CN111743906A - Application of alfacalcidol in preparation of external preparation for intervening skin photodamage - Google Patents
Application of alfacalcidol in preparation of external preparation for intervening skin photodamage Download PDFInfo
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- CN111743906A CN111743906A CN202010249523.5A CN202010249523A CN111743906A CN 111743906 A CN111743906 A CN 111743906A CN 202010249523 A CN202010249523 A CN 202010249523A CN 111743906 A CN111743906 A CN 111743906A
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- alfacalcidol
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/59—Compounds containing 9, 10- seco- cyclopenta[a]hydrophenanthrene ring systems
- A61K31/593—9,10-Secocholestane derivatives, e.g. cholecalciferol, i.e. vitamin D3
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/0012—Galenical forms characterised by the site of application
- A61K9/0014—Skin, i.e. galenical aspects of topical compositions
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P17/00—Drugs for dermatological disorders
- A61P17/16—Emollients or protectives, e.g. against radiation
Abstract
The invention discloses application of alfacalcidol in preparing an external medicine for preventing and/or treating skin photodamage. Alfacalcidol in the invention can improve the proliferation activity of fibroblasts; can also inhibit the expression of MMP-1 and MMP-3, and reduce the degradation of extracellular matrix; and can significantly alleviate the symptoms of photodamage to the skin; can be used for preparing external medicines for intervening skin photodamage, and has wide clinical application prospect.
Description
Technical Field
The invention particularly relates to application of alfacalcidol in preparing an external preparation for intervening skin photodamage.
Background
In everyday conditions, parts of the human skin are exposed to sunlight. Due to different lifestyle and type of work, excessive sun exposure causes a series of damages of the skin, mainly caused by excessive irradiation of ultraviolet rays in the sun, especially medium-wave ultraviolet rays (UVB). UVB has a wavelength of 290-320 nm and moderate penetrating power, most of UVB contained in sunlight is absorbed by an ozone layer, only less than 2% of UVB can reach the surface of the earth, and the UVB is particularly strong in summer and afternoon. UVB has a erythematous effect on the human body, and can promote mineral metabolism and vitamin D formation in the body, but a series of acute and chronic reactions such as erythema, blisters, inflammation, roughness, thickening, pigmentation, telangiectasia, photoaging and the like can be induced to the skin by long-term or excessive irradiation.
After the skin undergoes photodamage, the epidermis develops thickness variations, with focal atrophy or hyperplasia, and pigmentation of the melanin. The dermis shows a tendency to keratinize due to a decrease in fiber function and a decrease in proliferative capacity of fibroblasts. Depending on the degree of damage, a series of skin diseases from mild to severe can be induced, which mainly include some photosensitive skin diseases (such as light eruption, actinic dermatitis and the like), light-aggravated skin diseases (such as acne, chloasma and the like), and even light carcinogenesis (such as solar keratosis, squamous cell carcinoma and the like). The diseases are easy to repeat, difficult to cure and easy to induce canceration, seriously harm the physical and mental health of people and are a research problem in the field of skin diseases. At present, the protection against skin photodamage and photodamage skin diseases mainly aims at avoiding excessive sun exposure and using sun cream and sun protection measures; the treatment of malignant light carcinogenic diseases such as solar keratosis has fewer optional means, great harm to skin side effect and limited clinical effect, and a new therapeutic molecular therapeutic medicament is urgently needed to appear.
Alfacalcidol is an active metabolite of vitamin D and plays an important role in regulating calcium balance and skeletal metabolism. Alfacalcidol can be activated by 25-hydroxylase in the liver and exerts vitamin D effects in the whole body and osteoblasts. It exerts a unique pleiotropic effect in the intestine, bone, parathyroid, muscle and brain. Alfacalcidol can increase the calcium recovery of small intestine and renal tubule, inhibit parathyroid hyperplasia, reduce parathyroid hormone synthesis and release, and inhibit bone absorption; can also increase the synthesis of transforming growth factor and insulin-like growth factor, promote the synthesis of collagen and bone matrix protein; in addition, the medicine can also regulate muscle calcium metabolism, promote muscle cell differentiation, enhance muscle strength, increase neuromuscular coordination and reduce falling tendency. Based on these pharmacological mechanisms, alfacalcidol is currently used mainly for improving symptoms caused by abnormal vitamin D metabolism (such as hypocalcemia, convulsion, bone pain, and bone damage), and also for osteoporosis. No reports have been made to date on the use of alfacalcidol for the repair of photodamaged skin.
Disclosure of Invention
The purpose of the invention is as follows: in order to solve the technical problems, the inventor of the present invention has found that alfacalcidol shows better activity in intervening skin photodamage by accident in experiments. Based on experimental data, the invention provides the use of alfacalcidol or related external preparations containing alfacalcidol in the preparation of medicines for preventing and/or treating skin photodamage.
The technical scheme is as follows: the application of alfacalcidol in preparing the external medicine for preventing and/or treating skin photodamage comprises the following specific structure:
as an optimization: the external medicine is a substance for promoting proliferation of fibroblasts.
As an optimization: the external medicine is a substance for inhibiting the expression of matrix metalloproteinase MMP-1 and MMP-3.
As an optimization: the external medicine is a substance that inhibits degradation of extracellular matrix.
As an optimization: the topical medicine can increase the thickness of epidermis, and improve the arrangement of elastic fiber and collagen fiber of dermis.
As an optimization: the external medicine is a substance for increasing the thickness of the epidermis and improving the arrangement of elastic fibers and collagen fibers of the dermis.
As an optimization: photodamage to the skin refers to ultraviolet-induced damage to the skin.
An external preparation for preventing and/or treating skin photodamage is prepared from alfacalcidol or alfacalcidol and acceptable auxiliary materials through a certain preparation process, and the external preparation is prepared from the following components in percentage by mass: 3.5% of alfacalcidol, 44-49% of purified water, 9-10% of 1, 2-propylene glycol, 5-6% of ethanol, 7% of polydimethylsiloxane, 800025% of PEG, and 0.5% of methylparaben.
A preparation method of the external preparation for preventing and/or treating the skin photodamage comprises the following steps:
(1) heating alfacalcidol, 1, 2-propylene glycol, ethanol and methylparaben in the prescribed amount to 50 ℃ to fully dissolve the alfacalcidol, the 1, 2-propylene glycol, the ethanol and the methylparaben into clear and transparent solution, namely a mixture A;
(2) heating purified water, polydimethylsiloxane and PEG 8000 to 50 ℃, and mechanically stirring to fully homogenize to obtain a mixture B;
(3) the mixture A is slowly added into the mixture B while the mixture is hot under high-speed stirring, and the mixture is uniformly stirred and then cooled to room temperature, so that the external preparation of the embodiment is obtained.
According to the using method of the external preparation for preventing and/or treating the skin photodamage, a proper amount of the external preparation is directly applied to the skin once a day without cleaning, and moisturizing cream can be applied before use.
Has the advantages that: alfacalcidol in the invention can improve the proliferation activity of fibroblasts; can also inhibit the expression of MMP-1 and MMP-3, and reduce the degradation of extracellular matrix; and can significantly alleviate the symptoms of photodamage to the skin; can be used for preparing external medicines for intervening skin photodamage, and has wide clinical application prospect.
Drawings
FIG. 1 is a schematic representation of HE staining of mouse skin according to the invention;
wherein panel a is normal mouse skin; panel b is skin of control mice (i.e. subjected to UVB only, without alfacalcidol intervention); panel c is experimental group mouse skin (i.e. subjected to UVB irradiation and alfacalcidol intervention).
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below so that those skilled in the art can better understand the advantages and features of the present invention, and thus the scope of the present invention will be more clearly defined. The embodiments described herein are only a few embodiments of the present invention, rather than all embodiments, and all other embodiments that can be derived by one of ordinary skill in the art without inventive faculty based on the embodiments described herein are intended to fall within the scope of the present invention.
Example 1
A representative external preparation for intervening photodamage of skin is composed of the following components: alfacalcidol 3.5%, purified water 49%, 1, 2-propylene glycol 10%, ethanol 5%, polydimethylsiloxane 7%, PEG 800025%, and methylparaben 0.5%. The mass percentages of the above substances are mass ratios.
The preparation method comprises the following steps:
(1) heating alfacalcidol, 1, 2-propylene glycol, ethanol and methylparaben in the prescribed amount to 50 ℃ to fully dissolve the alfacalcidol, the 1, 2-propylene glycol, the ethanol and the methylparaben into clear and transparent solution, namely a mixture A;
(2) heating purified water, polydimethylsiloxane and PEG 8000 to 50 ℃, and mechanically stirring to fully homogenize to obtain a mixture B;
(3) the mixture A is slowly added into the mixture B while the mixture is hot under high-speed stirring, and the mixture is uniformly stirred and then cooled to room temperature, so that the external preparation of the embodiment is obtained.
The using method comprises the following steps: the appropriate amount of the cream can be directly applied to skin by external application once a day without cleaning, and the moisturizing cream can be applied before use.
Example 2
A representative external preparation for intervening photodamage of skin is composed of the following components: alfacalcidol 3.5%, purified water 44%, 1, 2-propylene glycol 9%, ethanol 6%, polydimethylsiloxane 7%, glyceryl monostearate 15%, polyoxyethylene castor oil (Cremophor EL) 15%, and methyl paraben 0.5%. The mass percentages of the above substances are mass ratios.
The preparation method comprises the following steps:
(1) heating alfacalcidol, 1, 2-propylene glycol, ethanol and methylparaben in the prescribed amount to 50 ℃ to fully dissolve the alfacalcidol, the 1, 2-propylene glycol, the ethanol and the methylparaben into clear and transparent solution, namely a mixture A;
(2) heating purified water, polydimethylsiloxane, glyceryl monostearate and polyoxyethylene castor oil (cremophor EL) to 50 deg.C, mechanically stirring, and homogenizing to obtain mixture B;
(3) the mixture A is slowly added into the mixture B while the mixture is hot under high-speed stirring, and the mixture is uniformly stirred and then cooled to room temperature, so that the external preparation of the embodiment is obtained.
The using method comprises the following steps: the appropriate amount of the cream can be directly applied to skin by external application once a day without cleaning, and the moisturizing cream can be applied before use.
The present invention is illustrated by the following comparative test examples 1-2, specifically as follows:
comparative test example 1 alfacalcidol intervention photodamage cell experiment
(1) Establishing cell photodamage model
Separating fibroblast from foreskin of children by conventional method, culturing, subculturing to 3-5 generations, freezing, inoculating to culture dish after cell recovery, adding 50 mg/L8-methoxsalen (8-MOP) into culture medium, discarding culture medium during irradiation, rinsing with PBS containing 50 mg/L8-MOP for 2 times, and irradiating with UVA at dose of 9J/cm2Irradiation was continued for 21 days. The cells were observed under a microscope to show characteristic changes of damage: i.e., cells become larger, intracellular granules increase, vacuolization of cytoplasm and proliferation are delayed.
(2) Experimental drugs: alfacalcidol
(3) Fibroblast proliferation promoting Activity (%)
The experimental method comprises the following steps: the fibroblasts after UVA irradiation are inoculated into a 96-well plate according to the number of 6 multiplied by 105/well, 10 mu L of alfacalcidol with the concentration of 0.01 percent, 0.02 percent and 0.04 percent respectively is added after 24 hours of culture, 10 mu L of CCK-8 reagent is added after 24 hours of continuous culture, and the absorbance (A) value of 450nm is measured by an enzyme linked immunosorbent assay detector after 3 hours of incubation. Repeat 3 times.
Cell viability (%) - [ a (test group) -a (0) ]/[ a (blank) -a (0) ] × 100%;
wherein A (test group) represents the absorbance of wells with cells, CCK-8 solution and alfacalcidol; a (0) represents the absorbance of wells with CCK-8 solution, but without cells and alfacalcidol; a (blank control) has the absorbance of cells, CCK-8 solution, but wells without alfacalcidol.
TABLE 1 Effect of different concentrations of alfacalcidol on fibroblast proliferation
As shown in Table 1, alfacalcidol can increase the cell proliferation activity after acting on fibroblasts, and the increase degree has statistical significance (p < 0.05).
(4) Inhibiting the expression of matrix metalloproteinases (MMP-1, MMP-3)
The test was divided into four groups, which were an untreated group (normal control group not treated with alfacalcidol and UVA irradiation), an alfacalcidol-treated group (fibroblasts + 0.01% alfacalcidol), a UVA-treated group, and a UVA + alfacalcidol-treated group (treated with 0.01% alfacalcidol after UVA irradiation), and after culturing for 48 hours, cell supernatants were taken, after extraction, sample addition, incubation, and washing were performed according to the ELISA kit instructions, enzyme addition, incubation, and washing were performed, and finally, absorbance (OD value) was measured at a wavelength of 450nm with a microplate reader, and concentrations of MMP-1 and MMP-3 in the samples were calculated from the standard curves.
TABLE 2 Effect of alfacalcidol on MMP-1 and MMP-3 expression
As can be seen from Table 2, alfacalcidol inhibits the expression of MMP-1 and MMP-3, while MMP-1 and MMP-3 are important enzymes for the breakdown of extracellular matrix components: wherein MMP-1 primarily hydrolyzes type I, II, and III collagen; MMP-3 hydrolyzes substrates more extensively, including collagen, proteoglycans, gelatin, and the like. Thus inhibiting both activities may reduce degradation of the extracellular matrix.
Comparative test example 2 animal experiments with alfacalcidol intervention photodamage
(1) Establishing animal photodamage model
Removing hair from the back of the mouse with scissors and an electric razor, and subjecting the skin to UVB (300 mJ/cm) after exposing the skin sufficiently2) Irradiating the mice once a day for 3 days continuously, finishing molding, and enabling the back skin of the successfully molded mice to have obvious erythema and scale change.
(2) Experiment grouping
The number of mice successfully modeled was 20, and each of the control group and the test group was 10. The control group was not treated and the test group was administered topical alfacalcidol external preparation (example 1) at a concentration of 3.5% every other day for 2 weeks.
(3) Skin damage status was scored on UVB-irradiated areas of the backs of mice (see table 3 for details), and the results are shown in table 4:
TABLE 3 evaluation criteria for skin damage status of UVB-irradiated area of mouse back
| Scoring | Skin appearance state |
| 0 | The skin is fleshy and smooth and plump |
| 0.5 | The skin appears flesh color, and the skin surface appears slightly rough |
| 1 | Erythema, moderate roughness and deepening skin lines |
| 1.5 | Erythema, obvious roughness, slight desquamation and small amount of wrinkles on the skin |
| 2 | The skin shows purplish red fester, rough and thick skin, a great deal of desquamation and wrinkle increase |
TABLE 4 Effect of alfacalcidol on score of skin lesions on the dorsal side of photodamaged mice
| Day 1 | Day 3 | Day 5 | Day 7 | Day 10 | Day 14 | |
| Experimental group | 1.80 | 1.50 | 1.30 | 0.40 | 0.40 | 0.25 |
| Control group | 1.80 | 1.80 | 1.75 | 1.50 | 1.30 | 1.30 |
As shown in Table 4, after 3 times of alfacalcidol treatment, the skin photodamage condition can be remarkably improved, and the damage states of erythema, ulceration, folds, desquamation and the like can be remarkably relieved.
(4) Mice were sacrificed and harvested and HE stained for evidence as shown in fig. 1 (a-c): normal mice have intact and delicate stratum corneum (fig. 1 a); the stratum corneum of the control mice was exfoliated to some extent, and some of the dermis was cavitated and fibrotic (fig. 1 b); in contrast, the experimental group greatly improved the skin condition, the stratum corneum began to recover, and the epidermal and dermal structures below the stratum corneum were well aligned (fig. 1 c).
The experimental results show that alfacalcidol can improve the proliferation activity of fibroblasts; can also inhibit the expression of MMP-1 and MMP-3, and reduce the degradation of extracellular matrix; and can significantly alleviate the symptoms of photodamage to the skin; can be used for preparing external medicines for intervening skin photodamage, and has wide clinical application prospect.
Obviously, many modifications, substitutions, and variations are possible in light of the above teachings of the invention, without departing from the basic technical spirit of the invention, as defined by the following claims.
Claims (10)
1. The application of alfacalcidol in preparing an external preparation for intervening skin photodamage is characterized in that: the alfacalcidol has the following specific structure:
2. use of alfacalcidol according to claim 1 for the preparation of a topical formulation for the intervention of photodamage to the skin, characterized in that: the external medicine is a substance for promoting proliferation of fibroblasts.
3. Use of alfacalcidol according to claim 1 for the preparation of a topical formulation for the intervention of photodamage to the skin, characterized in that: the external medicine is a substance for inhibiting the expression of matrix metalloproteinase MMP-1 and MMP-3.
4. Use of alfacalcidol according to claim 3 for the preparation of a topical formulation for the intervention of photodamage to the skin, characterized in that: the external medicine is a substance that inhibits degradation of extracellular matrix.
5. Use of alfacalcidol according to claim 1 for the preparation of a topical formulation for the intervention of photodamage to the skin, characterized in that: the topical medicine can increase the thickness of epidermis, and improve the arrangement of elastic fiber and collagen fiber of dermis.
6. Use of alfacalcidol according to claim 1 for the preparation of a topical formulation for the intervention of photodamage to the skin, characterized in that: the external medicine is a substance for increasing the thickness of the epidermis and improving the arrangement of elastic fibers and collagen fibers of the dermis.
7. Use of alfacalcidol according to claims 1 to 6 for the preparation of an external preparation for intervention in photodamage to the skin, characterized in that: photodamage to the skin refers to ultraviolet-induced damage to the skin.
8. An external preparation for preventing and/or treating photodamage to skin, characterized by: the external preparation is formed by alfacalcidol or alfacalcidol and acceptable auxiliary materials through a certain preparation process, and the external preparation is composed of the following components in percentage by mass: 3.5% of alfacalcidol, 44-49% of purified water, 9-10% of 1, 2-propylene glycol, 5-6% of ethanol, 7% of polydimethylsiloxane, 800025% of PEG, and 0.5% of methylparaben.
9. A method for preparing the external preparation for preventing and/or treating photodamage to skin according to claim 8, wherein the external preparation comprises: the preparation method comprises the following steps:
(1) heating alfacalcidol, 1, 2-propylene glycol, ethanol and methylparaben in the prescribed amount to 50 ℃ to fully dissolve the alfacalcidol, the 1, 2-propylene glycol, the ethanol and the methylparaben into clear and transparent solution, namely a mixture A;
(2) heating purified water, polydimethylsiloxane and PEG 8000 to 50 ℃, and mechanically stirring to fully homogenize to obtain a mixture B;
(3) the mixture A is slowly added into the mixture B while the mixture is hot under high-speed stirring, and the mixture is uniformly stirred and then cooled to room temperature, so that the external preparation of the embodiment is obtained.
10. A method of using the external preparation for preventing and/or treating photodamage to skin according to claim 8, wherein: the using method comprises the following steps: the appropriate amount of the cream can be directly applied to skin by external application once a day without cleaning, and the moisturizing cream can be applied before use.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113274382A (en) * | 2021-06-16 | 2021-08-20 | 南通大学 | Application of tricin in preparation of external preparation for intervening skin photodamage |
Citations (6)
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|---|---|---|---|---|
| US4610978A (en) * | 1983-03-22 | 1986-09-09 | Yissum Research Development Company Of The Hebrew University Of Jerusalem | Compositions containing 1α-hydroxycholecalciferol for topical treatment of skin disorders and methods employing same |
| JPH01249714A (en) * | 1988-03-30 | 1989-10-05 | Sunstar Inc | Skin cosmetic |
| EP0512814A1 (en) * | 1991-05-07 | 1992-11-11 | Unilever Plc | Cosmetic composition |
| JPH05246835A (en) * | 1992-03-03 | 1993-09-24 | Shiseido Co Ltd | External agent for skin |
| IN174131B (en) * | 1991-03-26 | 1994-09-24 | Lever Hindustan Ltd | |
| CN101541304A (en) * | 2006-11-30 | 2009-09-23 | 盖尔德玛公司 | Ointment compositions comprising a vitamin D derivative |
-
2020
- 2020-04-01 CN CN202010249523.5A patent/CN111743906A/en active Pending
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4610978A (en) * | 1983-03-22 | 1986-09-09 | Yissum Research Development Company Of The Hebrew University Of Jerusalem | Compositions containing 1α-hydroxycholecalciferol for topical treatment of skin disorders and methods employing same |
| JPH01249714A (en) * | 1988-03-30 | 1989-10-05 | Sunstar Inc | Skin cosmetic |
| IN174131B (en) * | 1991-03-26 | 1994-09-24 | Lever Hindustan Ltd | |
| EP0512814A1 (en) * | 1991-05-07 | 1992-11-11 | Unilever Plc | Cosmetic composition |
| JPH05246835A (en) * | 1992-03-03 | 1993-09-24 | Shiseido Co Ltd | External agent for skin |
| CN101541304A (en) * | 2006-11-30 | 2009-09-23 | 盖尔德玛公司 | Ointment compositions comprising a vitamin D derivative |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113274382A (en) * | 2021-06-16 | 2021-08-20 | 南通大学 | Application of tricin in preparation of external preparation for intervening skin photodamage |
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Application publication date: 20201009 |