CN112915198B

CN112915198B - An ointment containing lysozyme for treating psoriasis

Info

Publication number: CN112915198B
Application number: CN202110389695.7A
Authority: CN
Inventors: 潘宏涛; 卢亚萍
Original assignee: Zhejiang Aegis Biological Science & Technology Co ltd
Current assignee: Zhejiang Aegis Biological Science & Technology Co ltd
Priority date: 2016-03-11
Filing date: 2016-03-11
Publication date: 2022-05-24
Anticipated expiration: 2036-03-11
Also published as: CN105770869B; CN105770869A; CN112915198A

Abstract

The invention relates to a pharmaceutical composition for treating psoriasis, wherein the weight ratio of lysozyme to emodin as active ingredients is (1:10) - (10: 1). Compared with the common western medicine preparation, the lysozyme preparation provided by the invention has the advantages that the preparation method is simple, the compatibility is reasonable, the two active ingredients can generate a synergistic interaction effect while exerting respective effects, and the psoriasis is remarkably treated. The lysozyme and the emodin are natural in source, safe and non-toxic, and have no side effect; because the pharmaceutical composition contains no western medicine components, the medicine has no drug resistance.

Description

An ointment containing lysozyme for treating psoriasis

Technical Field

The invention relates to the field of pharmacy, in particular to a medicine for treating psoriasis.

Background

Psoriasis is also called psoriasis, is a skin disease with scales and erythema as clinical symptoms, and is accompanied by certain pruritus. Psoriasis is not infectious, but has high incidence rate, long course of disease, easy relapse, stubborn and difficult radical cure, has great influence on the physical health and spirit of patients, and is called as immortal cancer in dermatology.

The cause of psoriasis is not clear, modern studies suggest that it is associated with genetic, infectious, immunological, psychiatric, endocrine, traumatic, dietary and metabolic disorders, and so on, and immune mediation is now considered to be the major mechanism of occurrence. The clinical manifestations of the medicine are erythema scaling damage, early erythema pimple, layer upon layer of silvery white scale covering the epidermis, dry skin, desquamation and incrustation; some skin symptoms are connected into a sheet, and the sheet is like a map, and some skin symptoms are pruritus, pus flowing water and blood stain; psoriasis can be classified into the ordinary type, erythroderma type, abscess type and arthropathy type, with the ordinary type being the most common. The blood-heat syndrome skin rash in the traditional Chinese medicine is mainly pimple and maculopapule, new rash appears continuously, the color of the basal skin is bright red, scales are scraped off, punctate bleeding is caused, pruritus with different degrees can be caused, and the method is equivalent to the advanced stage of psoriasis in western medicine.

Psoriasis is the most common intractable disease of skin, the incidence rate of psoriasis in natural population is about 0.1% -3%, in China, the incidence rate is higher in the south in the north, higher in the city than in the countryside, and higher in the female in the male. According to incomplete statistics, the number of the psoriasis patients in China is more than 200 ten thousand. In recent years, the number of urban population in China is increased sharply, and with the addition of about 2000 ten thousand new population, according to epidemic research statistics, about 10 ten thousand psoriasis patients are increased each year in China. According to the data, the market space of the psoriasis medicine in China is about 20 hundred million yuan.

Methotrexate (MTX) is one of the most effective drugs for the treatment of psoriasis at present, and is effective in severe psoriasis. In the tretinoin, the abamectin is the 2 nd generation oral tretinoin which is the in vivo metabolite of abamectin A ester, the curative effect is equivalent to that of the abamectin A ester, but the biological activity is stronger, the half-life period is shorter, and the long-term in vivo accumulation adverse reaction is smaller. It binds to retinoic acid receptor, and has effects of treating psoriasis by regulating proliferation and differentiation of keratinocyte, anti-inflammatory action, anti-microvascular formation, etc. Cyclosporine is a potent immunosuppressant, acts mainly on pathogenic T cells in the skin lesion proliferative phase of psoriasis, and is effective on all types of psoriasis.

With the remarkable progress of the research on the pathogenesis of psoriasis and the great success of cyclosporine A in the treatment of psoriasis, new researches at present consider that T cells are a central link for maintaining the activity of psoriasis diseases. In recent years, it has been found that certain cytokines play an important role in the development of psoriasis, including interleukin 12, interleukin 23 and tumor necrosis factor secreted by helper T-cells 17 (Th-17). Interleukins 12 and 23 activate natural killer cells, T cells and CD4 lymphocytes, promoting differentiation of the latter into helper T cells 7 and 17. Helper T cell 1 produces inflammatory cytokines, primarily tumor necrosis factor. Helper T cell 17 produces different subsequent effects and various pro-inflammatory cytokines, and plays an important role in the formation and maintenance of psoriasis lesions. The current immune biologies for psoriasis are mainly classified as follows:

1. biological agents that inhibit T cell activation and inhibit proliferation of activated T cells: the alfacamide can eliminate the number of local and circulating pathological T cells, is firstly approved to be used for treating chronic moderate and severe plaque psoriasis, acts on human fusion protein of CD antigen on T cells, is used for blocking stimulation signals, and leads the stimulation signals to selectively induce and activate apoptosis of memory T cells, thereby achieving the aim of treating the psoriasis. Efaviruses are humanized anti-CD 11a mabs that bind to the surface molecule lfa.1 of T cells, block the secondary signal of T cell activation and T cell migration to peripheral skin tissues, and treat psoriasis by preventing T lymphocyte activation and its entry into the skin from the circulatory system.

2. Cytokine balance regulating drugs: the recombinant human interleukin 10(IL-10) is an important immunoregulation cell factor produced by various cells of a human body, has wide biological activities including immunosuppressive, anti-inflammatory and immunoregulation properties, has the main biological functions of limiting and stopping inflammatory reaction, regulating differentiation and proliferation of various immune cells, and also has the property of regulating immune stimulation and eliminating infected and non-infected particles. Etanercept is produced by linking the extracellular portion of the human Tumor Necrosis Factor (TNF) receptor P57 to the Fc fragment of IgG1, and is able to bind to and inactivate soluble TNF-D. In recent years, TNF has been found to be one of the most important cytokines in the pathogenesis of psoriasis. The food and drug administration of the united states was originally approved for rheumatoid arthritis and later officially for the treatment of psoriatic arthritis. The composition is suitable for moderate and severe plaque psoriasis, and can be used alone or in combination with methotrexate.

Modern Chinese medicine experts classify the ordinary psoriasis into 4 types of blood heat type, blood deficiency type, blood dryness type and blood stasis type according to the characteristics of skin lesions; in the specific type of psoriasis, the pustule type is generally classified as sepsis type, the erythroderma type is toxic heat type, and the joint type is cold-dampness type or wind-damp obstruction type. Blood heat type: is often found in the run period. It is usually manifested as rapid development of skin damage, continuous appearance of new rash, bright red rash, dripping (or spot-like) rash, excessive scales, severe pruritus, and punctate hemorrhage after scraping off scales. For heat-clearing and blood-cooling, herbs with heat-clearing and blood-cooling combined with toxicity-removing actions are usually used, such as Bai Mao Gen, Chi Shao, Fu Ling, Dan Pi, Sheng Huai Hua and Pu Gong Ying, etc. Blood dryness type: it is mostly seen in the resting stage. It is manifested as a long course of disease, few new eruptions, pale red rash, coin-like or large-piece fusion, more infiltrative thickening, less scales, pale tongue, thin white or little coating, deep, thready or wiry and thready pulse. For it is suitable to cool blood and moisten dryness, it is usually combined with spatholobus stem, clematis root, Chinese angelica root, dried rehmannia root, root of red rooted saliva, etc. Blood stasis type: it is common to see the resting stage. Clinically, the skin damage is persistent, large, fat, dark red and oyster shell-like, and the scales are white and dry, often accompanied by abdominal distension and dysmenorrhea, or arthralgia, failure to straighten, dark tongue with ecchymosis or petechia, and unsmooth pulse. For treatment, it is advisable to activate blood and resolve stasis, dispel wind and dredge collaterals. The Chinese medicines are selected from peach kernel and Chinese angelica root which have the functions of promoting blood circulation and nourishing blood, common burreed rhizome and zedoary which have the functions of promoting blood circulation and removing blood stasis, softening and resolving hard mass, clematis root, centipede, earthworm and the like which have the functions of dispelling wind, dredging collaterals and resolving hard mass. Blood deficiency type: it is mostly seen in the retirement phase. If the skin lesion is not cured for a long time, qi and yin will be damaged, and qi and blood will be deficient. The skin turns dark red after being dry, and although there is no new eruption of the rash, the surface of the skin is stained with a large amount of silvery white dry scales which are gradually removed from the skin layer by layer and gradually reduced. For it is suitable to nourish blood, dispel wind and moisten dryness, it is usually combined with qi and blood tonics such as Dang Gui, Shu Di Huang, Shi Hu and so on, and yin-nourishing and dryness-moistening herbs such as Tian Dong, Mai Dong, Xuan Shen and Huang Jing.

Psoriasis is chronic aeipathia, has a long course of disease, and is taken for a long time by a relatively fixed method to control and relieve the development of the disease. The commonly used method is based on the basic recipe, modified according to the syndrome. For blood heat type, it is advisable to clear heat and remove toxicity, cool blood and activate blood, and it is modified with blood-cooling and activating decoction (Baimaogen, Zicao, Sheng Di Huang, ban lan Gen, Sheng Gao, Chi Shao). For blood stasis type, it is advisable to activate blood and resolve stasis, move qi and soften hard mass, and the formula is modified with blood-activating and stasis-resolving decoction (red sage root, Chinese angelica root, Ligusticum wallichii, peach kernel, safflower, burred tuber, zedoary, bitter orange, dried orange peel and astragalus root). For blood dryness type, it is advisable to nourish yin and blood, activate blood and moisten dryness, modified blood-nourishing and toxicity-removing decoction (Dang Gui, He shou Wu, Huang Jing, Dan Shen, Ji Teng, Sheng Di, Shu Di, Tian Dong, Mai Dong).

Today, antipsoriatic drugs have been directed to natural drugs, synthetic drugs and biologics. Compared with the traditional medicine for treating psoriasis, the traditional Chinese medicine which is characterized by flexible medication and small adverse reaction and the biological preparation which is characterized by small adverse reaction and no immunosuppressive action provide an effective and safe scheme for treating psoriasis, and have wide clinical application prospect. In China, natural medicines, especially active parts of Chinese herbal medicines, and semisynthetic products thereof are developed, so that the Chinese herbal medicine has greater advantages and characteristics.

At present, the medicines for treating psoriasis on the market are mainly western medicine preparations, and the medicines are easy to generate drug resistance after long-term use. Therefore, it is necessary to develop a drug with natural source, no toxic and side effects and no drug resistance to replace western medicine preparations.

Lysozyme (also called muramidase) or N-acetylmuramidase hydrolase (N-acetylmuramidase glycohydrolase) is an alkaline enzyme that hydrolyzes mucopolysaccharides in pathogenic bacteria. The bacterial lysis is achieved by breaking the beta-1, 4 glycosidic bond between N-acetylmuramic acid and N-acetylglucosamine in the cell wall, breaking down the cell wall insoluble mucopolysaccharide into soluble glycopeptides, causing the contents of the broken cell wall to escape. Lysozyme can also be directly combined with virus protein with negative charge, and forms double salt with DNA, RNA and apoprotein to inactivate virus. Therefore, the enzyme has the functions of antibiosis, antiphlogosis, antivirus, etc. Lysozyme (also called lytic enzyme) is an enzyme with a molecular weight of 14.4kDa, which is capable of destroying the bacterial cell wall by catalyzing the hydrolysis of the 1, 4-beta chain between the N-acetylmuramic acid and the N-acetylglucosamine residues in peptidoglycans and between the N-acetylglucosamine residues in chitodextrins. In humans, lysozyme is widely found in cellular exudates, such as saliva, tears, and other body fluids; also present in the cytoplasmic granules in mitochondria. In addition, egg white also contains a large amount of lysozyme. Lysozyme can be used in food, feed, medicine and other industries because of its specific action on the basic hydrolase of cell wall of target microbe.

Lysozyme (lysozyme) is a pure basic globulin consisting of 18 129 amino acids and has a molecular weight of 14.4 KDa. The chemical properties are very stable. When the pH value is changed sharply within the range of 1.2-11.3, the structure is almost unchanged. In an acid environment, the lysozyme has strong thermal stability; when the pH value is 4-7, the activity of the proenzyme can still be maintained after treatment for 1min at 100 ℃; when the pH value is 3, the coating can resist heating treatment at 100 ℃ for 45 min; under the dry condition, the lysozyme can be stored at room temperature for a long time, and the pure product is white or yellowish. Yellow crystal or amorphous powder, no odor, and sweet taste. Is easily soluble in water, easily destroyed by alkali, and insoluble in acetone and diethyl ether. As a medical product, the lysozyme has the advantages of good biocompatibility, no irritation to tissues, no toxicity and the like.

Lysozyme is widely present in egg white of poultry and birds and tears, saliva, plasma, urine, milk, leukocytes and other body fluids and tissue (e.g., liver, kidney) cells of mammals; the lysozyme can also be separated from barley, fig, pawpaw, cabbage, radish and other plants, wherein the content of the lysozyme is the highest (about 0.3 percent) in egg white. The lysozyme activity in human milk, tears and saliva is much higher (about 3 times) than that in egg white or other sources. The lysozyme in egg white is a typical representative of animal lysozyme, is an important research object in lysozyme group, and is one of the best known lysozyme at present. In mammalian milk, the lysozyme content in human milk is 3000 times higher than that of cow's milk. According to different sources, the lysozyme can be divided into egg white lysozyme, animal lysozyme, plant lysozyme, lysozyme produced by microorganisms and lysozyme produced by bacteriophages; depending on the cell wall, lysozyme can be classified as bacterial cell wall lysozyme and fungal cell wall lysozyme.

Lysozyme is capable of specifically lysing the cell wall of the bacteria of interest. It mainly attacks peptidoglycans, hydrolyzing the beta-1, 4 glycosidic bond connecting the fourth carbon atom of N-acetylmuramic acid and N-acetylglucosamine. The whole process is that lysozyme is bound to the peptidoglycan molecule through the "groove" between its two domains; the substrate then forms a transition state conformation in the enzyme. Lysozyme binds to the glucan hexasaccharide according to the Phillips mechanism. Lysozyme then twisted the fourth sugar on the dextran hexaose into a half-chair conformation. In this distorted state (higher energy), the glycosidic bond is easily broken. The side chains of glutamic acid (Glu35) at position 35 and aspartic acid (Asp52) at position 52 of the lysozyme protein sequence are critical to lysozyme activity. Glu35 acts as a proton donor for glycosidic bonds, cleaving the C-O bond of the substrate; asp52 is involved in the formation of the glycosylase intermediate as a nucleophile. Subsequently, the glycosylase intermediate reacts with water molecules and is hydrolyzed to produce the product, while the enzyme remains unchanged.

The extraction method of the lysozyme comprises the following steps: 1. direct crystallization method: the method is simple to operate and low in cost, but cannot be used for separating trace lysozyme. 2. Ion exchange chromatography: the method has the advantages of simplicity, convenience, high efficiency, low cost and automatic continuous operation. 3. Affinity chromatography: the affinity chromatography has high separation and purification times and good quality, and can be widely applied to lysozyme from various sources and used for concentrating and refining trace lysozyme. However, the complex preparation of affinity adsorbent limits its industrial large-scale application. 4. Ultrafiltration combined with affinity chromatography: the combination of ultrafiltration and affinity chromatography for extracting lysozyme is a new method developed in recent years. Firstly, removing most of impure protein by ultrafiltration, retaining lysozyme, and obtaining a refined product by affinity chromatography. 5. Membrane chromatography technique: at the end of the 20 th century, 80 s, affinity membrane chromatography technology emerged. The technology has the characteristics of membrane separation and affinity separation. Compared with the traditional membrane separation and affinity chromatography, the membrane chromatography technology has the characteristics of high purification multiple, small pressure drop, short separation time, small variability probability of biomacromolecules in the separation process, high feeding speed allowance and the like, and is easier to realize large-scale production compared with the column affinity chromatography. 6. And (3) ultrafiltration: the major advantage of ultrafiltration technology over traditional biochemical separation techniques is the high yield of the product. However, despite the widespread use of ultrafiltration in reverse dialysis and concentration processes, ultrafiltration is still underutilized as a protein separation technique with great potential in the field of biological industry. 7. Reverse micelle extraction: the reverse micelle extraction is a new extraction method developed in recent years, which is a method for extracting reverse micelles by adding a small amount of surfactant into an organic solvent, develops a new separation technology for some bioactive substances such as enzyme, active protein and the like which cannot be extracted by the organic solvent, and expands the application range of the organic solvent extraction. 8. The biological separation technology comprises the following steps: affinity-based bioseparation techniques (immobilized metal affinity chromatography, affinity precipitation, affinity filtration) are also one of the lysozyme extraction methods.

Lysozyme has several unique and important pharmacological actions.

First, it is antibacterial and anti-inflammatory. The bacterial cell wall is composed of murein, which is a polymer composed of N-acetylglucosamine and N-acetylmuramic acid, and polypeptide, called peptidoglycan, can be connected to murein residue. Polysaccharides exist in a linear form, and polysaccharide chains adjacent to each other may be connected to each other via peptide chain moieties to form a three-dimensional structure. The lysozyme acts on the glycosidic bond between the N-acetylmuramic acid and the N-acetylglucosamine of the peptidoglycan molecule specifically, so that the cell wall of the bacteria becomes loose, the protection effect on the cells is lost, and finally the cells are dissolved and die; the lysozyme can directly kill gram-positive bacteria, and can also kill gram-negative bacteria under the participation of secretory immunoglobulin complement: such as Escherichia coli. The lysozyme can be combined with various acidic substances which induce inflammation, reduces the release of bacterial endotoxin, thereby relieving endotoxemia, slowing the inflammation process, having a certain synergistic effect on antibiotics, improving mucopolysaccharide metabolism of tissue matrix, and playing the roles of resisting bacteria, diminishing inflammation and repairing tissues. Experiments prove that the antibacterial ability of lysozyme can be obviously improved and the wound healing can be accelerated by coupling the human lysozyme and the proteolytic enzyme by using 1 cellulose copolymer.

Second, it is antiviral. The lysozyme has a large number of positive charges in the neutral body fluid environment, can directly act with virus proteins with negative charges, and is combined with DNA, RNA and desorption base proteins to form double salts, so that viruses are inactivated. The lysozyme is added into the culture solution of the Hela cells infected by the herpes virus to inhibit cell degeneration. Lysozyme can also inhibit the growth of adenovirus, and can be used for treating viral diseases such as herpes zoster, parotitis, chicken chickenpox, hepatitis and influenza.

Third, immunity is enhanced. The lysozyme participates in various immune reactions in the body, and plays an important role in the normal defense and the non-specific immunity of the body. It can improve and enhance phagocytic digestion function of macrophage, reduce leucopenia caused by cytostatic agent, combine with bacterial lipopolysaccharide, and relieve endotoxin, so as to achieve the purpose of enhancing body resistance. Lysozyme has a T cell epitope per se and can induce a type 2 (Th2) T helper cell response. The oral lysozyme can induce mice to generate systemic Th1 and Th2 immune response and enhance the immune function, so the oral lysozyme has the functions of controlling upper respiratory tract infection and treating dysentery. In addition, the lysozyme also has the function of activating blood platelets, can improve local blood circulation disorder of tissues, decompose pus and enhance the local defense function, thereby showing the effects of stopping bleeding, reducing swelling and accelerating the repair of damaged tissues of the body. It can also be used as a host resistance factor to protect tissue locally.

Researchers have accumulated a great deal of experience in the application of lysozyme in clinical trials. For example, the drug susceptibility test uses 3 drugs of compound lysozyme, fluconazole and nystatin as test drugs, studies the bacteriostatic effect of the test drugs on isolated candida albicans, and measures the MIC value of the drugs by NCCLSM27-A microdilution. Antifungal susceptibility testing of the three drugs showed that: the minimum inhibitory concentrations of the compound lysozyme, the fluconazole and the nystatin against the separated candida albicans are 2.03 mu g/ml, 3.19 mu g/ml and 6.03 mu g/ml respectively; the sensitivity of the candida albicans to the compound lysozyme is greater than that of fluconazole and nystatin; the observation and research of clinical curative effect show that: the effective rate of the compound lysozyme for treating the oral candidiasis of the children is 97.5 percent, which is obviously higher than that of a control nystatin group; the average treatment time of the compound lysozyme group is 6.46 days, which is obviously lower than 13.0 days of the control nystatin group; the research result shows that: candida albicans has the highest sensitivity to complex lysozyme, greater than fluconazole and nystatin. The compound lysozyme is used for treating children oral candidiasis, and has short treatment course and high cure rate. The antifungal in vitro drug sensitivity result of the compound lysozyme is consistent with the clinical curative effect, which shows that the drug sensitivity has certain correlation with the clinical curative effect. In addition, researches show that the symptoms of the infantile acute sinusitis can be quickly disappeared in about 14 days under the assistance of lysozyme, the nasal mucosa can be recovered to be normal, the healing time of the infantile acute sinusitis of a control group is obviously longer than that of a research group, and the lysozyme has a high-efficiency auxiliary antibacterial function; the research group and the control group have significant difference in cure rate and significant efficiency, and the lysozyme is proved to have significant acceleration of the cure speed of the infantile acute nasosinusitis, has no adverse reaction and has certain clinical popularization and application values. Because the lysozyme has good curative effect on the prevention and treatment of the diseases of the five sense organs, the lysozyme is on the market at present in China, and the lysozyme has wide application prospect in the treatment of the diseases of the five sense organs.

Rhubarb is a traditional Chinese medicine in China and is distributed in Shanxi, Sichuan, Hubei, Yunnan provinces and the like. Rhubarb, radix et rhizoma Rhei, the most commonly used medicine in clinic since ancient times, is bitter and cold in nature and enters spleen, stomach, large intestine, liver and pericardium channels. The record of Shen nong Ben Cao Jing: mainly aims at removing blood stasis, stopping blood, stopping cold and heat, breaking the symptoms and abdominal mass, remaining retained food, washing intestines and stomach, promoting aging to cause novelty, smoothing water and removing food, regulating middle energizer and resolving food, and harmonizing five internal organs. Compendium of materia Medica is called: da Huang can be indicated for dysentery with reddish and white discharge, tenesmus, abdominal pain, dribbling urine, excess heat and dryness, tidal fever and delirium, jaundice and other fire sores.

Emodin (emodin) has chemical name of 1,3, 8-trihydroxy-6-methylanthraquinone (1,3, 8-trihydr-oxy-6-methyl lanthraquinone) and molecular weight of 270.23. The chemical structure of the compound belongs to the hydroxy anthraquinone class, is a main effective monomer of the Chinese medicinal rhubarb, and has various pharmacological actions.

Emodin has good antibacterial effect. The emodin can effectively inhibit bacteria such as staphylococcus aureus, escherichia coli, shigella flexneri and the like. Staphylococcus aureus is less susceptible to resistance to emodin, streptococci are also sensitive to it, and emodin is also effective against diphtheria, hay, anthrax, paratyphoid, dysentery, large intestine, Bacillus influenzae, pneumococcus and catacoccus. The anti-anaerobic experiment shows that the emodin has strong inhibition effect on 100 clinically common anaerobic bacteria, the 8mg/ml can inhibit the growth of 76-91% of the anaerobic bacteria, and the most common bacteroides fragilis can inhibit the growth of 90-100% of the strains. Emodin can inhibit the growth of helicobacter pylori and the DNA damage caused by the helicobacter pylori in a dose-dependent manner. There are studies showing that emodin inhibits bacterial growth by effectively inhibiting bacterial collagenase.

The research reports that emodin 30 and 60mg/kg can inhibit the rat plantar swelling caused by carrageenan after the mice are orally taken for 1 time, the rat plantar swelling caused by carrageenan can be inhibited after the rats are orally taken for 60 and 100mg/kg, the inhibition effect is increased along with the increase of the dosage, the capillary permeability of the mice can be inhibited from increasing caused by acetic acid after the mice are orally taken for 1 time, and the exudation and the leukocyte migration of the rat acute pleurisy caused by the carrageenan can be inhibited by intraperitoneal injection of 20 and 40 mg/kg. The mechanism may be related to inhibition of NO synthesis. The research finds that the emodin can powerfully inhibit the expression of nuclear transcription factors and adhesion molecules, so that the emodin can treat various inflammations.

Emodin has effects of resisting oxidation and scavenging oxygen free radicals. Emodin has quenching effect on ultra-weak chemiluminescence caused by lipid peroxidation of rat brain homogenate. The mechanism of emodin for inhibiting lipid peroxidation is to inhibit the propagation of lipid peroxidation free radicals in a mitochondrial membrane, and the structure-activity relationship indicates that the substitution of ortho-position hydroxyl and meso-position hydroxyl of the emodin is necessary for the antioxidation of the substances.

Modern studies have found that emodin effects on guinea pig isolated gut tube are dose related, with contraction increasing with increasing dose below 29.16 μmol/L and decreasing to cessation of effect above 29.16 μmol/L. The appropriate dose of emodin (14.18. mu. mol/L) resulted in a marked increase in the contractile effects of acetylcholine (Ach) on guinea pig isolated ileum and colon compared to Ach alone, but increasing the dose of emodin reduced the contractile effects rather until the activity ceased. At this time, 0.127mmol/L calcium chloride is added to restore the effect of Ach on contracting intestinal canal smooth muscle. The experiment shows that the emodin has bidirectional regulation effect on the guinea pig in-vitro intestinal canal and is closely related to the participation of calcium ions. Research shows that the emodin has a bidirectional regulation effect on guinea pig, rat and rabbit isolated ileum, but the contraction frequency of the rabbit isolated ileum cannot be changed, the effect of the emodin on increasing the contraction amplitude of the ileum can be antagonized by atropine with low dose, and the emodin is proved to have similar acetylcholine-like effect but weak effect. The emodin is injected into the abdominal cavity, so that the aspirin can be inhibited from being combined with gastric ulcer of a rat caused by pylorus ligation, the concentration of gastric acid and gastric albumin is reduced, gastric mucus secretion is promoted, and the protein content in gastric juice is increased.

Research shows that emodin can reduce CCl₄And D-galactosamine induced liver damage, plays a role in protecting the liver, and histopathological detection shows that emodin can reduce lymphoid cellsCell, Kupffer cell, reduction of balloon-like changes, cell necrosis, and vitreoid changes. With CCl₄Primary culture rat liver cells damaged in vitro are used as an experimental model, and the influence of emodin on acute liver cell damage is observed. The results show that emodin is used for CCl₄The damaged primary culture rat cells have obvious protection effect on CCl₄The level of serum glutamic-pyruvic transaminase (ALT), glutamic-oxalacetic transaminase (AST) and lipid peroxide malonodiacetic acid (MDA) is improved, so that the effect of obviously inhibiting the serum glutamic-pyruvic transaminase (ALT), the serum glutamic-oxalacetic transaminase (AST) and the lipid peroxide malonodiacetic acid (MDA) is achieved, and the proliferative activity of damaged liver cells is obviously changed. Researchers also find that emodin can obviously reduce glutamic acid transaminase and alkaline phosphatase, obviously increase total protein albumin, obviously reduce serum hyaluronic acid and laminin, obviously reduce the content of collagen in liver tissues, obviously improve the fibrosis degree of the liver tissues and reduce the expression of alpha-muscle protein in the liver tissues.

The research shows that emodin causes apoptosis of 3 liver cancer cell lines of malignant liver cancer cell lines of Mahlavu PLC/PRF/5 and HepG2 by inducing increase of Reactive Oxygen Species (ROS), reduction of mitochondrial transmembrane potential and activation of Caspase. Research shows that the emodin has strong growth inhibition effect on human liver cancer cell strain smmc-7721, and flow cytometry, DNA gel electrophoresis and fluorescent staining methods prove that the emodin can induce the cell to generate G2/M phase retardation to die. Emodin can also inhibit proliferation of lung adenocarcinoma cells, inhibit growth of colon cancer by inhibiting Vascular Endothelial Growth Factor Receptor (VEGFR) tyrosine kinase activity, and can be used as an effective tumor angiogenesis inhibitor. The emodin and the astragalus polysaccharide which are jointly administered have certain prevention effect on the experimental liver cancer model of the rat. Emodin was also found to inhibit tumor cell migration by inhibiting the PI 3-kinase-Cdc 42/Rac1 pathway. Recent studies have also demonstrated that emodin down-regulates the male hormone receptor to inhibit prostate cancer cells, promotes apoptosis in human squamous cell lung carcinoma cells (CH27) and human non-small cell lung carcinoma cells (H460), probably by reducing the expression of protein kinase c (pkc).

At present, the application of the combination of lysozyme and emodin for preventing and treating psoriasis is not reported.

Disclosure of Invention

The invention aims to solve the technical problem of providing a pharmaceutical composition which can effectively prevent and treat psoriasis, has no toxic or side effect and drug resistance and has natural active ingredients.

The technical scheme adopted by the invention for solving the problems is to provide the pharmaceutical composition for treating psoriasis, wherein the weight ratio of lysozyme to emodin as active ingredients is (1:10) - (10: 1).

Preferably, in the pharmaceutical composition, the weight ratio of the lysozyme to the emodin is (1:1) - (1: 9).

More preferably, the weight ratio of lysozyme to emodin is 1: 3.

Preferably, the lysozyme is extracted from egg white.

Preferably, the pharmaceutical composition further comprises pharmaceutically acceptable auxiliary materials.

More preferably, the pharmaceutically acceptable auxiliary materials are selected from one or more of emulsifying agents, grease, chelating agents, moisture absorbing agents, thickening agents and purified water;

most preferably, the emulsifier is tween 60 or tween 80, the grease is lanolin or vaseline, the chelating agent is ethylenediamine tetraacetic acid or disodium ethylenediamine tetraacetic acid, the hygroscopic agent is talcum powder or zinc oxide, and the thickening agent is sodium carboxymethylcellulose or xanthan gum.

Preferably, the dosage form of the pharmaceutical composition is ointment, lotion, aerosol, liniment, and the like.

More preferably, the pharmaceutical composition is in the form of an ointment.

The invention also provides application of the pharmaceutical composition in preparing a medicament for treating psoriasis.

The invention has the positive and beneficial effects that:

surprisingly, through repeated experiments, the invention unexpectedly discovers that the combination of lysozyme and emodin has a synergistic effect on the prevention and treatment of psoriasis, and the effect is better when the lysozyme and the emodin are matched with other active ingredients. Compared with western medicine preparations, the lysozyme preparation provided by the invention has the advantages that the preparation method is simple, the compatibility is reasonable, the two active ingredients can generate a synergistic interaction effect while exerting respective effects, and the psoriasis can be remarkably treated. The lysozyme and the emodin are natural in source, safe and non-toxic, and have no side effect; the medicine composition has no western medicine component, so that the medicine has no drug resistance.

Detailed Description

The present invention will be further described with reference to the following examples, but the embodiments of the present invention are not limited thereto. The experimental procedures used in the following examples are all conventional procedures unless otherwise specified. The lysozyme used in examples and experimental examples was lysozyme extracted from egg white.

Example 1 preparation of an ointment containing emodin

Weighing 10 parts by weight of emodin, 50 parts by weight of sodium carboxymethylcellulose, 10 parts by weight of zinc oxide and 1 part by weight of disodium ethylene diamine tetraacetate, dispersing in 500 parts by weight of purified water, and uniformly stirring to obtain a water phase; weighing 50 parts by weight of vaseline and 10 parts by weight of tween 80 in another container, and mixing to obtain an oil phase; heating the water phase and oil phase respectively, mixing, homogenizing for 10min, cooling, and making into ointment A1 containing emodin.

Example 2 preparation of an ointment containing lysozyme

Weighing 10 parts by weight of lysozyme, 50 parts by weight of sodium carboxymethylcellulose, 10 parts by weight of zinc oxide and 1 part by weight of disodium ethylene diamine tetraacetate, dispersing in 500 parts by weight of purified water, and uniformly stirring to obtain a water phase; weighing 50 parts by weight of vaseline and 10 parts by weight of tween 80 in another container, and mixing to obtain an oil phase; heating the water phase and oil phase respectively, mixing, homogenizing for 10min, cooling, and making into ointment A2 containing lysozyme.

Example 3 preparation of an ointment containing Lysozyme and emodin (1:1)

Weighing 5 parts by weight of lysozyme, 5 parts by weight of emodin, 50 parts by weight of sodium carboxymethyl cellulose, 10 parts by weight of zinc oxide and 1 part by weight of disodium ethylene diamine tetraacetate, dispersing in 500 parts by weight of purified water, and uniformly stirring to obtain a water phase; weighing 50 parts by weight of vaseline and 10 parts by weight of tween 80 in another container, and mixing to obtain an oil phase; heating the water phase and oil phase respectively, mixing, homogenizing for 10min, cooling, and making into ointment A3 containing lysozyme and emodin (weight ratio of 1: 1).

Example 4 preparation of an ointment containing Lysozyme and emodin (1:3)

Weighing 2.5 parts by weight of lysozyme, 7.5 parts by weight of emodin, 50 parts by weight of sodium carboxymethylcellulose, 10 parts by weight of zinc oxide and 1 part by weight of disodium ethylene diamine tetraacetate, dispersing in 500 parts by weight of purified water, and uniformly stirring to obtain a water phase; weighing 50 parts by weight of vaseline and 10 parts by weight of tween 80 in another container, and mixing to obtain an oil phase; heating the water phase and oil phase respectively, mixing, homogenizing for 10min, cooling, and making into ointment A4 containing lysozyme and emodin (weight ratio of 1: 3).

Example 5 preparation of an ointment containing Lysozyme and emodin (1:9)

Weighing 1 part by weight of lysozyme, 9 parts by weight of emodin, 50 parts by weight of sodium carboxymethyl cellulose, 10 parts by weight of zinc oxide and 1 part by weight of disodium ethylene diamine tetraacetate, dispersing in 500 parts by weight of purified water, and uniformly stirring to obtain a water phase; weighing 50 parts by weight of vaseline and 10 parts by weight of tween 80 in another container, and mixing to obtain an oil phase; heating the water phase and oil phase respectively, mixing, homogenizing for 10min, cooling, and making into ointment A5 containing lysozyme and emodin (weight ratio of 1: 9).

Test example 1 use of ointment containing lysozyme/emodin for treating psoriasis

200 cases of psoriasis patients are selected as study objects, and the clinical symptoms are mainly manifested by red papules on different parts of the body or maculopapules, wherein the chestnut granules are as common as bean granules; clinical observation of patients shows that the red papules have a gradual expansion trend, white scales can be clearly seen on the surfaces, and a layer of bright white membranous substance appears after the removal. Red spots appear after removal of the film. All patients were divided into 5 trial groups of 40 patients each, half of both men and women, using a random number table. The data of sex, age, clinical symptoms, psoriasis type and the like of each group of patients are compared, and the difference of each group is confirmed to have no statistical significance (P is more than 0.05) and is comparable. Each test group of patients was treated by applying the same weight of ointment prepared in examples 1 to 5 to the affected part 1 time each day in the morning and evening for a period of 1 month. The evaluation of the treatment effect is carried out by adopting the index scoring standard of the psoriasis skin damage area and the severity degree, and the effect is obvious: the patient's symptoms disappeared 70%, the subjective itching sensation disappeared; improvement: the symptoms of the patient disappear by 30 percent, and the subjective itching feeling is relieved; and (4) invalidation: symptoms did not change before and after treatment, and subjective feeling was not improved. The test results were statistically analyzed, and the specific results are shown in table 1.

TABLE 1 results of the lysozyme/emodin ointment for psoriasis patients

	Obvious effect (example)	Improvement (example)	Invalid (example)
				A1 (emodin)	8	18	14
A2 (lysozyme)	4	21	15
				A3 (weight ratio of lysozyme to emodin 1:1)	12	20	8
A4 (weight ratio of lysozyme to emodin 1:3)	18	19	3
				A5 (weight ratio of lysozyme to emodin 1:9)	13	17	10

The test results show that the psoriasis of patients is obviously improved after the pharmaceutical composition of the invention with various proportions is used, and the pharmaceutical composition of the invention is proved to have obvious effect on preventing and treating the psoriasis. Particularly, under the condition that the total amount of the active ingredients is kept unchanged, compared with the single use of the pharmaceutical composition (A1 and A2) containing the lysozyme and the emodin, the pharmaceutical composition containing the lysozyme and the emodin (A3-A5) has the advantages that the cure rate of the psoriasis is obviously improved, the synergistic effect is generated by the combined use of the lysozyme and the emodin, and the use amount of the active ingredients is reduced. The pharmaceutical composition with the weight ratio of lysozyme to emodin of 1:3 has the most prominent effect, and produces unexpected excellent effect.

Claims

1. An ointment containing lysozyme and emodin is characterized in that the ointment is prepared by the following steps: weighing 2.5 parts by weight of lysozyme, 7.5 parts by weight of emodin, 50 parts by weight of sodium carboxymethylcellulose, 10 parts by weight of zinc oxide and 1 part by weight of disodium ethylene diamine tetraacetate, dispersing in 500 parts by weight of purified water, and uniformly stirring to obtain a water phase; weighing 50 parts by weight of vaseline and 10 parts by weight of tween 80 in another container, and mixing to obtain an oil phase; heating the water phase and oil phase respectively, mixing, homogenizing for 10min, cooling, and making into ointment containing lysozyme and emodin.