JP2006022121A - Atopic dermatitis retarder - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a new high-safety medicine that can retard atopic dermatitis. <P>SOLUTION: An atopic dermatitis retarder including astaxanthin and/or an ester thereof is provided. Since it has extremely low toxicity, the atopic dermatitis retarder according to this invention has high safety and can be used continuously for a long period of time. <P>COPYRIGHT: (C)2006,JPO&NCIPI

Description

  The present invention relates to an atopic dermatitis inhibitor.

  Atopic dermatitis is a disease whose main lesions are rash eczema and dry skin that repeats exacerbations and remissions. The most important clinical manifestation of atopic dermatitis is itching and associated skin damage. Eczema of atopic dermatitis is often severely intensified, and the rash is further exacerbated by scratching, and the vicious cycle of further itching and scratching is repeated, resulting in chronic skin damage and eczema lesions. Become. Since skin is a protective barrier on the outermost layer of the human body, its damage and reduced function lead to a decrease in physical, chemical and biological defenses, leading to secondary diseases such as infections. In addition, it is accompanied by itchiness, pain, and visual discomfort, greatly reducing QOL. Atopic dermatitis usually repeats onset over several years to several decades. Therefore, long-term treatment is necessary.

  In recent years, allergic diseases such as atopic dermatitis are increasing in developed countries, and it is considered that environmental factors are the cause in addition to genetic predisposition. As the cause, it is said that it is derived from a diet or a contact material such as a tick, and various things are mentioned. Conventionally, atopic dermatitis has been seen only in young people, but in recent years, the number of affected persons has increased rapidly from adults to elderly people, and establishment of treatment has become an urgent task.

  As a therapeutic agent for atopic dermatitis, treatment with an externally applied steroid is considered to be most effective, and various steroids are used depending on the type and degree of skin symptoms, site, age and the like. However, steroids have the drawback that dermatitis recurs after use and its withdrawal becomes difficult. In addition, since steroids have hormone-like effects, they often cause side effects and sometimes cause serious pathological conditions.

  As drugs other than steroids, antihistamines (H1 receptor antagonists) and antiallergic agents are used for itching skin. The latter is expected to lower the amount of use and strength of external steroids, but requires 2 to 4 weeks from the start of administration to the onset of effects. In atopic dermatitis, Staphylococcus aureus is present on the skin surface, and in particular, many inflammatory lesions such as the erosion surface. From such a viewpoint, an antibacterial agent is also applied to atopic dermatitis. However, it merely prevents infection and does not improve atopic dermatitis itself.

  Thus, at present, there is no effective means for treating or preventing atopic dermatitis without side effects and habits.

Carotenoids (carotenoids) are fat-soluble biological pigments that are widely distributed in animals, plants, and microorganisms, and have about 600 to yellow to orange to red colors. Astaxanthin, which is one type, is contained in crustaceans such as krill, shrimp and crabs, salmon and trout muscles and eggs (such as salmon roe), and body surfaces such as Thai, carp and goldfish. Astaxanthin is known not only to be provitamin A and to have a remarkable antioxidant effect, but also to have an anti-inflammatory effect (for example, Patent Documents 1 and 2). Patent Document 2 describes that the anti-inflammatory action is greatly enhanced by combining with a brown sugar extract having an anti-inflammatory action as compared with the case of using astaxanthin alone. The content of astaxanthin in the cosmetic in the example of Patent Document 2 is extremely low. Astaxanthin is also known to suppress the Th1 immune response and stimulate the Th2 immune response (Patent Document 3). However, the effect of astaxanthin alone on atopic dermatitis is not known at all.
JP 7-300421 A JP 2004-331512 A Special table 2003-510353 gazette

  An object of this invention is to provide the new highly safe chemical | medical agent which suppresses atopic dermatitis.

  As a result of various studies on astaxanthin which is known as an antioxidant, it was found that astaxanthin has an atopic dermatitis inhibitory action, and the present invention was completed.

  The present invention provides an atopic dermatitis inhibitor containing astaxanthin and / or an ester thereof.

  According to the present invention, a new atopic dermatitis inhibitor is provided. Since the atopic dermatitis inhibitor of this invention has very low toxicity, its safety is high.

  Astaxanthin and / or an ester thereof contained in the atopic dermatitis inhibitor of the present invention has the following formula:

(Here, R ¹ and R ² are each independently a hydrogen atom or a fatty acid residue). The ester of astaxanthin is not particularly limited. For example, saturated fatty acids such as palmitic acid and stearic acid, or oleic acid, linoleic acid, α-linolenic acid, γ-linolenic acid, bishomo-γ-linolenic acid, arachidonic acid, Examples thereof include monoesters or diesters of unsaturated fatty acids such as eicosapentaenoic acid and docosahexaenoic acid. These can be used alone or in appropriate combination. Astaxanthin has a structure having extra oxo groups and hydroxy groups at both ends of the skeleton of β-carotene, and therefore has low molecular stability unlike β-carotene. On the other hand, the ester body (for example, krill extract) in which the hydroxyl groups at both ends are esterified with unsaturated fatty acid or the like is more stable.

  Astaxanthin and / or its ester used in the present invention may be either chemically synthesized or derived from natural products. Examples of the latter natural products include red yeast containing astaxanthin and / or an ester thereof; shellfish shells such as tigliopath (red daphnia) and krill; and microalgae such as green algae. In the present invention, an extract containing astaxanthin and / or its ester produced by any method can be used as long as the characteristics of astaxanthin and / or its ester can be utilized. In general, extracts from these natural products are used, and the extract may be in the form of an extract or may be appropriately purified as necessary. In the present invention, such a crude extract or crushed powder containing astaxanthin and / or an ester thereof, or an appropriately purified or chemically synthesized product containing astaxanthin and / or its ester may be used alone or in appropriate combination. Can do. Considering the stability in the body, an ester form is preferably used.

  The administration route of the atopic dermatitis inhibitor of the present invention may be either oral administration or parenteral administration. The dosage form is appropriately selected depending on the administration route. For example, injections, infusions, powders, granules, tablets, capsules, pills, enteric solvents, troches, liquids for internal use, suspensions, emulsions, syrups, liquids for external use, poultices, nasal drops, ear drops Agents, eye drops, inhalants, ointments, lotions, suppositories, enteral nutrients and the like. This can be used alone or in combination depending on the symptoms. In these preparations, auxiliary agents usually used in the pharmaceutical preparation technical field such as excipients, binders, preservatives, oxidation stabilizers, disintegrants, lubricants, and corrigents are used as necessary. .

  The dose of the atopic dermatitis inhibitor of the present invention varies depending on the purpose of administration and the situation (sex, age, weight, etc.) of the administration subject. Usually, for adults, 0.1 mg to 2 g per day, preferably 4 mg to 500 mg per day in the case of oral administration, in terms of astaxanthin free body, while 0.01 mg to 1 g per day for parenteral administration, Preferably it can be administered at 0.1 mg to 500 mg.

  The atopic dermatitis inhibitor of the present invention can be used not only as a pharmaceutical product as described above, but also as a quasi-drug, cosmetic, functional food, nutritional supplement, food and drink, and the like. When used as a quasi-drug or cosmetic, it can be used together with various adjuvants commonly used in the technical field such as quasi-drug or cosmetic, if necessary. Or, when used as a functional food, nutritional supplement, or food and drink, it is usually used in foods such as sweeteners, spices, seasonings, preservatives, preservatives, bactericides, and antioxidants as necessary. You may use with the additive used. Further, it may be used in a desired shape such as solution, suspension, syrup, granule, cream, paste, jelly, etc., or as necessary. The ratio contained in these is not specifically limited, It can select suitably according to a use purpose, a usage form, and a usage-amount.

(Preparation Example 1: Preparation of astaxanthin monoester)
Astaxanthin monoester was prepared as follows. The Haematococcus pulvialis strain K0084 was cultured at 25 ° C. under irradiating gas containing 3% CO ₂ under light irradiation conditions, under nutrient stress (nitrogen source deficiency), and cysted. The cysted cells were crushed by means commonly used by those skilled in the art, and the oily fraction was extracted with ethanol. The extract contained lipids such as triglycerides in addition to astaxanthins. The extract was subjected to column chromatography using a synthetic resin adsorbent to obtain a purified product containing an astaxanthin monoester. This purified product was analyzed by HPLC, and it was confirmed that this purified astaxanthin monoester contained a monoester having a molecular weight of 858 as a main component, did not contain free and diester forms of astaxanthin, and contained a slight amount of diglyceride. .

(Example 1: Examination of effect on mite antigen-induced atopic dermatitis model mouse-1)
About the astaxanthin monoester obtained in the said preparation example 1, the effect with respect to a tick antigen induced atopic dermatitis model mouse was examined.

  The astaxanthin monoester obtained in Preparation Example 1 was dissolved in olive oil to prepare a 3 mg / mL astaxanthin monoester test solution.

  Twenty 7-week-old NC / NgaTndCrlj (SPF) male mice were prepared, and 5 μg / site (1 μL / site) of mite antigen (mite extract-Dp: (LS Co., Ltd.) was administered intradermally 7 times at intervals of 3 days. On the 7th day after the start of mite antigen administration, the mice were divided into a test group and a control group as 10 mice per group based on the ear thickness (degree of ear edema) measured as follows. From the 7th day to the 21st day of mite antigen administration, the test solution was orally administered once daily with 10 mL / kg (30 mg / kg) daily using an oral sonde. The control group was orally administered 10 mL / kg of olive oil.

The ear symptoms were observed before the start of mite antigen administration (day 0) and on days 7, 14, and 21 after the start of administration, and at the same time, the thickness of the left and right ears (ear edema was measured using a dial thickness gauge. ) And the rate of edema in the pinna (ear edema rate). Regarding the observation of the symptoms of the auricle, with reference to the evaluation criteria of clinical symptoms of human atopic dermatitis, it was scored as follows:
Evaluation items: pruritus, redness / bleeding, edema, abrasion / tissue defect, and dryness Score: 0: Asymptomatic 1: Mild 2: Moderate 3: Severely scored for each item, and the total score was used as the individual score . For each pinna score, ear thickness, and ear edema rate, mean ± standard error was calculated.

  FIG. 1 shows the results of the score of the ear symptom observation, FIG. 2 shows the measurement results of the ear edema, and FIG. As shown in the figure, in the test group, after the start of astaxanthin administration (on the 14th and 21st day of antigen administration), the score of the auricle was lower than that of the control group, and the degree of ear edema was reduced and the rate of ear edema Was also falling. Therefore, it was found that astaxanthin suppresses the symptoms of atopic dermatitis. In this example, there were deaths that were probably due to the large dose of olive oil during the administration period.

(Example 2: Examination of effect on mite antigen-induced atopic dermatitis model mouse-2)
As for the astaxanthin monoester obtained in Preparation Example 1, the amount of olive oil used for administration was reduced, while the amount of astaxanthin monoester was the same, as in Example 1, mite antigen-induced atopic dermatitis model mouse The effect on was investigated.

  The astaxanthin monoester obtained in Preparation Example 1 was dissolved in olive oil to prepare a 12 mg / mL astaxanthin monoester test solution.

  In the same manner as in Example 1, the mice were divided into 10 groups per test group and control group. From the 7th day to the 21st day from the start of mite antigen administration, the test solution was orally administered once a day by 2.5 mL / kg (30 mg / kg) daily using an oral sonde. The control group was orally administered with 2.5 mL / kg of olive oil.

  The ear symptoms were observed before the start of mite antigen administration (day 0) and on days 7, 14, and 21 after the start of administration, and at the same time, the thickness of the left and right ears (ear edema) was measured using a dial thickness gauge. ) And the rate of edema in the pinna (ear edema rate). The time course of the ear edema rate is shown in FIG. In this example, there were no deaths during the administration period.

  As shown in the figure, in the test group, the ear edema rate decreased significantly after the start of astaxanthin administration (on the 14th and 21st day of antigen administration) (* in the figure is p <0.01, Student's t test). In addition, the pinna score and the degree of ear edema were also lower than the control group (data not shown).

(Reference Example 1: Measurement of 50% lethal concentration against HUVEC)
Human umbilical vein endothelial cells (HUVEC) (ATCC CRL-1730) were obtained from American Type Culture Collection and endothelial cell growth containing 10% fetal bovine serum supplemented with 1% Antibiotic-Antimycotic (GIBCO BRL, USA) Medium (CELL APPLICATIONS, USA)) was pre-cultured at 37 ° C. in a 5% CO ₂ atmosphere.

  Matrigel matrix (BD Biosciences, USA) was thawed and kept on ice at 4 ° C., and 50 μL of the matrix was transferred to each well of a 96+ well tissue culture plate. The plate was incubated at 37 ° C. for at least 1 hour to solidify the matrix solution.

  On the other hand, the astaxanthin monoester obtained in Preparation Example 1 above was dissolved in dimethyl sulfoxide (DMSO) and then diluted with distilled water to obtain 25000, 2500, 250, 25, and 40% in 40 (v / v)% DMSO. A stock test solution containing 2.5 μM astaxanthin monoester was prepared.

100 μL of HUVEC suspension (approximately 2.5 × 10 ³ cells / well) was placed in a 96-well Matrigel plate at 37 ° C. in a 5% CO ₂ atmosphere. After 24 hours, 100 μL of growth medium and 2 μL of each of the above stock test solutions or vehicles (40 (v / v)% DMSO) were added to each of the two wells and incubated for a further 72 hours. The final concentrations of DMSO and astaxanthin monoester were 250, 25, 2.5, 0.25, and 0.025 μM.

  At the end of the incubation, 20 μL of 90% alamarBlue reagent was added to each well and incubated for an additional 6 hours. Next, the fluorescence intensity of each well was measured at an excitation wavelength of 530 nm and an emission wavelength of 590 nm using a Spectrafluor Plus plate reader, and the number of viable cells was counted. This is based on the ability of viable cells to change alamarBlue from a non-fluorescent oxidized form (blue) to a fluorescent reduced form (red). The 50% lethal concentration was calculated as the concentration at which 50% of the number of cells at the start of the experiment.

As a result, the 50% lethal concentration (LC ₅₀ ) of astaxanthin monoester with respect to HUVEC was 250 μM (maximum dissolution concentration in DMSO), indicating that the toxicity was low.

  According to the present invention, a new atopic dermatitis inhibitor is provided. Since the atopic dermatitis inhibitor of the present invention has very low toxicity, it is highly safe and can be used over a long period of time.

It is a graph which shows the time-dependent change of the symptom observation score of the ear pinna of the mouse | mouth in the test group and control group of an astaxanthin monoester. It is a graph which shows the time-dependent change of the ear edema (ear thickness) of a mouse | mouth in the test group of an astaxanthin monoester, and a control group. It is a graph which shows the time-dependent change of the ear edema rate of the mouse | mouth in the test group and control group of an astaxanthin monoester (Example 1). It is a graph which shows the time-dependent change of the ear edema rate of the mouse | mouth in the test group and control group of an astaxanthin monoester (Example 2).

Claims (1)

  An atopic dermatitis inhibitor comprising astaxanthin and / or an ester thereof.

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