CN110742888B - Application of bulleyaconitine A - Google Patents

Abstract

The invention relates to the technical field of medicaments, and discloses application of bulleyaconitine A. Animal model experiments and clinical experiments prove that the bulleyaconitine A can effectively reduce the thickness of the epidermis at the skin lesion, lighten the color of the skin lesion, reduce the area of rash, relieve pruritus and improve various physiological indexes, the related results have statistical differences, the bulleyaconitine A has obvious curative effect on treating psoriasis and high safety, and the invention provides the related application of the bulleyaconitine A in the aspect of treating the psoriasis.

Description

Application of bulleyaconitine A

Technical Field

The invention relates to the technical field of medicines, in particular to application of bulleyaconitine A.

Background

Psoriasis (Psoriasis), also known as Psoriasis, is one of the most common chronic inflammatory skin diseases, has a persistent state and a tendency to relapse, can cause damage to the systemic system, and has the main histopathological changes of abnormal keratinocyte proliferation, parakeratosis, inflammatory cell infiltration, dermal tissue vascular hyperplasia and the like. Meanwhile, psoriasis is also easy to cause various complications such as psoriatic arthritis, metabolic syndrome, malignant tumor and the like, secondary damage is brought to patients, and the life quality and even the life cycle of the patients are seriously affected. However, the exact pathogenesis of psoriasis is not yet clear. Research has shown that the occurrence of psoriasis is related to a variety of factors including genetic, immunological, environmental influences, etc., with immune mediation being its currently generally accepted pathogenesis.

At present, the medicines for treating psoriasis mainly comprise antitumor medicines, calcipotriol, glucocorticoid, immunosuppressant, biological agents and the like. The medicines can play a certain treatment role clinically, but after long-term administration, the medicines are easy to form drug resistance and dependence, and the biological preparation is expensive, so that huge economic burden is brought to patients. Therefore, researchers are actively searching for a new and effective anti-psoriasis drug in order to reduce side effects and burden on patients and society.

Disclosure of Invention

In view of the above, the present invention aims to provide an application of bulleyaconitine A in preparing a medicament for treating psoriasis. Preferably, the invention provides the application of bulleyaconitine A as the only active component in preparing the medicament for treating psoriasis.

Bulleyaconitine A is aconitine separated from Yunnan Sidodora (Aconitum Bulleyanum Diels), and has chemical name of (l alpha, 6 alpha, 14 alpha, 16 beta) tetrahydro-8, 13, 14-triol-20-ethyl-1, 6, 16-trimethoxy-4-methoxymethyl-8-acetoxy-14- (4' -p-methoxybenzyl ester) -aconitane, also known as Bulleyanonitine A. The structural formula is as follows:

the bulleyaconitine A has obvious analgesic and anti-inflammatory effects. The traditional Chinese medicine composition is clinically applied to rheumatic and rheumatoid arthritis, lumbar muscle strain, scapulohumeral periarthritis, limb sprain, contusion and the like. Rheumatic diseases, cervical spondylosis, nonspecific low back pain and the like belong to common chronic pain diseases of the old. The chinese schwann root et al report that aconitine-containing herbs mainly include yunaconitum yunnanensis (also called yunaconitum yunnanensis), aconitum kusnezoffii (a. forrestii Stapf), aconitum kusnezoffii (a. tatsienense fine & Gagnep.), aconitum kusnezoffii (a. transpectum Diels.), aconitum fistulosum (a. helminthepense var. fistulosa (tm.) wolf.f.), aconitum fischeri (a. fischeriana) w.t.wang), aconitum crassipes (a. fischeriana w.wang), aconitum yunnanensis (a. stapfianum hand-d.) and aconitum kusnezoffii (a. pacific. bulleyn.f.), aconitum kusnezoffii (a. pacific aconitum kusnezoffm.g.), aconitum kusnezoffii (a. pacific kusnezoff monkshood. kusnezoff monkshood.a. pacific aconitum (a. pacific.) and aconitum kusnezoff monkshood.

According to the invention, the bulleyaconitine A is found to be capable of remarkably improving the dorsal skin PASI score of psoriasis mice induced by imiquimod ointment, relieving weight loss and increasing the proportion of Treg cells in spleen and lymph nodes. The clinical curative effect research of the bulleyaconitine A and the placebo in random, double-blind and contrast shows that after the patient takes the bulleyaconitine A for treatment, the skin damage color becomes light, the rash disappears, the area is reduced, the pruritus symptom is relieved, the life quality is improved, and compared with the placebo control group, the bulleyaconitine A has statistical difference.

Meanwhile, the invention also carries out safety test on the bulleyaconitine A, SD rat gavage bulleyaconitine A tablet is carried out for 3 months under GLP experimental condition, the toxicity test result of multiple administration shows that the non-toxic reaction dose (NOAEL) of the product is 0.15mg/kg, the safe dose is 0.3mg/kg, and the conversion of the drug dose of adults and rats is about 6.15 times different, the non-toxic reaction dose (NOAEL) of the adults is 0.024mg/kg, and the safe dose of the adults is 0.049 mg/kg; if the adult is counted by 60kg, the non-toxic response dose (NOAEL) of the adult is 1.44mg/60kg, and the adult safe dose is 2.94mg/60kg, so that the effective dose in the non-toxic dose range and the safe dose range can achieve the effect of resisting psoriasis, and the psoriasis cannot be injured to a human body.

By combining the technical effects, the bulleyaconitine A has obvious curative effect on psoriasis and high safety, so the invention provides the application of the bulleyaconitine A in preparing the medicament for treating psoriasis.

According to the application provided by the invention, the invention also provides a medicine for treating psoriasis, which contains bulleyaconitine A and pharmaceutically acceptable auxiliary materials; the medicine can be any dosage form in clinic, including but not limited to tablets, granules (medicinal granules), pills, liquid preparations, capsules and the like; the choice of the excipients varies depending on the dosage form, and in the present embodiment, these excipients include, but are not limited to, lactose, microcrystalline cellulose, sodium carboxymethyl starch, magnesium stearate, powdered sugar, and hydroxypropyl cellulose, which are used to demonstrate the choice of excipients in tablets and granules.

In the animal model verification test, the medicine is prepared by polyethylene glycol and normal saline, and the concentration of the bulleyaconitine A is 0.125-0.5 mg/kg; in clinical trials, the drug is a tablet, and the dosage is below 2.94mg/60 kg.

According to the technical scheme, animal model tests and clinical tests prove that the bulleyaconitine A can effectively reduce the thickness of the epidermis of the skin lesion, lighten the color of the skin lesion, reduce the area of rash, relieve pruritus and improve various physiological indexes, the related results are statistically different, and the bulleyaconitine A has obvious curative effect and high safety in treating psoriasis, so that the bulleyaconitine A provides related application of the bulleyaconitine A in treating psoriasis.

Detailed Description

The invention discloses application of bulleyaconitine A, and can be realized by appropriately improving process parameters by taking the contents of bulleyaconitine A as reference by the technical personnel in the field. It is specifically noted that all such substitutions and modifications will be apparent to those skilled in the art and are intended to be included herein. While the invention has been described in terms of preferred embodiments, it will be apparent to those skilled in the art that variations and modifications, as appropriate, may be made to the applications and uses of the techniques disclosed herein without departing from the spirit, scope, and content of the invention.

The invention is further illustrated by the following examples.

Example 1: mouse model test

1. Laboratory animal

SPF grade BALB/C mice, body weight: 20 ± 2g, week old: male, free diet, temperature of 25 + -3 deg.C, and breeding in SPF animal room for 6-8 weeks.

2. Grouping animals

60 BALB/C mice were randomly divided into 5 groups, namely a normal group, a model group, an experimental group A, an experimental group B and an experimental group C, and each group had 12 mice.

3. Establishment of psoriasis model of mouse

The hair on the back of each mouse was shaved by electric hair clippers, and the remaining hair on the back of the mouse was removed by applying a depilatory cream to expose a skin area of about 3cm × 2 cm. Two days later, each mouse was applied with 62.5mg of 5% imiquimod ointment to the back skin daily for 7 consecutive days. The observation shows that the skin on the back of the mouse has obvious erythema, scale and thickening, namely the modeling is successful. Normal group mice were not given imiquimod ointment after dorsal epilation.

4. Test drugs and dosages

4.1 test drugs

Experimental group a: low dose group of bulleyaconitine A; experimental group B: the bulleyaconitine A is used as a traditional Chinese medicine; experimental group C: the bulleyaconitine A high dose group.

4.2 dosage administration

The test drugs of experimental groups A-C and normal group (no imiquimod ointment was given after depilating the back) were separately formulated with a normal saline solution containing 10% (v/v) polyethylene glycol 400 to a concentration equivalent to 5mg of the corresponding chemical component per ml and administered by intragastric administration. The amount of bulleyaconitine A administered in group A is 0.125 mg/kg; the bulleyaconitine A administered in group B is 0.25 mg/kg; experiment C group administered bulleyaconitine A0.5 mg/kg. 1 time daily for 7 days.

5. Experimental method

5.1 mice dorsal skin PASI score

The skin condition of the back of the mice was recorded daily using a digital camera and scored according to the PASI scoring criteria. The scoring items included erythema (erythema), scales (scales) and infiltration thickening (strickness) of the mouse skin lesions. Each was classified 0-4 according to severity and the PASI scoring criteria were as follows: 0-none; 1-mild; 2-moderate; 3-severe; 4-very severe. The scores of each mouse and the total scores of the three mice in each group are averaged and compared.

5.2 mouse weight determination

And weighing the weight of the mouse by using a small-sized weighing scale every day, comparing the weight with the weight of the mouse before modeling and administration treatment, and calculating the change condition of the weight of the mouse every day.

5.3 determination of epidermal thickness of mouse skin

After 7 days of administration after molding, the animals were sacrificed, and the skin of the mice was fixed in 4% neutral formaldehyde, soaked overnight, dehydrated in a programmed dehydrator, embedded in a paraffin embedding machine, and then the paraffin-embedded tissues were cut into 3 μm sections. Baking in a 65 deg.C oven for 1-2 hr, dewaxing to anhydrous, dyeing with hematoxylin for 10min, washing with tap water, dyeing with eosin hydrochloride for 15min, dehydrating, sealing with neutral gum, air drying overnight, observing with microscope, taking picture, and calculating skin epidermis thickness.

5.4 Treg cell ratio determination in spleen and lymph node of mouse

After sacrifice, the spleen and lymph nodes were removed, ground through a 40 mesh screen to make a cell suspension, centrifuged to remove the supernatant, and the spleen cells were treated with lysed erythrocytes and washed 2 times with PBS along with the lymph node cells. After cell counting, 106 cells per sample were resuspended in 100. mu.l PBS and 0.5. mu.l anti-CD4-FITC was added per sample and incubated for 30min at room temperature in the dark. Subsequently, the permeabilized cells are fixed, 0.5 mu l of anti-Foxp3-APC is added to each sample, the samples are incubated for 30min in the dark at room temperature, after PBS washing once, 300 mu l of PBS is used for cell resuspension, and the proportion of Treg cells is analyzed by adopting a flow cytometer.

6. Statistical treatment

All data were statistically analyzed using SPSS 25 statistical software, and the data were expressed as mean + -standard deviation.

7. Results of the experiment

7.1 mouse skin PASI score

Table 1 effect of bulleyaconitine a on the PASI score of the dorsal skin of mice (n ═ 12)

Group of	PASI score
		Normal group	0**
Model set	10.8±0.60
		Experimental group A	7.17±0.54*
Experimental group B	4.90±0.98**
		Experimental group C	5.60±0.43**

Note: p <0.05, P < 0.01 compared to model group.

The results in Table 1 show that the psoriasis model mice had a significant decrease in the dorsal skin PASI score (P <0.05 or P < 0.01) following drug treatment.

7.2 mouse weight changes

Table 2 effect of bulleyaconitine a on weight change in mice (n ═ 12)

Group of	Change in body weight
		Normal group	-0.05±0.62**
Model set	-4.90±0.70
		Experimental group A	-1.01±0.33**
Experimental group B	-0.97±0.52**
		Experimental group C	-1.06±0.43**

Note: p <0.05, P < 0.01 compared to model group.

The results in Table 2 show that the psoriasis model mice are significantly relieved of weight loss (P < 0.01) after drug treatment.

7.3 thickness of epidermal layer of mouse skin

Table 3 effect of bulleyaconitine a on thickness of skin epidermis in mice (n ═ 12)

Group of	Skin thickness (μm)
		Normal group	24.14±4.23**
Model set	142.37±6.74
		Experimental group A	45.38±3.74**
Experimental group B	34.67±1.32**
		Experimental group C	39.54±0.97**

Note: p <0.05, P < 0.01 compared to model group.

The results in Table 3 show that the epidermal thickness is significantly reduced (P < 0.01) after the drug treatment in psoriasis model mice.

7.4Treg cell proportion

Table 4 effect of bulleyaconitine a on Treg cells in mice (n ═ 12)

Note: p <0.05, P < 0.01 compared to model group.

The results in Table 4 show that the proportion of Treg cells in spleen and lymph node is increased significantly (P <0.05 or P < 0.01) after bulleyaconitine A treatment is given to psoriasis model mice.

8. Conclusion

The experimental result shows that after the psoriasis mouse induced by the imiquimod ointment is treated for 7 days by the bulleyaconitine A with the intragastric administration dosage of 0.125mg/kg, 0.25mg/kg and 0.5mg/kg, the PASI score of the back skin of the mouse can be obviously improved, the weight loss condition of the mouse is obviously relieved, the thickness of the epidermis of the mouse is obviously reduced, and the proportion of Treg cells in the spleen and lymph nodes of the mouse is obviously increased. Bulleyaconitine A can be used as medicine for treating psoriasis.

Example 2: clinical trial

1. Study object

The observed cases come from patients in outpatient clinics of dermatology department of traditional Chinese medicine hospital in Yunnan province from 12 months in 2017 to 5 months in 2019, and are all diagnosed as psoriasis vulgaris.

2. Diagnostic criteria

Made according to the Chinese clinical dermatology. The skin damage is red papule and macula, can be fused into pieces with obvious edges, and is covered with a plurality of layers of silvery white scales, a layer of semitransparent film can be seen after the scales are scraped off, and then the punctate bleeding phenomenon can be seen after the scales are removed.

2.1 grading Standard of disease severity

Grading the psoriasis disease degree according to the skin damage area:

mild symptoms: skin damage area < body surface area 10%;

the middle disease: the skin damage area is 10-30% of the body surface area;

severe disease: the area of skin damage is greater than 30% of the body surface area.

3. Inclusion and exclusion criteria

3.1 inclusion criteria

Meeting the diagnostic standard of psoriasis vulgaris;

② from 18 to 65 years old;

third, the severity of the disease: the degree of skin damage is mild to moderate (2< BSA < 10);

signing an informed consent form, volunteering to be tested, and conforming the informed consent process to the specification of GCP.

3.2 exclusion criteria

Women in pregnancy or lactation period have a birth control plan within three months;

② glucocorticoid and/or immunosuppressant drugs and retinoid drugs are taken orally in near one month or glucocorticoid preparations, retinoid drugs and vitamin D3 derivative preparations are taken orally and externally for two weeks;

③ patients with serious primary diseases such as cardiovascular, cerebrovascular, liver, kidney and hemopoietic system and mental disease;

fourthly, for the patients allergic to the research medicines;

patients participating in other medicine clinical tests.

3.3 termination of study cases

Firstly, a subject with serious adverse reaction judges that a clinical tester of the case should be stopped according to a researcher;

from the medical point of view, researchers need to stop testers;

thirdly, important deviation occurs in the implementation process of the clinical test scheme, such as poor compliance and the like, and the drug effect is difficult to evaluate;

and fourthly, the subject is unwilling to continue the clinical test in the implementation process of the clinical test scheme.

4. Case shedding and elimination

4.1 cases of exfoliation

4.1.1 shedding criteria

All subjects who filled out informed consent and screened for eligible entry into the trial had the right to exit the clinical trial at any time, and whenever they exited, subjects who did not complete the observation period specified by the protocol were called drop cases.

4.1.2 reasons for possible shedding

Adverse events, complications, or special physiological changes should not be continued, lack of efficacy, violation of protocol (including poor compliance), missed visits (including self-withdrawal of subject, change in subject's working environment, etc.), suspension by sponsor, and others. The statistical analysis should be combined with the actual situation for treatment, if the adverse reaction occurs, the statistics of the adverse reaction should be taken into account; the patients who fall off automatically due to ineffectiveness should be included in the therapeutic analysis.

4.1.3 treatment of cases of exfoliation

After the subject falls off, the investigator should contact him as much as possible to complete the assessment project that could be completed, record the last dose as much as possible, and fill out the test conclusion sheet. For subjects who are dropped due to adverse reactions, the test drug-related person is judged, and should be recorded in a CRF chart, and the person in charge is notified.

4.2 case removal

Firstly, patients can not follow up regularly to be a case of abscission after being brought in, and then are removed;

second, after the patients are brought into the hospital, the patients which do not meet the bringing standards are removed;

and eliminating the cases without any available data after the cases are brought into the database.

If one of the three symptoms is satisfied, the case is eliminated.

5. Quality control and data management

A leader group is set up, a perfect and detailed management system is established, and a standardized program for clinical research quality control is formed under the guidance of 'quality management standards of clinical medicine experiments'; the dermatologist, as the highest supervising executor of the research, supervises the whole implementation of the whole project; an inspector is set up to regularly monitor and process various research works; selecting cases strictly by inclusion criteria; signing an informed consent; randomly assigning the selected cases to each group; and a quality control and supervision mechanism is established to urge related personnel to complete a case report form according to requirements. And after the patients complete all follow-up visits, completing a case report table in time and performing data entry. The data obtained by the clinical observation is input and managed by a database management system made by Microsoft office-Access2007 software.

6. Test protocol

6.1 treatment group drug selection

The tablet of the embodiment 4 of the invention is orally taken, one tablet is taken at a time, three times a day, and each tablet contains 0.4mg of bulleyaconitine A.

6.2 control group placebo selection

The placebo adopts tablets without bulleyaconitine A, and the auxiliary materials and the preparation method are consistent with example 4.

The placebo with the same auxiliary materials and preparation methods as the control group and the treatment group has physical characteristics such as appearance, color, dosage form, weight, taste and smell similar to those of the test medicaments, does not contain active ingredients of research medicaments, and does not generate bias on judgment of a clinician on treatment effect, psychological acceptance degree of treatment of a patient and statistical analysis; and the psoriasis does not cause the disease aggravation caused by using the placebo, and has certain placebo effect, so the placebo is selected as a control and the effectiveness analysis is carried out.

The method comprises the following specific steps and time: the first day, visit, take photographs of skin lesions, perform examinations of blood routine, urine routine, liver function, kidney function, electrocardiogram, measure of skin barrier function, fill out psoriasis skin itch score (VAS), psoriasis skin lesion area and severity index (PASI), body surface area score (BSA);

the second, fourth and sixth weeks of the double-diagnosis are carried out, VAS scoring, PASI scoring and BSA scoring are carried out on the skin lesions, and the target skin lesions are photographed at each double-diagnosis;

and in the eighth week, blood routine, urine routine, liver function, kidney function and electrocardiogram are reviewed, skin barrier function is measured, VAS score, PASI score and BSA score are filled in, and the study is finished.

Adverse reactions: if any discomfort or any side effects occur during the trial, and if these side effects cannot be tolerated, the study medication is completely discontinued and the patient may be withdrawn from the study.

6.3 case number

The patients, 97 cases, aged 18-65 years, were divided into two groups, 50 in the treatment group and 47 in the control group.

7. Observation outcome indicator and observation time point

7.1 case Baseline level Observation

The demographic indexes are as follows: date of birth, age, gender, ethnicity, marital status, etc.;

② vital signs: body temperature, blood pressure, respiration, heart rate;

(iii) history of allergies: history of allergies to drugs, food, contacts;

fourthly, the existing medical history, the past history and the treatment history: psoriasis onset, family history, past treatment history, and other disease and treatment history;

auxiliary inspection: blood convention, liver function, kidney function, urine convention;

sixthly, psoriasis skin lesion area and severity index (PASI), psoriasis skin itch score (VAS), body surface area score (BAS).

7.2 evaluation of effectiveness and safety

7.2.1 effectiveness

The determination of the primary outcome index is based on the target lesion psoriatic lesion area and the disease severity score (PASI), as detailed in table 5.

TABLE 5 psoriasis skin lesion area and disease severity (PASI) score

The scoring method comprises the following steps:

target skin damage area (cm)²) Maximum length x maximum width of target skin damage

The percent reduction of the target lesion area (target lesion area value before treatment-target lesion area value after treatment)/target lesion area value before treatment × 100%

The target skin lesion area scoring method comprises the following steps:

the target skin lesion area before treatment is scored as 3 points uniformly, and according to the area reduction percentage, 2 points, 1 point and 0 point are sequentially recorded, specifically as follows,

the percentage of area reduction is more than or equal to 0 percent and less than 20 percent and 3 minutes

The percentage of area reduction is more than or equal to 20 percent and less than 60 percent and 2 minutes

The percentage of area reduction between 60 percent and 90 percent is 1 minute

The percentage of area reduction is more than or equal to 90 percent and less than or equal to 100 percent and 0 minute

Target lesion improvement PASI score area x (erythema + scaling + infiltration)

Evaluation of target lesion efficacy ═ (before treatment-after treatment)/before treatment × 100%

And (3) judging the clinical curative effect:

and (3) healing: the reduction of the PASI score is more than or equal to 95 percent; the effect is shown: the PASI score is reduced by 60-94%; the method has the following advantages: the PASI score is reduced by 30-59%; and (4) invalidation: reduction in PASI score < 30%.

Secondary outcome indicator:

PASI 50, PASI 75: i.e. after 8 weeks of treatment, the PASI score decreased to 50%, over 75% at week 0;

② psoriasis skin itch score (VAS), body surface area score (BAS).

7.2.2 safety observations and evaluation

Laboratory safety testing items include hematology, urinalysis, liver function, kidney function, electrocardiogram (evaluation at week 0, week 8, or upon recovery). Taking note of abnormal changes in laboratory test results, if necessary, reexamination and further judgment and evaluation of the relevance of the laboratory test results to study drugs are required, and if necessary, corresponding intervention measures are taken.

The symptoms, the occurrence time, the duration, the relevance (irrelevant, probable, relevant and relevant) to the test drug, the measures and the outcome of the adopted adverse reaction are recorded in detail, and the incidence rate of the adverse reaction is equal to the number of adverse reaction occurrences/total cases multiplied by 100%.

7.3 statistical analysis of data

The results of the clinical data are analyzed by a compliance program set (PPS), and the statistical analysis is calculated by SPSS 25 statistical analysis software. All statistical tests used a two-sided test, and a P value of 0.05 or less would be considered statistically significant for the differences tested. The related data description of each test group was statistically calculated by Mean ± standard deviation (Mean ± SD). Performing normality test on all data, wherein the data conform to normal distribution, the difference comparison before and after treatment in the same group adopts paired sample T test, and the difference comparison between groups adopts independent sample T test; data not conforming to the normal distribution were compared using the related sample rank sum test for the differences before and after treatment in the same group, and Wilcoxon rank sum test for the differences before and after treatment between the two groups.

8. Statistical results

The clinical observation test conforms to the inclusion standard, and 97 cases are total; patients in the cohort all completed 8 weeks of clinical observation.

8.1 target skin Damage improvement contrast

The improvement in target lesions, including erythema, scaling, infiltration and area, was evaluated in 97 patients using the Mann-Whitney test, and the results compared in Table 6 using the Wilcoxon signed rank test.

TABLE 6 comparison of target lesions in the treated and control groups

As can be seen from Table 6, the treatment groups were not statistically significantly different (P >0.05) from the control group for erythema, scaling, infiltration and area before the pharmaceutical composition of the invention was used, and the two groups were comparable. After 8 weeks of treatment, there was a significant statistical difference in area (P < 0.05). During follow-up, the area improvement of the treated group compared to the control group was statistically different at week 4.

8.2 comparison of target lesion PASI scores

The target lesions of the treatment group and the control group were scored for PASI, and the results are shown in table 7.

TABLE 7 comparison of target skin damage PASI

Time	Treatment group (M + -SD)	Control group (M + -SD)	Z	P value
					PASI (week 0)	18.8±7.45	18.6±7.24	-0.143	0.887
PASI (week 4)	13.2±7.28	15.4±7.18	-1.772	0.780
					PASI (week 8)	5.02±2.88	9.43±6.24	-4.046	0.00*

As can be seen from table 7, the mean score of the treated group was almost not different from that of the control group before the treatment, whereas the mean score of the treated group was statistically significantly different from that of the control group after 8 weeks of treatment.

8.3 comparison of target lesion PASI score efficacy before and after treatment

For the patients in the treatment group, 5 patients (10%), 40 patients with significant effect (80%), 3 patients with effect (6%), and 2 patients with no effect (4%) were cured in a clinical observation conducted for 8 weeks. The control group healed 1 (2.13%), the patients with significant effect 19 (40.4%), the patients with 11 effects (23.4%) and the patients with 16 effects (34.04%). The results are shown in Table 8.

TABLE 8 treatment Effect of target lesion PASI score before and after treatment

Group of	Healing process	Show effect	Is effective	Invalidation
					Treatment group	5(10％)	40(80％)	3(6％)	2(4％)
Control group	1(2.13％)	19(40.4％)	11(23.4％)	16(34.04％)

8.4 comparison of itching levels before and after treatment

The degree of pruritus is evaluated by a dermatosis pruritus score (VAS) scale, namely, the pruritus with different degrees is represented by a number of 0-10 on a line segment of 10 cm. The total weight is 0-III: grade 0, no itching; grade I, 1-3 points, mild pruritus (pruritus feeling, but normal life and sleep are not affected); grade II, 4-6 points, moderate pruritus (obvious pruritus, involuntary scratching, influence on sleep, but no taking of antipruritic medicine); grade III, 7-10 points, severe pruritus (severe pruritus, frequent scratching, severe influence on sleep and need of taking antipruritic medicines). The results of this clinical observation are shown in Table 9.

TABLE 9 comparison of itching levels before and after treatment

Time of follow-up	t	df	P value
				Week 2	1.824	96	0.089
Week 4	3.426	96	0.003
				Week 6	6.892	96	0.000
Week 8	8.650	96	0.000

As can be seen from table 9, the P values at weeks 4, 6 and 8 were 0.003, 0.000 and 0.000, respectively, all less than 0.05, indicating a statistical difference. The invention shows that the medicine composition has obvious effect of improving pruritus.

8.5 comparison of skin Barrier function

The skin barrier function detection of the clinical observation adopts a noninvasive examination method, and uses a SOFT 5.5 analyzer produced by CALLEGAR S.P.A to detect the skin temperature, the PH value, the skin surface moisture and the skin surface oil content of the target skin lesion.

Skin barrier function measurements were performed on the lesions of the treatment and control groups at weeks 0 and 8 using a SOFT 5.5 analyzer manufactured by calegas.p.a. to measure relevant parameters including skin temperature, pH, moisture, oil and fat, the average values of which are shown in table 10.

TABLE 10 mean values of skin barrier function between treatment and control before and after treatment

The experimental data obtained in table 10 were examined by rank-sum test, and the skin barrier function of the two groups of target lesions was examined before and after treatment, and the statistical results were examined by non-parametric test, and the results are shown in table 11.

TABLE 11 statistical results of skin barrier function of treatment group and control group before and after treatment

As can be seen from Table 11, the data of both groups before treatment were tested to be comparable without significant statistical difference (P > 0.05). After the treatment, two groups of data are subjected to rank sum test, the pH value and the moisture of the skin are obviously different (P <0.05), and the temperature and the grease are not obviously different (P > 0.05). Treatment groups: compared with the prior treatment and the post treatment, the skin temperature, the pH value, the skin moisture and the oil have obvious statistical differences (P is less than 0.05). Control group: compared with the prior and the post-treatment, all indexes have no obvious statistical difference (P is more than 0.05). The medicine has obvious effect on recovering the barrier function of the skin.

8.6 drug safety observations

The percentage of adverse reactions in this experiment was 12.4% (n-12), with 12% (n-6) in the treatment group and 12.8% (n-6) in the control group. The adverse reactions are respectively as follows: itching, burning, desquamation, rash, and increased dryness. There was no statistical difference between the treated group and the control group (P > 0.05).

In conclusion, the study of the clinical efficacy of the pharmaceutical composition and placebo in the randomized, double-blind and controlled manner showed that the total effective rate of the pharmaceutical composition in the study is 96% and the total effective rate of the controlled group is 65.96%, which are statistically different. The color of the skin lesion of the patient is lightened, the rash is eliminated, the area is reduced, the pruritus symptom is relieved, and the life quality is improved. The external preparation of the traditional Chinese medicine has obvious curative effect on psoriasis vulgaris and good safety.

Example 3: bulleyaconitine A safety test

Toxicity test data of SD rat gavage bulleyaconitine A tablet for multiple times of administration for 3 months under GLP test conditions of traditional Chinese medicine research institute (drug safety evaluation center in Chongqing city) show that the non-toxic reaction dose (NOAEL) of bulleyaconitine A is 0.15mg/kg, the safe dose is 0.3mg/kg, and the difference of drug dose conversion between an adult and a rat is about 6.15 times, the non-toxic reaction dose (NOAEL) of the adult is 0.024mg/kg, and the safe dose of the adult is 0.049 mg/kg; if the adult is counted as 60kg, the non-toxic response dose (NOAEL) of the adult is 1.44mg/60kg, and the adult safety dose is 2.94mg/60 kg. The specific test is as follows:

the test observation indexes are as follows: general conditions of animal appearance signs, behavior and activity, food intake, drinking, urine, excrement and the like are observed every day during the experiment; weighing body weight and food intake 1 time per week; detecting indexes of hematology, hematochemistry, urine, ophthalmology and the like at the end of administration and in the recovery period; animals were treated in each group at the end of dosing and during recovery, both male and female halves, and gross dissection and histopathological examination of the organs were performed.

And (3) test results: under the test conditions, SD rats are continuously administered 0.15, 0.3 and 0.8mg/kg of bulleyaconitine tablets by intragastric administration for 3 months (the administration day 50 is adjusted to be 0.6mg/kg), which are equivalent to 7.5, 15 and 40 times of the clinical dose of human (the administration day 50 is adjusted to be 30 times), and are recovered for 30 days after drug withdrawal. The results show that animals in the high-dose group die, the clinical observation of the animals mainly shows that the symptoms of perioral hair dampness (excessive salivary secretion) appear in an interval of 15-50 minutes after the drug administration, and the average weight value and the weight growth value of the animals are lower than those of a blank control group and are recovered after the drug administration is stopped; the food intake, hematology, blood biochemistry, urine, ophthalmology, organ weight, organ coefficient and histopathology examination of each dosage group have no abnormal change related to the test article; no target organs with obvious toxicity related to the drugs are found.

Example 4: bulleyaconitine A tablet

60g of lactose, 64g of microcrystalline cellulose, 14g of sodium carboxymethyl starch, 1.6g of magnesium stearate and 0.4g of bulleyaconitine A, and the components are directly pressed into 1000 tablets after being uniformly mixed, wherein each tablet contains 0.4mg of bulleyaconitine A.

Example 5: bulleyaconitine A capsule

60g of lactose, 64g of microcrystalline cellulose, 14g of sodium carboxymethyl starch, 1.6g of magnesium stearate and 0.4g of bulleyaconitine A are uniformly mixed, dry granulation is adopted, and the obtained granules are prepared into 1000 capsules, wherein each capsule contains 0.4mg of bulleyaconitine A.

Example 6: bulleyaconitine A granule

60g of lactose, 20g of powdered sugar, 40g of hydroxypropyl cellulose, 64g of microcrystalline cellulose, 14g of sodium carboxymethyl starch, 1.6g of magnesium stearate and 0.4g of bulleyaconitine A, uniformly mixing, performing dry granulation, and preparing 1000 bags of obtained granules, wherein each bag contains 0.4mg of bulleyaconitine A.

The foregoing is only a preferred embodiment of the present invention, and it should be noted that, for those skilled in the art, various modifications and amendments can be made without departing from the principle of the present invention, and these modifications and amendments should also be considered as the protection scope of the present invention.

Claims (4)

1. Use of bulleyaconitine A as the sole active ingredient in preparing medicine for treating psoriasis vulgaris is provided.

2. The use as claimed in claim 1, wherein the medicament for the treatment of psoriasis vulgaris comprises bulleyaconitine A and pharmaceutically acceptable excipients.

3. Use according to claim 2, wherein the pharmaceutical dosage form is a tablet, capsule, granule, pill or liquid formulation.

4. The use of claim 2, wherein the adjuvant is one or more selected from polyethylene glycol, physiological saline, lactose, microcrystalline cellulose, sodium carboxymethyl starch, magnesium stearate, sugar powder, and hydroxypropyl cellulose.