene glycol 6000 and polyethylene glycol 8000; the weight ratio of the low molecular weight polyethylene glycol to the high molecular weight polyethylene glycol is 59-68:30-39.

2. The carboxyamide triazole water-soluble ointment according to claim 1, characterized in that The composition comprises the following components in parts by weight: 0.5-8 parts of carboxyamine triazole and 92-99.5 parts of polyethylene glycol.

3. The carboxyamide triazole water-soluble ointment according to claim 1, characterized in that The carboxyamide triazole water-soluble ointment also includes an antioxidant component; the components include, by weight: 0.5-8 parts of carboxyamide triazole, 91.9-99.499 parts of polyethylene glycol and 0.001-0.5 parts of antioxidant; the antioxidant is selected from at least one of tocopherol, BHA and BHT.

4. The carboxyamide triazole water-soluble ointment according to claim 1, characterized in that The carboxyamide triazole water-soluble ointment also includes a pH regulator component to adjust the pH value of the carboxyamide triazole water-soluble ointment to 4-9; the pH regulator is selected from at least one of phosphate buffer, citric acid solution, sodium hydroxide solution and hydrochloric acid solution.

5. The carboxyamide triazole water-soluble ointment according to claim 1, characterized in that The carboxyamide triazole water-soluble ointment comprises the following ingredients: carboxyamide triazole, polyethylene glycol, an antioxidant, a pH regulator and a penetration enhancer.

6. The carboxyamide triazole water-soluble ointment according to claim 5, characterized in that The composition comprises the following components in parts by weight: 0.5-3 parts of carboxyamide triazole, 90-98 parts of polyethylene glycol, 2-10 parts of a penetration enhancer and 0.001-0.5 parts of an antioxidant. The pH value of the carboxyamide triazole water-soluble ointment is adjusted to 4-9 by a pH adjuster.

7. The carboxyamide triazole water-soluble ointment according to claim 6, characterized in that The composition comprises the following components by weight: 1-2.5 parts of carboxyamide triazole, 92-93 parts of polyethylene glycol, 2-10 parts of a penetration enhancer, 0.001-0.1 parts of an antioxidant, and a pH regulator to adjust the pH value of the carboxyamide triazole water-soluble ointment to 5-7.

8. The carboxyamide triazole water-soluble ointment according to any one of claims 5 to 7, characterized in that: The polyethylene glycol is a mixture of low molecular weight polyethylene glycol and high molecular weight polyethylene glycol; the low molecular weight polyethylene glycol is polyethylene glycol 400, and the high molecular weight polyethylene glycol is selected from at least one of polyethylene glycol 6000 and polyethylene glycol 8000; the penetration enhancer is selected from at least one of dimethyl sulfoxide, oleic acid, azone and diethylene glycol monoethyl ether; the antioxidant is selected from at least one of tocopherol, BHA and BHT; and the pH adjuster is selected from at least one of phosphate buffer, citric acid solution, sodium hydroxide solution and hydrochloric acid solution.

9. The method for preparing the carboxyamide triazole water-soluble ointment according to any one of claims 1 to 8, characterized in that: Includes steps: (1) mixing part of the low molecular weight polyethylene glycol with all of the high molecular weight polyethylene glycol, heating to melt, stirring, and cooling to obtain a PEG matrix; (2) mixing the carboxyamide triazole with the remaining weight portion of low molecular weight polyethylene glycol and the PEG matrix under heat preservation conditions, and stirring to homogenize and disperse; (3) Stir, cool and bottle.

10. Use of polyethylene glycol 400 and / or polyethylene glycol 6000 and / or polyethylene glycol 8000 in preparing a highly stable carboxyamide triazole water-soluble ointment.

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