CN114886938B

CN114886938B - Composite composition suitable for psoriasis and preparation method thereof

Info

Publication number: CN114886938B
Application number: CN202210483829.6A
Authority: CN
Inventors: 请求不公布姓名
Original assignee: Niten Guangzhou Biotechnology Co ltd
Current assignee: Niten Guangzhou Biotechnology Co ltd
Priority date: 2022-05-05
Filing date: 2022-05-05
Publication date: 2023-07-18
Anticipated expiration: 2042-05-05
Also published as: CN114886938A

Abstract

The invention discloses a composite composition suitable for psoriasis and a preparation method thereof, and belongs to the technical field of medicines. The composition is prepared from caprylic/capric triglyceride, perilla seed oil, ascorbate, cholesterol, white beeswax, ceramide, hydrogenated lecithin and squalane, and forms a pure oil ointment with the functions of epidermal symbiotic bacteria prebiotics and permeation barrier. After the psoriasis patient uses the composition, a layer of stable solid grease film can be formed, the skin permeation physical barrier function is improved, the abundance of beneficial bacteria such as staphylococcus epidermidis, propionibacteria, malassezia and the like in a skin damage area is obviously increased, the abundance of pathogenic bacteria such as staphylococcus aureus, streptococcus, candida and the like is reduced, the microecology on the skin surface is recovered, the skin adaptive immune response is improved, scales and pruritus are obviously reduced, and the severity of psoriasis is reduced. The product is solid at normal temperature, is easy to spread and paint on the skin surface to form a film, and avoids liquefying into oil due to body temperature.

Description

Composite composition suitable for psoriasis and preparation method thereof

Technical Field

The invention belongs to the technical field of medicines, and particularly relates to a composite composition suitable for psoriasis and a preparation method thereof.

Background

Psoriasis is a chronic inflammatory disease, has a long disease course and is easy to recur, and has adverse effects on the physical health and mental condition of patients. According to the statistics of world health organization, about 1.25 hundred million people worldwide suffer from psoriasis, but the cause of the psoriasis is not yet studied clearly.

At present, no method for completely radically curing psoriasis exists, and proper means such as external medicines, physical treatment or systemic medicines are selected to effectively control the disease condition and improve the life quality of patients. According to the psoriasis diagnosis and treatment guide, for localized psoriasis with skin damage less than 3% of body surface area, external medicine treatment can be independently adopted; 80% of patients with clinical psoriasis are mild, the external medicine treatment can relieve the illness state, and 20% of patients are moderate and severe psoriasis, so that the combination treatment is often required. Therefore, the external medicine plays a very important role in psoriasis treatment.

The external medicines which are marketed at present mainly comprise glucocorticoids, coal tar, vitamin A acids, vitamin D3 analogues, calcineurin inhibitors, heat-clearing traditional Chinese medicines and the like. The medicines mainly play roles in resisting inflammation, inhibiting immune response, loosening cutin and the like, have a certain curative effect, but are easy to relapse, have more adverse reactions after long-term use, and are urgently needed to be marketed as a brand new medicine with safer and definite curative effect.

At present, more and more researches find that the surface symbiotic bacteria of the skin lesion area of psoriasis are disordered, the abundance of staphylococcus aureus, streptococcus and candida is increased, the abundance of staphylococcus epidermidis, propionibacterium and malassezia is obviously reduced, the staphylococcus aureus, streptococcus and candida have superantigens, can induce strong induction of Th17 reaction, generate cytokines such as IL-17A, IL-17F, IL22 and the like, cause abnormal T cell immune reaction of skin, and cause skin lesion and keratinization abnormality. Beneficial bacteria such as staphylococcus epidermidis, propionibacteria, and malassezia can decompose triglyceride in sebum, generate short chain fatty acid and other bacterial metabolites, and regulate CD ₄ ⁺ T cells maintain skin adaptive immune homeostasis. Additional studies have found that reduced sebaceous gland content in psoriatic plaques is an important factor in bacteria that cause skin commensal bacteria disorders.

The loss of sebum, keratosis, and keratosis in the skin lesions of psoriasis patients directly lead to the disruption of the skin's epidermal permeability barrier. Abnormal epidermal penetration barrier in turn leads to abnormal proliferation and differentiation of keratinocytes, leading to keratinization; abnormal permeation barrier can also cause loss of skin moisture or infiltration of moisture and foreign matters in the environment, moisture infiltration causes hydration of cutin, and the barrier function of the foreign matters is reduced; abnormal permeation barrier causes antigen invasion, facilitates migration of skin immune cells, and simultaneously provides convenience for migration and aggravates immune response. Many factors interweave, the penetration barrier is destroyed to enter the vicious circle, and finally the skin is thickened and dried, and the disease is difficult to heal.

However, no drugs for improving the symbiotic bacteria of the epidermis of the skin and improving the permeation barrier are currently marketed.

Disclosure of Invention

The invention aims to solve the technical problems that the abnormal conditions of skin symbiotic bacteria disorder, epidermis permeation barrier damage and the like of psoriasis patients cannot be improved and the healing effect is poor in the prior art, and provides a composite composition suitable for psoriasis and a preparation method thereof.

The technical scheme adopted by the invention is as follows: a composite composition suitable for psoriasis, the composition being formed by mixing prebiotic oils, permeation barrier oils and melting point regulating oils;

the prebiotic oil is prepared from the following raw materials in parts by weight: 39-60 parts of caprylic/capric triglyceride, 0.4-4 parts of perilla seed oil and 3-15 parts of ascorbate;

the permeability barrier grease is prepared from the following raw materials in parts by mass: 7-15 parts of cholesterol, 19-36 parts of white beeswax, 0.1-1 part of ceramide and 0.1-2 parts of hydrogenated lecithin;

the melting point regulating grease is 2-9.5 parts of squalane.

Preferably, the prebiotic oil is prepared from the following raw materials in parts by weight: 45-52 parts of caprylic/capric triglyceride, 1.5-3 parts of perilla seed oil and 5-13 parts of ascorbate;

preferably, 48 parts of caprylic/capric triglyceride, 2.9 parts of perilla seed oil and 8 parts of ascorbate.

Preferably, the permeability barrier grease is prepared from the following raw materials in parts by mass: 9-12 parts of cholesterol, 23.5-31 parts of white beeswax, 0.4-0.9 part of ceramide and 0.5-1.7 parts of hydrogenated lecithin;

preferably, the permeability barrier grease is prepared from the following raw materials in parts by weight: 10 parts of cholesterol, 26 parts of white beeswax, 0.5 part of ceramide and 1.2 parts of hydrogenated lecithin.

Preferably, the melting point regulating grease is 4-9 parts of squalane;

preferably, the melting point adjusting oil is squalane 6 parts.

Preferably, the ascorbate comprises one or more of ascorbyl palmitate, ascorbyl dipalmitate, ascorbyl methylsilanol pectate, ascorbyl tocopheryl maleate, ascorbyl tetraisopalmitate, ascorbyl stearate, ascorbyl tetrahexyldecanol, and aminopropanol ascorbyl phosphate;

ascorbyl palmitate is preferred.

Preferably, the ceramide is ceramide phosphorylcholine and/or ceramide phosphorylethanolamine.

Preferably, the hydrogenated lecithin is soybean hydrogenated lecithin.

A method of preparing a complex composition suitable for psoriasis, the method comprising:

weighing caprylic/capric triglyceride, perilla seed oil, ascorbate, cholesterin, white beeswax, ceramide, hydrogenated lecithin and squalane according to the corresponding mass fractions, mixing, stirring and cooling.

Preferably, the mixing and stirring conditions are as follows:

stirring at 400-500r/min and 130-150deg.C for 2-3min.

Preferably, the cooling temperature of each raw material after mixing and stirring is 40-50 ℃.

The beneficial effects of the invention are as follows:

the invention provides a composite composition suitable for psoriasis, which is formed by combining caprylic/capric triglyceride, perilla seed oil, ascorbyl ester, cholesterol, white beeswax, ceramide, hydrogenated lecithin and squalane. The composition is a grease ointment preparation for external use.

Wherein, the caprylic acid/capric triglyceride is the main component of sebum and is also the main nutrient substance of the symbiotic bacteria on the body surface, and the perilla seed oil and the ascorbate have the function of inhibiting the proliferation of pathogenic bacteria and are also the nutrient substance of the symbiotic bacteria on the body surface. The mixture of caprylic acid/capric acid triglyceride, perilla seed oil and ascorbate produces the optimal effect of skin symbiotic bacteria prebiotics, obviously increases the abundance of probiotics such as staphylococcus epidermidis, propionibacterium, lipophilic malassezia and the like in the skin damage area of psoriasis, and reduces the abundance of harmful bacteria such as staphylococcus aureus, streptococcus, candida and the like.

White beeswax is waxy with good sealing property, strong barrier effect, high melting point, close to the waxy property in epidermis, stable chemical property, high viscosity, high brittleness and insufficient plasticity; cholesterol is the main component of the cell membrane of the organism, has high melting point, stable chemical property, good adhesion with the skin horny layer, and poor viscosity and film forming property; ceramide is a matrix between skin keratinocytes, has high melting point, stable chemical property and good adhesion with skin horny layer, and has the defects of high viscosity and poor film formation; hydrogenated lecithin is a component of cell membranes, has a high melting point, high chemical stability and good air permeability, and has the disadvantages of poor sealing property and poor barrier property. The 4 kinds of grease are compounded, make up for the advantages of high melting point, stability, sealing property, plasticity and skin affinity, good adhesion with skin and cutin, easy filling between loose cutin when being smeared on the surface of psoriasis skin damage area, stable solid grease film forming, and playing the best penetration physical barrier function (barrier function), not only slowing down the penetration of moisture, but also blocking the loss of moisture in the skin, and also blocking the invasion of other foreign matters including pathogenic bacteria and other antigens.

Squalane is stable liquid oil, has high skin affinity, good air permeability, easy spreading, no greasiness, low melting point, and no solidification below-38deg.C. The test composition detects that when the physical characteristic melting point of the oil-mixed product is in the range of 45-65 ℃, the ointment is easy to smear and form a film, the film has good stability at the body surface temperature, is not easy to liquefy, and has good use feeling. Therefore, the product selects squalane as the melting point of the mixed grease, and adopts proper dosage according to the proportion of other raw materials.

And (II) the formulation is built through a plurality of experiments, so that the pure oil ointment with the functions of the epidermal symbiotic bacteria prebiotics and the permeation barrier is formed. After bathing, phototherapy, daily morning and before sleeping, a proper amount of ointment is smeared on the skin damage part of a psoriasis patient, a layer of stable solid grease film can be formed, the skin permeation physical barrier (barrier) function is improved, the abundance of beneficial bacteria such as staphylococcus epidermidis, propionibacteria, malassezia and the like in the skin damage area can be obviously increased, the abundance of pathogenic bacteria such as staphylococcus aureus, streptococcus, candida and the like is reduced, the microecology of the skin surface is recovered, the skin adaptive immune response is improved, the scales and the pruritus are obviously reduced, and the severity of psoriasis is reduced.

The composition is used for solving the defects that the existing external preparation is easy to relapse and has more adverse effects after long-term use mainly through pharmacological actions such as anti-inflammatory, immune response inhibition and keratolytic action, and is specially used for improving the stable state of skin symbiotic bacteria, improving adaptive immunity, supplementing sebum, improving the skin permeation barrier function and the like aiming at abnormal conditions such as 'skin symbiotic bacteria disorder', 'skin permeation barrier destruction' and the like existing at the skin damage part of a psoriasis patient, and can remarkably reduce scales and pruritus, delay relapse and reduce the severity of psoriasis.

(IV) the product of the application belongs to a brand new psoriasis treatment pharmaceutical preparation, and has the following beneficial effects compared with the prior art: 1) Can improve symbiotic bacteria disorder at skin injury part of psoriasis, increase the abundance of probiotics such as staphylococcus epidermidis, propionibacterium, lipophilic malassezia, etc., reduce the abundance of harmful bacteria such as staphylococcus aureus, streptococcus, candida, etc., and improve T cell immune response; 2) The grease ointment can be smeared on the skin damage surface of psoriasis to form a stable solid grease film with good air permeability, supplements sebum of sebum at the skin damage part of psoriasis, plays a role of permeation physical barrier (barrier), can reduce the permeation and loss of moisture, and can also block the invasion of other foreign matters including pathogenic bacteria and other antigens; 3) The product of the invention has obvious differences from the existing external psoriasis medicines on the market in terms of the raw materials of the prescription and the treatment mechanism, can obviously reduce psoriasis scales and pruritus, delay relapse and reduce the severity of psoriasis; 4) The product contains no chemical surfactant (emulsifier), preservative, pigment, essence and water. Use of emollients under dry skin conditions: consensus declaration (2013): the emulsifier-containing product can aggravate skin itch and reduce skin permeability barrier; the antiseptic, pigment and essence are all used as foreign matters to aggravate itching and symptoms of psoriasis. The moisture can swell the loose horny layer of the psoriasis skin, aggravate the damage of the skin barrier function; 5) The mixed grease of the product is solid at normal temperature, is easy to spread and smear on the surface of skin to form a film, prevents the formation of oil due to body temperature liquefaction, prevents greasy feeling, and adopts raw materials with higher affinity with the skin, so that the product has better use feeling than the pure grease product on the market, and achieves the effect of grease without greasy feeling.

Drawings

FIG. 1 is a diagram of experimental examples for the study of the D6 skin lesions of a psoriatic mouse;

FIG. 2 shows the results of experimental investigation of skin lesions pathology in psoriatic mice.

Detailed Description

The invention will be further illustrated with reference to specific examples. It will be appreciated by those skilled in the art that the embodiments described below are some, but not all, of the embodiments of the present invention and are merely illustrative of the present invention and should not be construed as limiting the scope of the present invention. The reagents used were all conventional products commercially available.

Example 1:

the formula is as follows: 45 g of caprylic/capric triglyceride, 2.9 g of perilla seed oil, 15 g of ascorbyl palmitate, 7 g of cholesterol, 19 g of white beeswax, 0.1 g of ceramide phosphorylcholine, 2g of soybean hydrogenated lecithin and 9 g of squalane;

weighing the raw materials according to the corresponding mass, adding the raw materials into mixing and stirring equipment, continuously stirring for 2min under the condition that the temperature is gradually increased to 130 ℃ at the rotating speed of 400-500r/min, cooling and stirring to 50 ℃, and discharging to obtain the grease ointment with the melting point of 50-55 ℃.

Example 2:

the formula is as follows: caprylic/capric triglyceride 60 g, perilla seed oil 0.5 g, ascorbyl dipalmitate 3 g, cholesterol 15 g, white beeswax 19 g, ceramide phosphoethanolamine 0.4 g, soybean hydrogenated lecithin 0.1 g and squalane 2 g;

weighing the raw materials according to the corresponding mass, adding the raw materials into mixing and stirring equipment, continuously stirring for 3min under the condition that the temperature is gradually increased to 150 ℃ at the rotating speed of 400-500r/min, cooling and stirring to 40 ℃, and discharging to obtain the grease ointment with the melting point of 45-51 ℃.

Example 3:

the formula is as follows: 52 g of caprylic/capric triglyceride, 4 g of perilla seed oil, 5 g of ascorbyl methylsilanol pectate, 10 g of cholesterol, 23.5 g of white beeswax, 1 g of ceramide phosphorylcholine, 0.5 g of soybean hydrogenated lecithin and 4 g of squalane;

weighing the raw materials according to the corresponding mass, adding the raw materials into mixing and stirring equipment, continuously stirring for 2min under the condition that the temperature is gradually increased to 130 ℃ at the rotating speed of 400-500r/min, cooling and stirring to 50 ℃, and discharging to obtain the grease ointment with the melting point of 45-52 ℃.

Example 4:

the formula is as follows: 45 g of caprylic/capric triglyceride, 4 g of perilla seed oil, 5 g of tocopheryl ascorbate maleate, 7 g of cholesterol, 36 g of white beeswax, 0.5 g of ceramide phosphoethanolamine, 0.5 g of soybean hydrogenated lecithin and 9.5 g of squalane;

weighing the raw materials according to the corresponding mass, adding the raw materials into mixing and stirring equipment, continuously stirring for 3min under the condition that the temperature is gradually increased to 150 ℃ at the rotating speed of 400-500r/min, cooling and stirring to 40 ℃, and discharging to obtain the grease ointment with the melting point of 51-56 ℃.

Example 5:

the formula is as follows: 39 g of caprylic/capric triglyceride, 0.4 g of perilla seed oil, 13 g of tetraiso-palmitate ascorbate, 10 g of cholesterol, 31 g of white beeswax, 0.9 g of ceramide phosphorylcholine, 1.7 g of soybean hydrogenated lecithin and 4 g of squalane;

weighing the raw materials according to the corresponding mass, adding the raw materials into mixing and stirring equipment, continuously stirring for 2min under the condition that the temperature is gradually increased to 130 ℃ at the rotating speed of 400-500r/min, cooling and stirring to 50 ℃, and discharging to obtain the grease ointment with the melting point of 62-65 ℃.

Example 6:

the formula is as follows: 45 g of caprylic/capric triglyceride, 1.5 g of perilla seed oil, 5 g of ascorbyl stearate, 9 g of cholesterol, 23.5 g of white beeswax, 0.4 g of ceramide phosphoethanolamine, 0.5 g of soybean hydrogenated lecithin and 4 g of squalane;

weighing the raw materials according to the corresponding mass, adding the raw materials into mixing and stirring equipment, continuously stirring for 3min under the condition that the temperature is gradually increased to 130 ℃ at the rotating speed of 400-500r/min, cooling and stirring to 40 ℃, and discharging to obtain the grease ointment with the melting point of 48-54 ℃.

Example 7:

the formula is as follows: 52 g of caprylic/capric triglyceride, 3 g of perilla seed oil, 13 g of tetrahexyldecanol anti-ascorbate, 12 g of cholesterol, 31 g of white beeswax, 0.9 g of ceramide phosphorylcholine, 1.7 g of soybean hydrogenated lecithin and 9 g of squalane;

weighing the raw materials according to the corresponding mass, adding the raw materials into mixing and stirring equipment, continuously stirring for 2min under the condition that the temperature is gradually increased to 150 ℃ at the rotating speed of 400-500r/min, cooling and stirring to 50 ℃, and discharging to obtain the grease ointment with the melting point of 53-59 ℃.

Example 8:

the formula is as follows: 48 g of caprylic/capric triglyceride, 2.9 g of perilla seed oil, 8 g of aminopropanol anti-ascorbic acid phosphate, 10 g of cholesterol, 26 g of white beeswax, 0.5 g of ceramide phosphoethanolamine, 1.2 g of soybean hydrogenated lecithin and 6 g of squalane;

weighing the raw materials according to the corresponding mass, adding the raw materials into mixing and stirring equipment, continuously stirring for 3min under the condition that the temperature is gradually increased to 150 ℃ at the rotating speed of 400-500r/min, cooling and stirring to 40 ℃, and discharging to obtain the grease ointment with the melting point of 50-56 ℃.

Based on the formulation of example 3, a control was designed

Comparative example 1:

the formula is as follows: 52 g of caprylic/capric triglyceride, 4 parts of olive oil and 5 parts of ascorbyl palmitate; 23.5 parts of white beeswax, 10 parts of cholesterol, 0.5 part of hydrogenated lecithin, 1 part of ceramide and 4 parts of squalane;

weighing the raw materials according to the corresponding mass, adding the raw materials into mixing and stirring equipment, continuously stirring for 2min under the condition that the temperature is gradually increased to 130-150 ℃ at the rotating speed of 400-500r/min, cooling and stirring to 50 ℃, and discharging to obtain a finished product.

To verify the efficacy of perilla seed oil, the present comparative example 1 was obtained by replacing perilla seed oil with olive oil in example 3.

Comparative example 2:

the formula is as follows: 57 parts of caprylic/capric triglyceride and 4 parts of perilla seed oil; 10 parts of cholesterol, 23.5 parts of white bee wax, 1 part of ceramide, 0.5 part of hydrogenated lecithin and 4 parts of squalane;

To verify the efficacy of ascorbate, ascorbate in example 3 was eliminated to give comparative example 2.

Comparative example 3:

the formula is as follows: 52 parts of caprylic/capric triglyceride, 4 parts of perilla seed oil and 5 parts of ascorbate; 10 parts of cholesterol, 23.5 parts of white beeswax, 1 part of ceramide and 4 parts of squalane;

To verify the efficacy of the hydrogenated lecithin, the hydrogenated lecithin material in example 3 was removed to obtain comparative example 3.

Comparative example 4:

the formula is as follows: 52 parts of caprylic/capric triglyceride, 4 parts of perilla seed oil and 5 parts of ascorbate; 10 parts of cholesterol, 23.5 parts of white beeswax, 0.5 part of hydrogenated lecithin and 4 parts of squalane;

To verify the efficacy of ceramide, ceramide material in example 3 was removed to obtain comparative example 4.

Comparative example:

"A sebum membrane-like component and its preparation method" example 1 "in the data, as a comparative example:

the raw materials comprise the following components in percentage by weight: 60 parts of triglyceride (15 parts of sun-sunflower seed oil, chicory seed oil, butter fruit, and grape seed oil respectively), 23 parts of wax (5 parts of Brazil wax palm acid wax, 10 parts of beeswax, 8 parts of candelilla wax), 12 parts of squalane, and 5 parts of dihydrocholesterol.

Weighing wax and dihydrocholesterol, premixing at 150deg.C, stirring at 400-500r/min, cooling to 90deg.C, slowly adding the rest materials, stirring to 30deg.C, and discharging to obtain 30 deg.C liquid final product, and making into ointment with melting point of 12-18deg.C.

Experimental example:

study one: evaluation of therapeutic Effect of psoriasis imiquimod cream mouse model

SPF grade C57BL/6J mice, male, 20-22g quality, purchased from Beijing Wanfukang Biotechnology Co., ltd, animal qualification number SCXK (Beijing) 2014-0004, imiquimod cream (Sichuan Mingxin pharmaceutical Co., ltd); peroxidase blocking solution, DAB color development solution and fluorescent blocking agent (Zhonghua gold bridge); anti-CD 3 antibodies (Abcam), alexa488 goat anti-rabbit IgG (Guanzhong technology Co., ltd.), BCA kit (Thermo), tissue dehydrator (Leica-ASP 300S), embedding machine (Leica-EG 1140C) orthofluorescence microscope and image analysis system (Zeiss-IMAGERZ 2).

The literature [ vander Fits L, mourits S, voerman JS, et al, iminomodificated PSORIASIS-like skin inflammation in mice is mediated via the IL-23/IL-17axis.J Immunol,2009,182 (9): 5836-5845 imiquimod induces a classical model of psoriasis-like skin lesions mice were anesthetized with intraperitoneal injection of 80mg/kg sodium pentobarbital, and dorsal scrape Mao Yao cm×2cm.

The randomization was divided into 6 groups: blank control group; a model group; example 3 (experimental group); comparative example 1; comparative example 2; comparative example group. 5 animals are fed in a single cage in each group. 62.5mg of 5% imiquimod cream was administered topically 1 time daily except for the blank. After 6 days, mice were sacrificed. The administration mode is as follows: the imiquimod cream is smeared with 50mg of paste to be detected for 2h and 12h, and is continuously administrated for 5 days.

And (3) observing the indexes:

1) Skin lesion performance and PASI score in mice: mice were photographed daily for skin lesions and evaluated for severity scores based on erythema, scaling and infiltration, respectively, in terms of 3 aspects of erythema, infiltration and scaling, and rated 5 according to severity: no score of 0, mild score of 1, moderate score of 2, severe score of 3, very severe score of 4, total score of 12.

2) Detection of skin epidermis moisture content of mice: before drawing materials, the skin moisture content of the skin lesions of the backs of the mice in each group is detected by a water-oil pen, repeated for 3 times, and an average value is obtained.

3) Pathological changes of skin lesions in mice the tissues were fixed, dehydrated, and then prepared into 5 μm paraffin sections for HE staining. The epidermis thickness was measured using a ZEN image analysis system for pictures under a 40-fold objective.

4) Immunohistochemical assay detects infiltration of mouse skin lesions cd3+ T cells:

after dewaxing and antigen retrieval of paraffin sections, peroxidase blocking solution was blocked at room temperature for 10min,10% goat serum was incubated at 37℃for 30min, and when the addition of lorecrin (1:50) I antibody was incubated overnight at 4 ℃. After incubation with goat anti-rabbit IgG antibody for 1h at 37 ℃ the plate was developed. Infiltration of cd3+ T cells was observed under an overhead light microscope.

Study results:

1) Skin lesions of psoriasis-like mice show:

as shown in fig. 1, the skin of the back of the mice in the blank group was smooth and had a reddish flesh color. Along with the smearing of imiquimod, the skin lesions of mice in the model group show psoriasis-like manifestations of erythema, scales, infiltration and the like.

TABLE 1 PASI score (mean value) for skin lesions in psoriatic mice

Grouping	The next day	Third day	Sixth day
				Blank control	0	0	0
Model group	3	6	11
				Example 3	2	3	2*
Comparative example 1	3	4	5*△
				Comparative example 2	3	5	6*△
Comparative example	3	6	9△

Note that: * : after rank sum test, compared with the model group, the model group has significant difference (p is less than 0.05); delta: after rank sum test, there was a significant difference (p < 0.05) compared to example 3.

As shown in table 1, each group of mice showed erythema, scaling, infiltration, etc. on day 2 of dosing compared to the model group; the mice in each group developed a tendency to differ in PASI score 3 days after dosing.

2) Skin damage part moisture content and skin damage thickness

As shown in fig. 2, pathological observation can see that the skin epidermis of the mice in the blank group on the 6 th day is thin, the dermal lymphocyte infiltration is less, the epidermis of the mice in the model group is obviously thickened (P < 0.05), the epidermis is not fully keratinized, and the dermis is infiltrated by a large amount of lymphocytes; the other groups had thinner epidermis and less dermal lymphocyte infiltration than the model group; the skin thickness of the comparative example was thicker (P < 0.05) than that of the comparative example 1-2, and the skin thickness of the example 3 was significantly lower than that of the comparative example 1-2 (P < 0.05).

TABLE 2 skin moisture content and skin loss thickness

Grouping	Skin moisture content of skin injury part (%)	Skin thickness (mum)
			Blank control	34.65±1.02	12.34±0.25
Model group	17.24±0.89	89.36±6.41
			Example 3	33.26±0.93*	13.95±0.47*
Comparative example 1	29.74±1.59*	31.01±2.83*△
			Comparative example 2	28.38±1.16*	33.79±3.84*△
Comparative example	18.43±1.42	78.42±7.35△

Note that: * : compared with the model group, there was a significant difference (p < 0.05); delta: compared with example 3, there was a significant difference (p < 0.05).

As shown in table 2, the water content of the epidermis was significantly reduced (P < 0.05) on day 6 in the model mice compared to the blank group, suggesting that the epidermal permeation barrier was compromised; the epidermis water content of the mice in other groups is increased compared with the model group.

3) T lymphocyte activated infiltration of psoriasis-like mice skin lesions

TABLE 3 count of CD3+ T cell activation at the skin lesion site in psoriatic mice

Grouping	CD3+T (number of positive cells)
		Blank control	1.41±0.37
Model group	13.42±1.95
		Example 3	3.06±0.24*
Comparative example 1	7.38±2.03*△
		Comparative example 2	7.57±2.93*△
Comparative example	12.03±1.76△

As shown in table 3, CD3 marks activated T lymphocytes in the skin, with positives appearing brown. In the blank control group, a small amount of brown positive particles are visible in dermis, and the positive points of the model group are obviously increased (P < 0.05); the cd3+ T cells were significantly reduced (P < 0.05) in the example 3 group compared to the model group, and the control 1-2 groups were also reduced, but not to the same extent as in example 3, the control group being comparable to the model group.

The results of the above study show that:

1) The effects of example 3 (with caprylic capric triglyceride, perilla seed oil, and ascorbate) and control 1 (with caprylic capric triglyceride, without perilla seed oil) and control 2 (with caprylic capric triglyceride, without ascorbate) were very remarkable, and the effects were also somewhat remarkable, differing from those of example 3, for the improvement of symptoms (PASI score) such as psoriasis scales, erythema, and the like. The perilla seed oil and the ascorbate cooperate with caprylic acid and capric acid triglyceride to have therapeutic effects on psoriasis, and the combination of the three components generates synergistic effects with the best effect;

2) The detection result of the water content of the psoriasis skin damage area shows that: example 3 (with caprylic capric triglyceride, perilla seed oil and ascorbate) and comparative example 1 (with caprylic capric triglyceride, no perilla seed oil) and comparative example 2 (with caprylic capric triglyceride, no ascorbate) can significantly improve skin moisture content (reflect skin permeation barrier), but the effect among the three is not significantly different, which indicates that the influence of perilla seed oil and anti-ascorbate on skin permeation barrier function is less;

3) The detection results of the thickness of the skin lesion area of psoriasis, inflammatory cell infiltration, CD3T cell positive and the like show that: example 3 (caprylic capric triglyceride, perilla seed oil, and ascorbate) significantly improved the inflammatory response in the psoriatic lesions. The effect of the composition was not as good as that of example 3 (p < 0.05), but it was demonstrated that the synergistic effect of the combination of the perilla seed oil and the ascorbate with caprylic-capric triglyceride was effective in suppressing inflammatory immune response, as compared with that of comparative example 1 (caprylic-capric triglyceride without perilla seed oil) and comparative example 2 (caprylic-capric triglyceride without ascorbate).

4) The comparative examples have weak improvement effects on PASI score, skin lesion area water content, epidermis thickness, inflammatory cell infiltration, CD3T positive cells and the like, and cannot be used as external preparations for psoriasis;

in a word, the product of the invention has definite curative effect as an external preparation for psoriasis, and the caprylic/capric triglyceride, the perilla seed oil and the ascorbyl ester are indispensable key raw materials in the raw materials of the product of the invention.

Study two: evaluation of improving permeable skin Barrier function

Evaluation of skin permeation Barrier function of mice model of imiquimod cream for psoriasis

Mice were modeled as mice with imiquimod cream psoriasis, and were divided into 6 groups of blank, model, example 3, control 4, comparative example, etc., 10 mice per group, and single-cage rearing per group.

The barrier effect of the skin lesion site on water penetration was examined.

The administration method comprises the following steps: each group was prepared by taking 5, except for the blank control group, 62.5mg of 5% imiquimod cream for external use, and the other groups except for the model group were respectively coated with 50mg of the corresponding test cream (the ointment prepared in example 3 or comparative example 3/4 or comparative example) on the corresponding group after 2 hours of coating the imiquimod cream, covering the skin damage area with a water saturated cotton cloth strip after 1 hour, fixing with an adhesive tape, performing continuous intervention 1 time daily for 5 days, and dying at 6 days.

Examining the barrier effect of the skin damage part on imiquimod penetration, and the administration method comprises the following steps: each group was given 5, except the blank control group, 50mg of test cream was first administered, 62.5mg of 5% imiquimod cream was applied after 1h, 1 time daily, 5 days of continuous intervention, and the animals were sacrificed on day 6.

And (3) observing the indexes: PASI scoring and skin moisture and skin thickness measurements were performed on skin lesions of mice; the modeling method and PASI score, skin moisture content, and skin thickness measurements were the same as "study one". The research results show that:

1) Blocking barrier effect of psoriasis lesions on water penetration

TABLE 4 skin loss PASI score, moisture content and thickness (mean) of psoriatic mice

As shown in table 4, the skin lesions had a weak permeation barrier to moisture in the environment, and the skin moisture content and skin lesion thickness of the model group were significantly increased, with a higher PASI score. Whereas example 3 was able to block water penetration, reduce the PASI score, the skin thickness was closest to the blank, and the improvement of control examples 3-4 was weaker than example 3, but stronger than the control and model groups.

2) Penetration barrier effect of psoriasis skin lesion part on imiquimod cream

TABLE 5 skin loss PASI score, moisture content and thickness of psoriatic mice

As shown in table 5, the PASI score of the model group was significantly increased, the skin moisture content was significantly reduced and the skin lesion thickness was significantly increased compared to the blank group, whereas the PASI, skin moisture content and thickness were significantly improved by applying the imiquimod cream of example 3 first and then. The improvement effect of comparative examples 3-4 was weaker than that of example 3, but stronger than that of the comparative example and model group.

The results of the above study two show that:

1) The effect of example 3 (with hydrogenated lecithin and ceramide, white beeswax, cholesterol) was very remarkable, and the effect of comparative example 3 (without hydrogenated lecithin, with ceramide, white beeswax, cholesterol) and comparative example 4 (without ceramide, with hydrogenated lecithin, white beeswax, cholesterol) were also remarkable, differing from that of example 3. The hydrogenated lecithin and the ceramide cooperate with each other to have the synergistic effect of the hydrogenated lecithin and the ceramide cooperate with the white beeswax and cholesterol to treat psoriasis, and the combination of the hydrogenated lecithin and the ceramide has the synergistic effect with the best effect;

2) The detection result of the water content of the psoriasis skin damage area shows that: example 3 (with hydrogenated lecithin and ceramide, white beeswax, cholesterol) and comparative example 3 (without hydrogenated lecithin, with ceramide, white beeswax, cholesterol) and comparative example 4 (without ceramide, with hydrogenated lecithin, white beeswax, cholesterol) can significantly block the penetration of external moisture and imiquimod (reflect skin permeation barrier), but comparative example 3 and comparative example 4 have significant differences from example 3, which means that hydrogenated lecithin and ceramide cooperate with white beeswax and cholesterol to improve skin permeation barrier function, and the four have the best synergistic effect;

3) The comparative examples have weaker effects on PASI score, skin lesion area water penetration blocking and imiquimod penetration, and cannot be used as external preparations for psoriasis;

in a word, the product of the invention has definite curative effect as an external preparation for psoriasis, and hydrogenated lecithin, ceramide, white beeswax and cholesterol are indispensable key raw materials in the raw materials of the product of the invention.

Study three: functional evaluation study affecting skin' epidermosymbiotic

Evaluation of efficacy evaluation of skin "epidermotoga" affecting the mice model of imiquimod cream for psoriasis

Culturing skin surface symbiotic bacteria, namely standard strains staphylococcus aureus ATCC25923, candida albicans ATCC10231, streptococcus pyogenes ATCC19615, staphylococcus epidermidis ATCC12228, propionibacterium ATCC6919 and salad bacterium ATCC44344, wherein the standard bacteria are all purchased from Barceicosis organisms, culturing for 20 hours by adopting nutrient broth, and mixing bacterial solutions of each bacteria with the same volume to prepare a composite bacterial solution.

Mice models of imiquimod cream psoriasis were established, and animals were divided into 7 groups of blank, model, example 3, comparative examples 1-3, comparative example, etc., 5 animals per group, and single-cage rearing per group. 62.5mg of 5% imiquimod cream was administered topically 1 time daily except for the blank. After the imiquimod cream is smeared for 4 hours on the first day, 1ml of smeared composite bacterial liquid is taken out by a cotton swab. Day 2-6, the imiquimod cream was applied 2 hours later, 60mg of test cream was applied once a day, and sacrificed on day 7.

And (3) observing the indexes: before the ointment to be measured is smeared on the skin lesion surface on the 2 nd and sixth days, a microbial sample is taken for gene sequencing (Meiger V1V 9) by using a cotton swab, and various bacterial abundances are analyzed. Performing PASI scoring and skin moisture content and skin damage thickness detection on skin damage of the mice before applying the paste to be detected on the 2 nd and 6 th days; the other operations were the same as "efficacy evaluation study 1".

Three results of the study:

1) Analysis of abundance of 'epidermosymbiotic bacteria' at skin injury part of psoriasis mice

TABLE 6 epidermal symbiotic bacteria abundance of psoriatic mice skin lesions

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Note that: * : compared to the same group of D2 bacterial abundances, D6 bacterial abundances were significantly different (p < 0.05) 2) in the psoriatic mouse skin lesion site PASI score, moisture content and skin lesion thickness.

As shown in table 6, the change in bacterial abundance on the sixth day was examined with reference to the bacterial abundance on the second day. The abundance of staphylococcus aureus, candida and streptococcus is obviously increased after the skin damage of the model group and the comparison example group is generated, the abundance of staphylococcus epidermidis, propionibacterium, mala salad bacteria and the like is obviously reduced, and both the example 3 and the comparison examples 1-3 have prebiotic effects, wherein the effects of the example 3 and the comparison example 3 are stronger than those of the comparison examples 1-2; no prebiotic effect was demonstrated for the comparative examples.

TABLE 7 skin lesion PASI score, moisture content and skin lesion thickness in psoriatic mice

As shown in table 7, the PASI score was significantly increased, the skin moisture content was significantly reduced and the skin lesion thickness was significantly increased for the model and comparative examples compared to the blank, while the PASI, skin moisture content and thickness were significantly improved for example 3 and comparative examples 1 and 2, wherein example 3 was more effective than comparative examples 1-2; the effect of comparative example 3 and comparative example was poor.

The three results of the study showed that:

1) The symbiotic bacteria in the skin damage area of the psoriasis in the example 3 and the control 3 (both containing perilla seed oil and ascorbate and caprylic-capric triglyceride) have the strongest prebiotic effect, and the control 1 (without perilla seed oil and containing ascorbate and caprylic-capric triglyceride) and the control 2 (without ascorbate and containing perilla seed oil and ascorbate and caprylic-capric triglyceride) also have certain prebiotic effect, and the control has no prebiotic effect. The purple perilla seed oil, the ascorbyl ester and the caprylic/capric triglyceride are combined as key raw materials with prebiotic effect, and the prebiotic function of the three raw materials used in combination is optimal.

2) In aspects of water content in skin lesion areas of psoriasis and the like, the effect of the example 3 is very remarkable with the comparative examples 1-2 (with hydrogenated lecithin and ceramide, cholesterol and white beeswax), and the effect of the comparative example 3 (without hydrogenated lecithin, with ceramide, cholesterol and white beeswax) on preventing water loss is reduced, which indicates that the hydrogenated lecithin is very important for permeability barrier recovery.

3) In terms of psoriasis lesion thickness and PASI score, example 3 had very significant effects compared to comparative examples 1-2, and comparative example 3 (containing perilla seed oil, ascorbate, no hydrogenated lecithin) had poor effects. It is explained that the function of 'epidermolysis symbiotic prebiotic' and the function of 'skin permeation barrier' are the key to ensure the treatment of psoriasis.

In a word, the product of the invention is used as the external preparation of psoriasis to treat the curative effect definitely, and among the raw materials of the product of the invention, raw materials such as hydrogenated lecithin, ceramide, perilla seed oil, ascorbate, cholesterol, white beeswax, caprylic/capric triglyceride, etc. are all indispensable key raw materials of the product of the invention.

Study IV: evaluation of melting Point of Mixed fat on skin feel in use

The ointments of Table 8 were evaluated for skin feel by following the table below to prepare a number of formulations of different melting points (see Table 8). The testing method for crowd assessment comprises the following steps: 18-45 years old volunteers were selected for 30 cases. The test period was six weeks, daily, and weekly evaluation of an ointment was performed by: a proper amount of sebum membrane-like component is smeared on the back of a hand to experience skin feel, and the evaluation result of the crowd is shown in Table 9.

TABLE 8 raw material ratios and melting Point ranges for different greases

The evaluation of the skin feel index was made by classification. The specific grading standards for skin feel after use are as follows:

stage 1: the skin has no adverse reaction after use, does not dry, and is fresh and moisture-keeping.

2 stages: the skin has no adverse reaction after use, is not very dry and does not feel moist.

3 stages: the skin has no adverse reaction after use, and is greasy or dry.

TABLE 9 evaluation of skin feel after use of greases of different melting points

Evaluation index

Ointment 1

Ointment 2

Ointment 3

Example 3

Example 5

Ointment 7

Skin feel after use

3 grade

Level 2

Level 1

Level 2

The number of people

30 people

The results of the study show that the melting point of the mixed oil and fat is adjusted to be in the range of 45-65 ℃ so as to optimize the use feeling.

In a word, the product of the invention has the effects of the symbiotic bacteria of epidermis in the skin lesion area of psoriasis, improves the permeation barrier function, improves the symptoms such as itch, scales, erythema and the like, reduces the activation of T cells, reduces the recurrence and reduces the severity of psoriasis, and is a brand-new external preparation for treating psoriasis. In the formula of the product, caprylic acid and capric triglyceride, perilla seed oil and ascorbate are key raw materials for playing a prebiotic function of the product, white beeswax, cholesterol, ceramide, hydrogenated lecithin and the like are key raw materials for playing a permeation physical barrier function, and squalane is an important raw material for adjusting the melting point of mixed grease and improving the skin affinity of a solid grease film. The therapeutic effect of the product for treating psoriasis can be ensured as long as the key raw materials are regulated and prepared within the scope of the application, so that the invention still belongs to the protection scope of the invention by adding a small amount of other auxiliary components (including spices and other animal and vegetable oils) on the basis of the proportion of the key raw materials and the important raw materials.

The invention is not limited to the above-described alternative embodiments, and any person who may obtain other products in various forms under the teaching of the invention falls within the scope of protection of the invention. The above detailed description should not be construed as limiting the scope of the invention, and it should be understood by those skilled in the art that the technical solutions described in the foregoing embodiments may be modified or some or all of the technical features may be replaced equally without departing from the scope of the technical solutions of the embodiments of the invention.

Claims

1. A composite composition suitable for psoriasis, characterized in that the composition is mixed by prebiotic oil, penetration barrier oil and melting point regulating oil;

the prebiotic oil is prepared from the following raw materials in parts by weight: 52 parts of caprylic/capric triglyceride, 4 parts of perilla seed oil and 5 parts of ascorbic acid methylsilanol pectate;

the permeability barrier grease is prepared from the following raw materials in parts by mass: 10 parts of cholesterol, 23.5 parts of white beeswax, 1 part of ceramide phosphorylcholine and 0.5 part of soybean hydrogenated lecithin;

4 parts of melting point regulating grease;

the physical melting point of the composition is 45-52 ℃.

2. A method of preparing a composite composition suitable for psoriasis according to claim 1, comprising:

weighing caprylic/capric triglyceride, perilla seed oil, ascorbyl methyl silanol pectate, cholesterol, white beeswax, ceramide phosphorylcholine, soybean hydrogenated lecithin and squalane according to the corresponding mass fraction, mixing and stirring for 2-3min under the conditions that the rotating speed is 400-500r/min and the temperature is increased to 130-150 ℃, and cooling to 40-50 ℃.