TW201436800A - Oil-extracted products of indigo naturalis, and the preparation process and uses thereof - Google Patents

Abstract

Disclosed herein are an oil-extracted product of indigo naturalis, and the preparation process thereof. Also disclosed is that the oil-extracted product of indigo naturalis can be used in the treatment of psoriasis, especially skin psoriasis and nail psoriasis.

Description

Oil extracted product of barley and its preparation method and use

The invention relates to an oil-extracted product of barley and a preparation method thereof. The invention also relates to the use of the oil extracted product of the barley to treat dryness (especially dry skin and nail dryness).

Psoriasis [also known as Yin Xie Bing] is a genetically associated chronic inflammatory skin disease that invades the skin of the body. The more common sites include : the scalp, the outside of the limbs, the elbows, and the knees. Cognac patients usually have redness and obvious papular or plaque on the skin, and the surface of the skin lesions is covered with a silvery-white scale, and some patients even Will be accompanied by symptoms such as arthritis.

At present, in the clinic, cognac can be divided into the following four common types according to the patient's skin condition: (1) chronic plaque psoriasis (also known as vulgaris psoriasis), It is the most common of all cognac types, with approximately 80% to 90% of cognac patients belonging to this type; (2) Guttate psoriasis, which is characterized by a majority of the skin of the patient. Small pink or red teardrop-shaped lesions usually occur in children between the ages of 7 and 10 years, and most patients have suffered a chain within 1 to 2 weeks before onset. Streptococcal infection; (3) erythrodermic psoriasis, characterized by erythema or exfoliation of the skin of the patient or over 90% of the body surface area, and Severe cases may have overall itching, accompanied by fever and malaise, and even cause systemic acute pustular psoriasis; and (4) pustular psoriasis It can be divided into generalized pustular psoriasis and localized pustular psoriasis [eg, pustulosis palmaris et plantaris and acrodermatitis perstans]

In addition, approximately 10% of patients with cognac are complicated with psoriatic arthritis, a ligaments, tendons, fascia, and spinal or peripheral joints that affect the patient. Peripheral joints) CD8 T cell-driven autoimmune inflammation; and up to 90% of patients with cognac have suffered from nail psoriasis.

Because the pathological condition and symptoms of dryness are quite complicated, it is still unclear about its pathogenesis. Studies have shown that inheritance plays an important role in the pathogenesis of dryness, while trauma, Infection, stress, endocrine factors, metabolic factors, climate, and drugs can all induce or exacerbate the onset of cognac. Once a dry patient is ill, the disease will eventually recur in one lifetime and cannot be eradicated.

Most Western medicine today adopts the most appropriate treatment according to the age, sex, occupation, intelligence, shape and distribution of the disease, the response to the previously used therapy, and other related medical history, and the main treatment methods. These include: (1) topical therapy: for example, corticosteroids, anthralin (trade name: Margiton ^® ), coal tar (trade name Polytar ^® ), Calcitriol (trade name: Silkis ^® ), tazarotene (trade name is Tazorac ^® ), and salicylic acid, etc., which are suitable for patients with mild conditions; (2 Systemic therapy: for example, oral preparations such as methotrexate (MTX), cyclosporin, and retinoids, which are suitable for moderate to severe Coronary patients with conditions; (3) Injection of biologics: for example, alefacept (trade name Amevive ^® ), eleucilumab (trade name) Known as Raptiva ^® ), etanercept (trade name Enbrel ^® ) and Adalimumab (trade name Humira ^® ), they are also suitable for patients with moderate to severe conditions of cognac; And (4) phototherapy: for example, ultraviolet B phototherapy [ultraviolet B (UVB) phototherapy], photochemotherapy (such as psoralen plus ultraviolet A (PUVA)] Etc., they are suitable for patients with dry conditions who have serious conditions.

However, long-term use of the above treatments may result in severe side effects or drug tolerances in the patient, which in turn reduces compliance. In view of this, many researchers have tried to find active components that can be used to treat cognac from traditional Chinese medicines (TCM).

青黛[English name: indigo naturalis or natural indigo; Hanyu Pinyin: qing dai; Chinese common name: dian hua, qing ge fen, qing gang hua, blue dew (lan lou), dian hua, and dian mo hua, etc. are from indigo-producing plants [such as Baphicacanthus cusia (Nees) Dark blue plant pigment extracted from the leaves of Bremek.), Polygonum tinctorium Lour., and Isatis indigotica Fort. Qinglan has the function of clearing the heat and cooling the blood in the clinical use of traditional Chinese medicine, and is often used in the clinical use of modern medicine to treat chronic myeloid leukemia, cognac, Mumps, various ulcers (eg, peptic ulcer, oral ulcer, etc.), hepatitis, herpes zoster, and otitis external.

It is known that various active ingredients can be isolated from barley obtained from different indigo-producing plants, among which indigo, indirubin, and isoindigo are more common. Studies have found that indirubin has anti-tumor, anti-inflammation and immunomodulation effects and is considered to be developed for the treatment of Alzheimer's disease. The potential of drugs (G. Eisenbrand et al. (2000), J. Cancer Res . Clin . Oncol ., 130: 627-635; T. Kunikata et al. (2000), European Journal of Pharmacology , 410: 93 -100; NKMak et al. (2004), Biochemical Pharmacology , 67: 167-174; S. Leclerc et al. (2001), The Journal of biological Chemistry , 276: 251-260).

It has been reported in the literature that the water solubility of barley is poor, so the organic solvent is most often used to extract barley and obtain various active ingredients. In QS Zhang et al. (2006), J Guangxi Normal University: Natural Science Edition , 24:58-60, QSZhang et al. mainly used 75% ethanol, chloroform, ethyl acetate and acetone, respectively. (acetone) to extract the barley, thereby obtaining four different organic extracts, and then comparing the content of indirubin contained in the organic extracts. The results of the experiment showed that when ethyl acetate was used for the extraction of barley, the content of indirubin was the highest, followed by acetone, chloroform and 75% ethanol, respectively.

Studies have found that the organic solvent-extracted product of indigo naturalis has significant anti-inflammatory or anti-psoriasis activity. For example, in YKLin et al. (2009), Journal of Dermatological Science , 54: 168-174, YKLin et al. dissolved barley powder in a ratio of 1:10 (w/v) in dimethylsulfoxide. In DMSO), it was then sterilized by filtration to obtain a guanidine-derived product of guanidine. The DMSO-extracted product of the barley was confirmed to reduce the number of cells of normal human epidermal keratinocyte (NHEK), and the cell cycle of the NHEK cell was arrested in the G0/G1 phase (G0). /G1 phase). In addition, the two main active ingredients (ie, sputum blue and indirubin) contained in the DMSO-extracted product of barley are also confirmed to cause cell cycle arrest of keratinocytes, and only indigo can be used. Regulation of proliferating cell nuclear antigen (PCNA) and expression of involucrin. Therefore, YKLin et al. believe that the DMSO-extracted product of the barley is mainly composed of indirubin as an active ingredient to regulate the proliferation and differentiation of keratinocytes, thereby achieving the anti-drying effect.

Although the organic solvent-extracted product of barley has anti-drying effect, it is necessary to further remove the organic solvent in clinical use to be made into a drug for human use. Therefore, the Applicant has attempted to utilize various vegetable oils for the extraction of barley, and the resulting oil-extracted product of indigo naturalis has been proven to be effective in treating cognac after clinical trials. (especially dry skin and dry nails).

Summary of invention

Thus, in a first aspect, the present invention provides an oil-extracted product of barley, which is obtained by extracting barley with an oil under heating.

In a second aspect, the present invention provides a method for preparing an oil-extracted product of barley, which comprises: extracting barley with an oil under heating.

The oil-extracted product of the barley according to the present invention has been confirmed by clinical trials to have significant therapeutic effects on dryness of human subjects (especially dry skin and nail dryness). Accordingly, in a third aspect, the present invention provides a pharmaceutical composition for treating dryness comprising an oil-extracted product of barley as described above.

In a fourth aspect, the present invention provides a method for treating an individual having or suspected of having dryness, comprising administering to the individual a petroleum extracted product of barley as described above.

The above and other objects, features and advantages of the present invention will become apparent from

Detailed description of the invention

It is to be understood that if any of the previous publications is quoted here, the prior publication does not constitute an acknowledgement that in Taiwan or any other country, the pre-existing publication forms a common general knowledge in the art. Part of it.

For the purposes of this specification, it will be clearly understood that the term "comprising" means "including but not limited to" and the term "comprises" has a corresponding meaning.

All technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which the invention pertains, unless otherwise defined. A person skilled in the art will recognize many methods and materials similar or equivalent to those described herein, which can be used to practice the invention. Of course, the invention is in no way limited by the methods and materials described.

As used herein, the term "psoriasis" is intended to encompass all types of cognac known to those skilled in the art, including chronic plaque psoriasis [also known as vulgaris vulgaris]. Psoriasis)], guttate psoriasis, erythrodermic psoriasis, pustular psoriasis, inverse psoriasis [also known as flexor dryness] (flexural psoriasis)], nail psoriasis, and psoriatic arthritis.

Traditionally, organic solvents have been used to extract various active ingredients in barley. It has been found that the organic solvent-extracted product of indigo naturalis can regulate the proliferation and differentiation of keratinocytes. Achieve anti-drying effect. However, the organic solvent contained in the organic solvent-extracted product of the barley may have an undesired side effect on the human body in clinical application, and also increases the cost of the drug process. Therefore, in order to develop an anti-dry drug that is safer and more convenient to use, the applicant has attempted to extract the barley with various vegetable oils and various extraction operating conditions.

Thus, the present invention provides an oil-extracted product of indigo naturalis which is obtained by extracting barley with an oil under heating.

According to the present invention, the oil-extracted product of the barley contains indirubin as an active ingredient.

The present invention also provides a method for preparing an oil-extracted product of barley, which comprises: extracting barley with an oil under heating.

According to the present invention, the oil suitable for extracting barley is selected from the group consisting of animal oils, vegetable oils, mineral oils, and combinations thereof. In a preferred embodiment of the invention, the extraction of the barley is carried out by using a vegetable oil.

According to the present invention, the vegetable oil may be, but not limited to, cottonseed oil, olive oil, sesame oil, sunflower seed oil, peanut oil, wheat germ. (wheat germ oil), soybean oil, jojoba oil, evening primrose oil, coconut oil, sesame oil, palm oil ,sweet almond Sweet almond oil, aloe oil, apricot kernel oil, avocado oil, borage oil, hemp seed oil, Hawaii Macadamia nut oil, rose hip oil, pecan oil, hazelunt oil, sasanqua oil, rice bran oil, shea butter Shea butter, corn oil, camellia oil, grape seed oil, canola oil, and castor oil. In a preferred embodiment of the invention, the vegetable oil is olive oil.

According to the present invention, the extraction ratio of the barley to the olive oil falls within the range of 1:10 (w/v) to 1:40 (w/v). In a preferred embodiment of the invention, the extraction ratio of the barley to the olive oil is 1:10 (w/v).

According to the invention, the barley is extracted at a temperature ranging from 100 ° C to 155 ° C. In a preferred embodiment of the invention, the barley is extracted at a temperature of 145 °C.

According to the invention, the barley can be further subjected to a filtration treatment after being heat-extracted.

According to the present invention, the oil-extracted product of the barley can be removed by the filtration treatment to remove indigo and impurities. This filtration process can be carried out using techniques well known and customary to those skilled in the art. For example, the filtration treatment can be carried out by using a cheese cloth or a filter paper or a filter membrane having a selected pore size. In a preferred embodiment of the invention, the filtering process is carried out by using a filter paper having a pore size of 6 μm.

The oil-extracted product of barley according to the present invention has been confirmed by clinical trials: it has significant therapeutic effects on dry lesions of patients (especially dry skin lesions and nail dryness lesions) and does not affect the skin of patients. Produces a severe irritating response. Based on the above, the oil-extracted product of the barley is expected to have a high potential for treating dryness. force. Accordingly, the present invention provides a pharmaceutical composition for treating dryness comprising an oil-extracted product of barley as described above.

In accordance with the present invention, the pharmaceutical composition can be formulated into a dosage form suitable for parenterally, topically or orally administration using techniques well known to those skilled in the art (dosage). Form), including but not limited to: sterile powder, tablet, troche, pill, capsule, external preparations, suppositoriums ) and similar things.

The pharmaceutical composition according to the present invention may further comprise a pharmaceutically acceptable carrier which is widely used in pharmaceutical manufacturing techniques. For example, the pharmaceutically acceptable carrier can comprise one or more agents selected from the group consisting of solvents, emulsifiers, suspending agents, decomposers, and binding agents. Agent), excipient, stabilizing agent, chelating agent, diluent, gelling agent, preservative, lubricant, Absorption delaying agents, liposomes, and the like.

In a preferred embodiment of the invention, the pharmaceutical composition is manufactured as an external preparation that can be applied directly to a cognac lesion.

According to the present invention, the external preparation may be, but not limited to, an emulsion, a gel, an ointment, a cream, a patch, an embrocation, an aerosol. (aerosol), spray, lotion, serum, paste, foam, and drop.

According to the present invention, the external preparation is prepared by mixing the oil-extracted product of the barley of the present invention with a base well known to those skilled in the art.

According to the invention, the substrate may comprise one or more additives selected from the group consisting of hydrocarbons [such as petroleum jelly). And white petrolatum], waxes (such as paraffin and yellow wax), preserving agents, antioxidants, surfactants, absorption enhancers Absorption enhancers, stabilizing agents, gelling agents, active agents, odor absorbers, and fragrances. The selection and quantity of these additives falls within the professional literacy and routine skills of those skilled in the art.

In a preferred embodiment of the invention, the pharmaceutical composition is manufactured in the form of an ointment. In another preferred embodiment of the invention, the pharmaceutical composition is manufactured in the form of a drop.

The invention also provides a method of treating an individual having or suspected of having dryness, comprising administering to the individual a petroleum extracted product of barley as described above.

According to the present invention, the dosage and the number of administrations of the oil-extracted product of the barley may vary depending on factors such as the severity of the disease to be treated, the route of administration, and the body weight, age, and physical condition of the individual to be treated. And reaction. In general, the daily dose of the pharmaceutical composition according to the present invention is usually from 0.01 mg/kg to 1 mg/kg body weight, in a single dose or divided into several doses, and can be administered orally or non-menstrually. Intestines are administered.

According to the present invention, the oil-extracted product of the barley can be administered alone or in combination with other drugs. Suitable for indigo of the present invention is oil-extracted product pharmaceutical combined together include, but are not limited to: corticosteroids (corticosteroids), dithranol (anthralin) (trade name Margiton ^®), coal tar (coal tar) ( The trade name is Polytar ^® ), calcitriol (trade name is Silkis ^® ), tazarotene (trade name is Tazorac ^® ), salicylic acid, and amine sputum ( Methotrexate, MTX), cyclosporin, retinoids, alefacept (trade name Amevive ^® ), efalizumab (trade name Raptiva ^® ), Etanercept (trade name Enbrel ^® ) and adalimumab (Adalimumab).

Figure 1 shows a comparison standard picture for assessing the severity of scales, erythema and thickness; Figures 2A to 2D show the position of the lesions respectively applied with the crude ointment of the present invention and the refined ointment, respectively, as measured over time. Individual average scores for scales, erythema, and thickness, and total average scores for the three items; Figure 3 shows the average of lesion locations measured over time with the crude ointment and refined ointment of the present invention applied separately The percentage value of the lesion area relative to the reference value; and Figure 4 shows the percent improvement in the appearance of the lesion measured over time with the lesion position of the crude ointment of the present invention and the refined ointment, respectively.

Detailed description of the preferred embodiment

The invention is further described in the following examples, but it should be understood that these examples are for illustrative purposes only and are not to be construed as limiting.

Example Experimental Materials:

1. The barley powder used in the following examples was purchased from Guang Sheng Trading, Taipei, Taiwan.

2. The various vegetable oils used in the examples below were purchased from Sigma, USA.

General experimental method:

1. Quantitative analysis of oil-extracted products in barley by high performance liquid chromatography (HPLC) Indirubin in oil-extracted product of indigo naturalis: It is known that indirubin is one of the main active ingredients of barley, so applicants use indirubin as an indicator component to evaluate different Operating conditions for the extraction efficiency of the oil-extracted product of barley.

The oil-extracted product of the barley obtained in the following examples was subjected to high performance liquid chromatography (HPLC), and indirubin (0.1 mg/mL) in chloroform (purchased from Alexis, Lausen). , Switzerland) was taken as a control standard and subjected to the same HPLC analysis. HPLC quantitative analysis was performed using an Agilent 1100 series liquid chromatography system (Agilent Technologies, Palo Alto, CA, USA), and the various operating conditions of HPLC are shown in Table 1 below. .

Example 1. Effect of different extraction conditions on the content of indirubin red in oil-extracted product of indigo naturalis experimental method: A. The effect of different vegetable oils on the indirubin content of the oil-extracted product of barley:

First, for 7 plant oils [including cottonseed oil, olive oil, sesame oil, sunflower seed oil, peanut oil, wheat germ oil (wheat germ oil) And soybean oil (2) each taken 2 mL and mixed with 0.1 g of barley powder, respectively, and the resulting mixture was heated at 110 ° C for 30 minutes, thereby obtaining oil extract of 7 kinds of barley. Oil-extracted crude product of indigo naturalis (all of which are bluish-purple). Thereafter, the crude oil-extracted crude products of the seven kinds of barley were filtered with a syringe filter (Millipore, France) and the filtrate was collected, thereby obtaining oil-extracted products of seven kinds of barley. (They are all purple). In addition, for comparison, two conventional organic solvents (including ethyl acetate and ethanol) were separately subjected to the same extraction step, thereby obtaining an organic solvent-extracted product of two kinds of barley. Product of indigo naturalis).

Then, 100 μL of each of the obtained oil-extracted products of the barley and the organic solvent-extracted products of the two kinds of barley were separately diluted 10 times with chloroform, and then according to the first item of the "general experimental method" above. The method described in "Quantitative analysis of indirubin in the oil-extracted product of barley by high performance liquid chromatography" was used to measure the content of indirubin.

result:

Table 2 shows the results of quantitative analysis of indirubin by the extraction product of barley obtained by using different extraction solvents. As can be seen from Table 2, the content of indirubin obtained when vegetable oil was used to extract green barley was between those using ethyl acetate and ethanol. In addition, among these vegetable oils, the content of indirubin obtained by extracting barley with sesame oil is the highest, followed by olive oil. However, the ingredients contained in sesame oil are complex and odorous, and it has been reported that sesame oil may cause an allergic reaction in an individual. Therefore, the applicant believes that the product extracted from olive oil by olive oil is better and safer in clinical application. complete.

B. Effect of heating temperature on the indirubin content of the product extracted from olive oil by olive oil:

First, 1 g of barley powder was mixed with 10 mL of olive oil, and then the resulting mixture was heated at 100 ° C, 110 ° C, 125 ° C, 145 ° C, and 155 ° C for 30 minutes, and then applied by a syringe filter. The filtrate was filtered and collected, whereby five olive oil-derived products of barley were obtained, respectively. Thereafter, 100 μL of each of the five extracts of the green peony oil extracted with olive oil was diluted 10 times with chloroform, and then quantitatively analyzed by high performance liquid chromatography according to the first item of the "general experimental method" above. The method described in the Saitama Red in the oil-extracted product of the barley is used to measure the content of indirubin.

result:

Table 3 shows the results of quantitative analysis of indirubin red by the olive oil-derived product of barley obtained at different heating temperatures. As can be seen from Table 3, it can be extracted at various heating temperatures. In particular, the indirubin content obtained by extracting barley at 145 ° C is the highest.

C. Screening the optimal extraction ratio of barley and olive oil:

First, 1 g, 0.5 g, and 0.25 g of barley powder were mixed with 10 mL of olive oil, respectively, whereby the extraction ratios of barley and olive oil were 1:10 (w/v) and 1:20 (w/, respectively). v) and 1:40 (w/v). Thereafter, the resulting mixture was heated at 145 ° C for 30 minutes, and then filtered with a syringe filter and the filtrate was collected, whereby three olive oil-derived products of barley were respectively obtained. Then, 100 μL of each of the three extracts of the green barley oil extracted with olive oil was diluted 10 times with chloroform, and then quantitatively analyzed by high performance liquid chromatography according to the first item of the "general experimental method" above. The method described in the Saitama Red in the oil-extracted product of the barley is used to measure the content of indirubin.

result:

Table 4 shows the results of quantitative analysis of the indirubin red by the olive oil-derived product obtained by extraction with different extraction ratios of barley and olive oil. It can be seen from Table 4 that the content of indirubin obtained when extracting barley in various extraction ratios is not much different. However, the extraction of barley is carried out at an extraction ratio of 1:20 (w/v) and 1:40 (w/v). When the amount of barley is fixed, the amount of olive oil added does not increase in the barley. The content of indirubin in the product extracted from olive oil, but will cause the extract of the barley to be extracted with olive oil. The concentration of the product is relatively low. Therefore, the applicant believes that the extraction of a barley with a higher concentration of indirubin can be obtained by extraction with a ratio of 1:10 (w/v) of barley and olive oil. .

Based on the experimental results of items A to C above, Applicants summarized the preferred extraction operating conditions in Table 5 below.

Example 2. Large scale preparation of the olive oil extracted product of the barley of the present invention.

First, according to the extraction operating conditions shown in Table 5 above, 90 g of barley powder and 900 mL of olive oil were placed in a stainless steel pot and mixed, and then the resulting mixture was heated at 145 ° C for 30 minutes. This gave a crude product of olive oil extracted from blue-purple barley. In addition, another crude product extracted from the olive oil obtained according to the above-mentioned barley is filtered with a filter paper having a pore size of 6 μm, and the filtrate is collected, thereby obtaining a product of a purplish red barley extracted with olive oil. . The crude product extracted from the olive oil obtained from the above-mentioned barley and the olive oil-extracted product of the barley isolated therefrom were subjected to the following clinical experiments.

In addition, in order to understand the main component distribution of the olive oil-extracted product of the barley of the present invention, the applicant takes the olive oil-extracted product of the barley obtained above for high performance liquid chromatography, according to YKLin et al. . (2009) the product of indigo was extracted with DMSO (same as above) among methods disclosed are used to obtain the same HPLC analysis for comparison. HPLC analysis was performed using an Agilent 1100 Series liquid chromatography system, and the HPLC operating conditions were as shown in Table 1 above. From the obtained HPLC elution profile, it was found that the olive oil-derived product of the barley obtained according to the present invention is different from the DMSO-extracted product of barley obtained according to the method disclosed by YK Lin et al. The elution peaks indicate that the two extracts contain different ingredients (data not shown).

Example 3. Evaluation of the efficacy, safety and convenience of the indigo naturalis ointment of the present invention in improving human skin dryness

In order to understand the therapeutic effect of the crude product extracted from the olive oil extracted from the barley according to the present invention and the olive oil extracted from the barley isolated therefrom, the therapeutic effect of the human subject suffering from dry skin and the use thereof. Safety and convenience, the following clinical trials were conducted.

Experimental materials, individuals, and methods: A. Preparation of crude ointment and refined ointment of barley:

The crude product extracted from the olive oil obtained in the above Example 2 and the olive oil-extracted product of the barley were each taken 900 mL, and then 90 g of yellow wax and 90 g of petroleum glue were added to both products. (ptroleum jelly), whereby an indigo naturalis crude ointment and an indigo naturalis refined ointment are separately used.

B. Screening and clinical information of experimental individuals:

The subjects involved in this clinical trial were from Chang Gung Memorial Hospital (Chang Gung). Memorial Hospital) Chinese individual in the Chinese medicine outpatient department, they must meet each of the inclusion criteria as shown in Table 6 below, and do not have the exclusion criteria shown in Table 6. Any of the (exclusion criteria). In addition, the trial was approved by the Institutional Review Board of Chang Gung Memorial Hospital and all of the individuals participating in the trial were informed of their informed consent.

A total of 38 patients participated in the trial, and clinical information about these patients [such as gender, age, duration, psoriasis area and severity index (PASI), and body surface area of the cognac lesions (body surface area, BSA)] is shown in Table 7 below.

C. Human clinical trials:

First, from each of the patient's body parts (excluding the head and neck), two lesions are similar in the extent of the disease and are photographed and measured separately, and then applied separately to the above A The crude ointment and refined ointment prepared in the item (dose: 1 g of ointment per 100 cm 2 of area, once a day), the test time lasted for 8 weeks. At the end of the 2nd, 4th, 6th and 8th week after the application of the ointment, the crude ointment treatment and the treated ointment treated parts of all patients were photographed and measured, respectively, according to the following D and E The method described in the item is used for evaluation.

Throughout the trial, a total of 35 patients completed all trials, while 3 patients completed only partial trials. One patient developed itching at the 5th week after the start of the trial because of the location of the treatment. In the case of the case, the other two patients withdrew from the fourth week and the sixth week after the start of the trial because of work factors and the inability to return to the hospital.

D. Assessment of therapeutic utility: 1. Assessment of the severity of scaling, erythema, and thickness:

The crude ointment treatment and the treated ointment treated site of each patient were performed before the application of the ointment (that is, at the 0th week) and at the end of the 2nd, 4th, 6th and 8th weeks after the application of the ointment. The photos taken by the photographs were taken by a dermatologist and compared with a standard picture (see Figure 1). The comparison items included the severity of scales, erythema, and thickness, and scored in the following five categories. To be the basis for the difference: score 0, indicating normal; score 1, indicating slight; score 2, indicating moderate; score 3, indicating serious; and score 4, indicating very serious. Thus, the individual average scores of scales, erythema, and thickness at the different test time points of the crude ointment treatment and the treated ointment and the total average score of the three items were obtained.

The experimental data obtained are expressed as mean ± standard deviation (SD). All data were analyzed by two-way analysis of variance (two-way ANOVA) to assess the differences between the various ointment treatment sites at various test time points, and The difference between the treatment with the crude ointment and the site treated with the refined ointment at the same test time point. If the resulting statistical alignment result is p < 0.05, it represents statistical significance.

2. Changes in lesion area:

At the end of the application of the ointment (ie at the 0th week) and at the end of the 2nd, 4th, 6th and 8th weeks after the application of the ointment, the treatment of the crude ointment and the refined ointment for each patient were respectively The lesion area (in square centimeters) is measured, and the lesion area of the two treatment sites changes with time as the average lesion area measured at different test time points relative to the baseline value (baseline value) The percentage value is expressed as the average value of the lesion measured before the application of the ointment (i.e., at week 0). The percentage value of the average lesion area relative to the reference value is calculated by referring to the following formula (1): Formula (1): A = (B/C) × 100

Where: A = percentage of the average lesion area relative to the reference value (%)

B = average lesion area (cm ² ) measured at week 0 and at the end of weeks 2, 4, 6 and 8

C = average lesion area measured at week 0 (cm ² )

The experimental data obtained are expressed as mean ± standard deviation (S.D.), and statistical analysis was performed in the manner described in the above point 1.

3. The improvement percentage of lesion appearance:

The total average score of the three items of scale, erythema, and thickness measured at each test time point in point 1 above and the average lesion area measured at each test time point in point 2 above. The percentage value of the reference value is multiplied, thereby obtaining the area and severity score at different test time points of the portion treated with the crude ointment and the refined ointment. The percentage improvement in the appearance of the lesions at the different treatment time points of the two treatment sites was calculated by substituting the obtained area and severity score values into the following formula (2): formula (2): D =[(EF)/E]×100

Where: D = percentage improvement in the appearance of the lesion (%)

E = area and severity score measured at week 0

F = area and severity scores measured at the end of weeks 2, 4, 6 and 8

The experimental data obtained are expressed as mean ± standard deviation (S.D.), and statistical analysis was performed in the manner described in the above point 1.

4. Assessment of the overall improvement in the appearance of the lesion:

A photograph taken by a dermatologist to visually compare the results of 35 patients who had completed the trial at the end of the 8th week and the 0th week (ie, before the ointment was administered). In this way, the overall improvement of the appearance of the lesions at the site treated by the crude ointment and the refined ointment can be evaluated, and the following six kinds of scores are used as the basis for the difference: a score of 0 indicates deterioration of the lesion; a score of 1, indicates No improvement; score 2 indicates a slight improvement; score 3 indicates a moderate improvement; score 4 indicates a near disappearance of the lesion; and score 5 indicates a disappearance of the lesion. In addition, the patients also visually observed the overall improvement of the appearance of the lesions at the two treatment sites, and scored in the manner described above.

After that, the number of patients corresponding to the six scores obtained by the patient's self-assessment is evaluated by the dermatologist, and the number of patients with each score is calculated relative to the total number of patients. The percentage value. The experimental data obtained are expressed as a percentage and analyzed by the McNemar test to understand the correlation between the results evaluated by the physician and the results of the patient's self-assessment.

E. Assessment of safety and convenience:

The safety assessment was at the end of weeks 2, 4, 6 and 8 after the ointment was administered and the investigator asked and recorded if the patient had any skin irritation reactions [including itching). , irritation and erythema, and use the following four kinds of scores as the basis for distinction: score 0, meaning no; score 1, indicating slight; score 2, indicating moderate; and scoring 3, said serious. The convenience assessment was made by comparing the convenience of the crude ointment and refined ointment for each patient involved in the trial after the end of the trial.

result: A. Evaluation of the therapeutic utility of the crude ointment and refined ointment of the barley according to the present invention:

2A to 2D respectively show individual average scores of scales, erythema, and thickness measured by time of the lesion position of the crude ointment and the refined ointment of the present invention, respectively, and the total average score of the three items. value. As can be seen from Figures 2A to 2D, as measured by the lesion position measured at the 0th week (i.e., prior to the application of the ointment), the lesions were treated with the crude ointment and the refined ointment. Scales, erythema, thickness The average score and the total average score of the three items will decrease significantly with time, but the scores measured under the same time point are almost no. difference.

Fig. 3 and Fig. 4 respectively show the percentage values of the average lesion area with respect to the reference value and the percentage improvement of the appearance of the lesion measured by the lesion position of the crude ointment of the present invention and the refined ointment, respectively, as a function of time. As can be seen from Fig. 3 and Fig. 4, the area of the lesions treated with the crude ointment and the refined ointment, respectively, will be significantly smaller with time, and the percentage improvement of the appearance of the lesion will increase significantly with time. However, there is almost no difference in the area percentage value and the improvement percentage value measured between the two treatment sites at the same time point. The above experimental results show that the crude ointment and the refined ointment of the barley according to the present invention can significantly improve the skin dryness of the patient.

In addition, after the end of the trial, the dermatologist evaluated each patient with an overall improvement in the appearance of the lesion, and the results obtained are shown in Table 8 below. As can be seen from Table 8, most of the dermatologists and the patients participating in the trial considered that the crude ointment and the refined ointment according to the present invention can effectively improve the cognac lesion, and the two ointments have similar therapeutic effects.

B. Evaluation of the safety and convenience of the crude ointment and refined ointment of the barley according to the present invention:

Table 9 shows the number of patients who developed skin irritation and the scores at the end of weeks 2, 4, 6 and 8 after administration of the ointment. As can be seen from Table 9, only a small number of patients developed transient and mild itching, irritation or erythema after application of the crude ointment of the present invention and the refined ointment throughout the trial, but this did not affect them. Receive follow-up treatment. In particular, the crude ointment has a higher total score than the refined ointment, which means that the crude product extracted from olive oil of the present invention can be removed by filtration to remove impurities and pigments, thereby reducing the patient's use. Possible skin irritation.

In addition, a total of 35 patients completed the entire trial and compared the ease of use of the two ointments. Among them, 31 patients (88.6%) considered the refined ointment to be superior to the crude ointment in use. Three patients (8.6%) thought there was no difference between the two ointments. Only one patient (2.9%) considered the crude ointment to be preferred. According to this, the applicant believes that the product extracted from the olive oil extracted from the crude oil extracted from the barley oil by the olive oil is less contaminated due to less impurities and lighter color. The affected skin causes irritation or severe staining on the clothes, thereby increasing the convenience and willingness of the patient during use.

Example 4. Evaluation of the efficacy and safety of the indigo naturalis drop of the present invention in improving human nails (nail psoriasis)

In order to understand the therapeutic effect of the olive oil-extracted product of the barley according to the present invention on human nail dryness and the safety in use, the following clinical trials were conducted.

Experimental materials, individuals, and methods: A. Preparation of barley drops and olive oil drops:

The olive oil-extracted product of the barley obtained in the above Example 2 was directly dispensed into dropping bottles and used as a barley drop in the following clinical trial. In addition, olive oil was dispensed into a dropper bottle and used as an olive oil drop in the following clinical trials.

B. Screening and clinical information of experimental individuals:

The subjects participating in this clinical trial were human subjects from the TCM clinic of Chang Gung Memorial Hospital, who must meet each of the inclusion criteria as shown in Table 10 below, and did not have the exclusion criteria shown in Table 10. Any of them. In addition, the trial was approved by the Human Rights Trial Ethics Committee of Chang Gung Memorial Hospital and all of the individuals involved in the trial were informed of their consent.

A total of 31 patients participated in the trial, and clinical information about these patients (such as gender, age, duration of disease, PASI, and BSA of cognac lesions, etc.) are shown in Table 11 below.

C. Human clinical trials:

First, the left and right hands of each patient were randomly divided into the experimental group and the control group, and the lesion position of the nail dryness was photographed, and then the green scorpion drops prepared in the above item A were applied to the experimental group. Nail nail plate, nail wrinkles (nail fold) and hypopichium (dose: 0.05 to 0.1 mL of drops per nail, 2 times a day for at least 30 minutes each), the test time lasted for a total of 24 weeks. In addition, the clinical trial of the nail of the control group was carried out in accordance with the above method, except that the nail was applied during the test from the start of the test to the end of the 12th week to be prepared in the above item A. Olive oil drops, and during the test period from the beginning of the 13th week to the end of the 24th week, the treatment is carried out with a drop of green peony. At the end of the 4th, 8th, 12th, 16th, 20th and 24th weeks after the administration of the drops, the nails of the control group and the experimental group of all the patients were photographed according to the items D and E below. The method described is used for evaluation.

Throughout the trial, a total of 27 patients completed all trials, while 4 patients completed only partial trials and they withdrew from trials at 8, 8, 20, and 24 weeks.

D. Assessment of therapeutic utility: 1. Evaluation of nail psoriasis severity index (NAPSI):

Control group and experimental group of each patient before the administration of the drops (ie at the time of week 0) and at the end of weeks 4, 8, 12, 16, 20 and 24 after the administration of the drops The photos taken by the nails were taken for NAPSI evaluation, and the NAPSI evaluation was based on the method described in Rich P. and Scher RK (2003), J. Am . Acad . Dermatol , 49: 206-212. get on. Briefly, the nails of the control group and the experimental group are respectively divided into an quaternary horizontal line and an imaginary longitudinal line into quadrants, and then the fourth step is performed according to the following steps. The clinical characteristics of the nail dryness [including the nail matrix psoriasis and the nail bed psoriasis] (see Table 12) were scored:

(a) Evaluation of the clinical features of the nail matrix dryness: In each of the four aliquots of the nail, each clinical feature of the nail matrix dryness is evaluated and counted in the following five categories. Divided into the basis of the distinction: score 0, indicating that the four aliquots did not show any clinical features; score 1, indicating that there is an aliquot in any of the four aliquots showing any of the clinical features; score 2, Indicates that there is a halved score in any of the aliquots that exhibits any of the clinical features; a score of 3 indicates that there are three aliquots in the aliquot that exhibit any of the clinical features; and a score of 4 indicates a four Each of the aliquots exhibits any of the clinical features. Accordingly, each nail can obtain a score (0 to 4) regarding the clinical characteristics of the nail matrix dryness.

(b) Assessment of the clinical characteristics of the nail bed dryness: in each of the four aliquots of the nail, evaluate each clinical feature of the nail bed dryness and refer to the method described in point (a) above. Score. Accordingly, each nail can obtain a score (0 to 4) regarding the clinical features of the nail bed dryness.

(c) Adding the scores measured at the above two points, thereby obtaining the total score (0-8) of each nail.

(d) The total scores measured for each nail (n=5) of the experimental group and the control group were summed, thereby obtaining the NAPSI total scores (0 to 40) of the experimental group and the control group, respectively.

According to the above manner, the NAPSI total average scores of the nails of the experimental group and the control group of all patients at different test time points were respectively obtained.

In addition, before the administration of the drops (that is, at the 0th week), the clinical features of the nails of all patients (see Table 12) were separately counted, and it was found in the experimental group as well as The nails of the control group correspond to the number of the 8 clinical features, respectively. Then, the NAPSI was evaluated for the clinical characteristics of the nails of the experimental group and the control group, respectively, and the following five kinds of scores were used as the basis for the difference: a score of 0 indicates that the four equal parts of the nail did not show The clinical feature; score 1 indicates that one of the four aliquots of the nail exhibits the clinical feature; score 2 indicates that the aliquot in the four aliquots of the nail exhibits the clinical appearance. Feature; score 3, indicating that there are three aliquots in the four aliquots of the nail exhibiting the clinical feature; and score 4 indicating that the four aliquots of the nail exhibit the clinical feature. Accordingly, the nails of the experimental group and the control group were each given a NAPSI average score (0 to 4) for their clinical characteristics.

At the end of the 12th week after the administration of the drops, the nails of the experimental group and the control group were scored separately in accordance with the above-described manner. Accordingly, the nails of the experimental group and the control group were each given a NAPSI average score (0 to 4) for their clinical characteristics. After that, the nails of the experimental group and the control group were compared at the 0th week and the NAPSI average scores measured at the end of the 12th week, thereby observing the experimental group and the control group of the patient. The various clinical features exhibited by the nails are improved after administration of the green barley drops of the present invention.

The experimental data obtained are expressed as mean ± standard deviation (SD). All data were statistically analyzed by the mixed effect model followed by the Bonferroni test to assess the differences between test points in the experimental or control group. And the difference between the experimental group and the control group under the same test time point. If the resulting statistical alignment is p < 0.05, it is statistically significant.

2. Evaluation of the modified target NAPSI (modified target NAPSI):

Select one of the target nails from the experimental group and the control group of each patient to the extent of the disease, and then apply the target nails before the application of the drops (ie, at week 0). Photographs taken at the end of weeks 4, 8, 12, 16, 20, and 24 after administration of the drops were used for improved target NAPSI evaluation, and the improved target NAPSI evaluation This was carried out in accordance with the method described in Parrish CA et al. (2005), J. Am . Acad . Dermatol , 53: 745-746. Briefly, the target nails of the experimental group and the control group of each patient were respectively divided into four equal parts by an imaginary horizontal line and an imaginary vertical line, and then 8 items of the four-part nail drying were performed according to the following steps. The clinical features (see Table 12) and their severity are scored:

(a) Evaluation of clinical features of nail dryness: In each of the four aliquots of the target nail, each of the clinical features shown in Table 12 above was evaluated separately, and the following five scores were used as The basis of the difference: score 0, indicating that the four aliquots did not show the clinical features; score 1, indicating that there is an aliquot in the four aliquots showing the clinical features; score 2, indicating in the aliquot There are two aliquots showing this clinical feature; a score of 3 indicates that the aliquot has a clinical score in the fourth aliquot; and a score of 4 indicates that the aliquot exhibits this clinical feature. Accordingly, the target nail will receive a score (0~4) for each clinical feature.

(b) Assessment of the severity of nail dryness: The severity of each clinical feature appearing on the target nail is evaluated and the following four scores are used as the basis for the difference: a score of 0 indicates normal Score 1 for slight; score 2 for moderate; and score 3 for severe. Accordingly, the target nail will each receive a score (0 to 3) for the severity of each clinical feature.

(c) Multiplying the score values measured at the above two points, thereby obtaining individual total scores (0 to 12) of the eight clinical features.

(d) The total scores of the eight clinical features measured by the target nails are summed to obtain a modified target NAPSI total score (0 to 96).

According to the above method, the modified target NAPSI total average scores of the target nails of the experimental group and the control group of all patients at different test time points were respectively obtained.

The experimental data obtained are expressed as mean ± standard deviation (S.D.), and statistical analysis was performed in the manner described in the above point 1.

3. Percentage improvement in appearance of lesions:

The percentage improvement in the appearance of the lesions of the nails treated with the barley drops and the olive oil drops is obtained by applying the drops to the test group and the control group before the application of the drops (ie, at the 0th week) The total NAPSI total score score or the modified target NAPSI total average score value measured at the end of the 12th and 24th weeks after the agent is calculated by substituting the following formula (3): Formula (3) :G=(HI/H)×100

Where: G = percentage improvement in the appearance of the lesion (%)

H = total NAPSI average score measured at week 0 or modified target NAPSI total average score

I = total NAPSI average score or modified target NAPSI total average score at the end of weeks 12 and 24, respectively

The experimental data obtained are expressed as an average value, and statistical analysis was performed in the manner described in the above point 1.

4. Assessment of the overall improvement in the appearance of the lesion:

The photographs taken by the dermatologists at the end of the 12th and 24th weeks after the administration of the drops were visually compared with the photographs taken at the 0th week, respectively. This can be used to evaluate the overall improvement of the appearance of the lesions in the experimental group and the control group, and the following six kinds of scores are used as the basis for the difference: score 0, indicating deterioration of the lesion; score 1 indicating no improvement; score 2, Indicates a slight improvement; a score of 3 indicates a moderate improvement; a score of 4 indicates that the lesion has nearly disappeared; and a score of 5 indicates that the lesion has disappeared. In addition, the patients also visually observed the overall improvement of the appearance of the lesion of the nail of the experimental group and the control group, and scored by referring to the above manner.

After that, the number of patients corresponding to the six scoring values obtained by the patient's self-assessment is evaluated by the dermatologist, and the disease of each score is calculated. The percentage of the number of patients relative to the total number of patients. The experimental data obtained is expressed as a percentage.

E. Assessment of safety:

The safety assessment was at the end of weeks 4, 8, 12, 16, 20, and 24 after the administration of the drops, and the investigator asked and recorded any signs of skin irritation (including itching, irritation, and Red spot), and the following four kinds of scores are used as the basis for the difference: score 0, indicating no; score 1, indicating slight; score 2, indicating moderate; and score 3, indicating serious.

result: A. Evaluation of the therapeutic utility of barley drops according to the present invention:

Table 13 shows the NAPSI total average scores, the modified target NAPSI total average scores, and the percent improvement in lesion appearance measured for the nails of the experimental and control groups over time, respectively. As can be seen from Table 13, the nails of the experimental group were measured at the end of the 4th, 8th and 12th weeks compared to the scores measured at the 0th week (ie before the application of the drops). Both the total NAPSI average score and the improved target NAPSI total average score will decrease significantly over time. However, the individual scores measured by the control group did not change significantly with time. In particular, at the end of the 8th and 12th weeks, the scores measured by the nails of the experimental group were significantly lower than those of the control nails, and at the end of the 12th week, the lesions of the nails of the experimental group The percentage improvement in appearance was significantly higher than that of the control nails.

In addition, at the end of the 16th, 20th, and 24th weeks, the scores measured by the nails of the experimental group and the control group were significantly reduced with time, and at the end of the 24th week, the experiment was performed. The percent improvement in the appearance of the lesions measured by the nails of the group and the control group also showed a significant increase, and the data measured by the two groups did not differ much. In particular, the percentage improvement in the control nail at the end of week 24 was significantly higher than at the end of week 12. The above experimental results show that the barley drops according to the present invention can effectively improve nail dryness lesions.

In addition, before the administration of the drops (that is, at the 0th week) and at the end of the 12th week after the administration of the drops, the experimental group of the 30 patients and the control nails were taken separately for the nails. Eight clinical features were evaluated for NAPSI and the results obtained are shown in Table 14 below. It can be seen from Table 14 that among the clinical features of the nail matrix dryness, the incidence of nail depression is the highest, followed by the deck fragmentation; and in the clinical features of the nail bed dryness, the incidence of the nail bed separation The highest is followed by the discoloration of the nail bed, and the hyperkeratosis of the underarm. In addition, compared with the data measured at the 0th week, at the end of the 12th week, the scores of the experimental group nails measured for each of the 8 clinical features were reduced. In particular, at the end of the 12th week, the degree of improvement of nail separation in the nail bed, nail bed discoloration, and nail depression in the experimental group was significantly higher than that in the control group. The results of this experiment show that the barley drops according to the present invention have an improved utility for the clinical features of the nail dryness of the patient, particularly the separation of the nail bed, the discoloration of the nail bed, and the nail depression.

a: indicates the number of nails showing specific clinical characteristics; b: indicates the average score measured after NAPSI evaluation for a specific clinical feature; and **: when the experimental group ends at the 12th week The measured scores were compared with those of the control group, p < 0.01.

In addition, at the end of the 12th and 24th weeks after the administration of the drops, the dermatologist and each patient were evaluated separately for the overall improvement in the appearance of the lesion, and the results obtained are shown in Table 15 below. . As can be seen from Table 15, at the end of the 12th week, most of the dermatologists and the patients participating in the trial thought that the appearance of the nails of the experimental group was significantly improved compared with the control group; at the end of the 24th week, the skin The physician and the patient considered that the appearance of the lesions of the nails of the experimental group and the control group were significantly improved. In particular, the overall improvement in lesions measured at the end of Week 24 at the end of Week 24 was significantly better than at the end of Week 12.

B. Assessment of the safety of barley drops according to the invention:

All of the nails administered with the barley drops of the present invention did not produce any skin irritation response throughout the test (data not shown). The Applicant accordingly believes that the olive oil-extracted product of the barley of the present invention has a high potential to develop into a drug against nail dryness and can be used for a long period of time without undesired side effects.

All of the patents and documents cited in this specification are hereby incorporated by reference in their entirety. In the event of a conflict, the detailed description of the case (including definitions) will prevail.

While the invention has been described with respect to the specific embodiments of the invention, it will be understood that many modifications and changes can be made without departing from the scope and spirit of the invention. It is therefore intended that the invention be limited only by the scope of the appended claims.

Claims (14)

An oil-extracted product of barley, which is obtained by extracting barley with an oil under heating. An oil-extracted product of barley, as claimed in claim 1, which comprises indirubin as an active ingredient. An oil-extracted product of barley, as claimed in claim 1, wherein the barley is further subjected to a filtration treatment after being heated and extracted. An oil-extracted product of barley, as claimed in claim 1, wherein the barley is heated and extracted at a temperature ranging from 100 ° C to 155 ° C. An oil-extracted product of barley, as claimed in claim 1, wherein the oil is selected from the group consisting of animal oils, vegetable oils, mineral oils, and combinations thereof. An oil-extracted product of barley, as claimed in claim 5, wherein the oil is a vegetable oil and the vegetable oil is selected from the group consisting of cottonseed oil, olive oil, sesame oil, sunflower oil, peanut oil , wheat germ oil, soybean oil, jojoba oil, evening primrose oil, coconut oil, sesame oil, palm oil, sweet almond oil, aloe oil, apricot walnut oil, avocado oil, borage oil, hemp seed oil, macadamia Kernel oil, rosehip oil, walnut oil, hazelnut oil, camellia oil, rice bran oil, shea butter, corn oil, bitter tea oil, grape seed oil, canola oil and castor oil. A pharmaceutical composition for treating cognac comprising an oil-extracted product of barley as claimed in claim 1. A pharmaceutical composition according to claim 7 of the patent application, which is in a form for parenteral administration. A pharmaceutical composition according to claim 7 of the patent application, which is in the form of a dosage form for oral administration. A method for preparing an oil-extracted product of barley, which comprises: extracting barley with an oil under heating. The method of claim 10, wherein the barley is further subjected to a filtration treatment after being heated and extracted. The method of claim 10, wherein the barley is heated and extracted at a temperature ranging from 100 ° C to 155 ° C. The method of claim 10, wherein the oil is selected from the group consisting of animal oils, vegetable oils, mineral oils, and combinations thereof. The method of claim 13, wherein the oil is a vegetable oil and the vegetable oil is selected from the group consisting of cottonseed oil, olive oil, sesame oil, sunflower oil, peanut oil, wheat germ oil, large Soybean oil, jojoba oil, evening primrose oil, coconut oil, sesame oil, palm oil, sweet almond oil, aloe oil, apricot walnut oil, avocado oil, borage oil, hemp oil, macadamia nut oil, rosehip oil, Pecan oil, hazelnut oil, camellia oil, rice bran oil, shea butter, corn oil, bitter tea oil, grape seed oil, canola oil and castor oil.