CN103070849B - Application of oxyresveratrol in preparing ovary aging resistant medicine - Google Patents
Application of oxyresveratrol in preparing ovary aging resistant medicine Download PDFInfo
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- CN103070849B CN103070849B CN201310032742.8A CN201310032742A CN103070849B CN 103070849 B CN103070849 B CN 103070849B CN 201310032742 A CN201310032742 A CN 201310032742A CN 103070849 B CN103070849 B CN 103070849B
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Abstract
The invention discloses an application of oxyresveratrol in preparing an ovary aging resistant medicine. The oxyresveratrol has significant treatment effects on various ovary aging diseases such as polycystic ovarian syndrome, dysfunctional uterine bleeding, premature ovarian failure and climacteric syndrome. The oxyresveratrol is prepared into the oral, transdermal or injection medicine for treating the ovary aging diseases.
Description
Technical field
The present invention relates to the medical usage of oxidized resveratrol, oxidized resveratrol is in the application of effectively treating in multiple ovary decline disease medicament.
Background technology
The complicated bioprocess that ovary aging is a multifactor interaction, build up.Constantly consume along with day in year passing follicle, it is the main cause of ovary aging that Follicle number declines; And machine interior metabolism product is piled up microenvironment in ovary and is changed, as free radical, the damage of advanced glycation end products to follicle, and the continuous accumulation of the factor such as Mitochondrial DNA Mutation, telomere shortening, the quality of residue follicle also reduces simultaneously; The expression of series of genes and changing function in ovary, and the function of the secretion precision control ovary of H-P-O axle (Hypothalamus-Pituitary-Ovary Axis, hypothalamic-pituitary-ovarian axis) hormone.Ovarian function decline is ovary aging, and take menopause as performance, menopause in the past in 40 years old is called premature ovarian failure (Premature Ovarian Failure, POF), i.e. pathologic ovary aging.Ovarian function decline can cause ovarian dysfunction disease (as polycystic ovarian syndrome, dysfunctional uterine bleeding etc.), then exhaustion causes perimenopausal syndrome, there is each tract senile disease in these diseases: in blood circulation, coronary heart disease, cerebrovascular incidence rate obviously improve in succession, in central nervous system, easily send out presenile dementia, depression, because calcium loss sharply increases, osteoporosis is occurred frequently etc.In recent years, along with rhythm of life accelerate and learning pressure excessive, occur in ovary in the women trend that disease association disease has been significantly increased that fails.Because there are youth, middle age or elderly woman in each family, therefore the old and feeble above-mentioned disease producing of ovary has become the social problem being generally concerned about, the research of this disease has been become to focus and the difficult point of domestic and international reproductive development circle.
At present both at home and abroad research think more ovary aging and heredity, immunity, metabolism, viral infection and the factor such as iatrogenic relevant.Be summarised as the following aspects: 1 karyotype is abnormal, by the organon phase to the old and feeble patient of chromosomal abnormality ovary, some syndrome genes mainly concentrate on X chromosome and autosome, karyotype abnormal how many relevant with Inheritance, the gene mutation that environmental factors causes is in addition also the key factor that causes this disease.2 immunity factors, may with cytokine induction granular cell and lutein cell, be mainly histocompatibility complex (MHC) class antigen presentation, bring out the autoimmune response of body, granular cell and follicle are damaged relevant.In addition, think in addition ovarian function decline and ovary autoantibody generation and merge the autoimmune disease of other endocrine bodies of gland or system, as relevant in autoimmune thyroiditis, systemic lupus erythematosus (sle), myasthenia gravis, parathyroid function decline, rheumatoid arthritis, idiopathic thrombocytopenic purpura, diabetes etc.3 promoting sexual gland hormone and receptor abnormality thereof, the conduction defect of promoting sexual gland hormone FSH, LH and receptor thereof (FSHR, LHR) can cause ovarian function decline.4 azymias can cause galactosemia, and galactosemia occurs relevant with ovarian function decline.Galactose can directly damage oocyte, and its metabolite can produce infringement to ovary essence again, and the infiltration of galactose molecule also can change the activity of promoting sexual gland hormone, thereby causes the exhaustion of folliculus ovarii.5 ovary destructive factors, day by day frequent along with women's social activity, being not intended to Long Term Contact reflected ray stimulates, or accepts lonizing radiation heavy dose of or over a long time because of work, disease, contingency, all can destroy ovary.6 other dangerous factors, smoking, depression, the exogenous E of long-term a large amount of contact
2there is close relationship with nullipara and ovary decline.And impact research or the blank of heavy metal class environmental hormone on ovarian function decline, along with China's rapid development of economy, this class environmental factors will can produce long-range adverse effect to Chinese people people healthy reproduction.
Hormone replacement therapy (Hormone replacement therapy, HRT) be the important method of the old and feeble relevant symptoms of current modern medicine treatment ovary and disease, but existing research obviously shows that it has multiple side effect and carcinogenecity at a specified future date, is subject to significant restrictions its clinical practice.Corresponding this state, follow Traditional Chinese medical theory in China, advocate " the kidney invigorating tonifying YIN, soothing the liver nourishing blood " and delay ovary aging, the treatment of employing Chinese medicinal formulae, but current in the situation that also not getting its cause of disease, pathology clear, exist the mistaken ideas of abusing medicine, random treatment, and it is unclear that compound medicine treatment has effective substance basis, the defect that mechanism of action is not clear, cannot make Chinese medicine move towards modernization, moves towards internationalization.
Oxidized resveratrol (Oxyresveratrol), have another name called 4-[(E)-2-(3, 5-dihydroxy phenyl) vinyl] benzene-1, 3-glycol, belong to trans stilbene compounds, in various plants, find at present its existence, as the Fructus Artocarpi Heterophylli in Moraceae (Moraceae) belongs to (Artocarpus), Morus (Moraceae), orange Morus (Maclura), the smilax (Smilax) of Liliaceae (Liliaceae), the Gnetum (Gnetum) of Veratrum (Veraturm) and Gnetaceae (Gnetaceae) etc.Oxidized resveratrol is yellow solid, molecular formula: C
14h
12o
4molecular weight: 244.24; 199～204 ℃ of fusing points; its structure is suc as formula I; it is a kind of effectively tyrosinase inhibitor, in addition also has very strong skin pigment precipitation, and herpes, antiviral, antiinflammatory, antioxidation, neuroprotective and the effect of inhibited apoptosis in the time that cerebral ischemia occurs; owing to thering is hypotoxicity and good water solublity, be applicable to carrying out drug development.Report oxidized resveratrol for the preparation of the application in the old and feeble medicine of anti-ovary but not yet have at present.
Summary of the invention
Technical problem to be solved by this invention is to provide the application of oxidized resveratrol in the old and feeble medicine of the anti-ovary of preparation.Described oxidized resveratrol (Oxyresveratrol) has following structure:
The present invention relates to the application of the anti-ovary diseases of aging of oxidized resveratrol (Oxyresveratrol), particularly the diseases such as polycystic ovarian syndrome, dysfunctional uterine bleeding, premature ovarian failure or menopausal syndrome etc. are had to significant curative effect, and when for such use, have the safety of use and can long-term taking etc. feature.Therefore, oxidized resveratrol can be applied in the old and feeble medicine of the anti-ovary of preparation, and this medicine has the purposes of the multiple ovary decline for the treatment of disease.
Oxidized resveratrol can derive from commercially available or adopt prior art to prepare.
The oxidized resveratrol of the present invention's application can be made medicine with any one adjuvant or the drug excipient that pharmaceutically allow, its preparation can be any one dosage form pharmaceutically allowing, and includes but not limited to liquid preparation, granule, tablet, electuary, soft capsule, soft capsule, drop pill, ointment or injection.
Medicine provided by the invention, its form of medication mainly comprises oral administration, percutaneous dosing or drug administration by injection.
The dosage of oxidized resveratrol in the present invention, because of patient's state, body weight, administering mode etc. different different.It is characterized in that oxidized resveratrol and pharmaceutical carrier that described medicine contains 5-200mg.
Beneficial effect of the present invention compared with the prior art: oxidized resveratrol has significant proliferation function in vitro gonad granulocyte, can significantly improve the estrogen secretion level of in vitro gonad granulocyte, can significantly increase the estrogenic secretion of naturally-aged rat, anti-apoptotic effect to gonad granulocyte is obvious, can reduce Model of Polycystic Ovarian Syndrome Mus FSH, LH, T level, can have sex hormone regulation effect to caused by cyclophosphamide premature ovarian failure rat.Therefore, oxidized resveratrol can be applied in the old and feeble medicine of the anti-ovary of preparation, in the medicine that is particularly premature ovarian failure and resistant ovary syndrome at the anti-ovary senile symptom of preparation, applies.The present invention is the white Herba chenopodii of oxidation
Reed alcohol provides novel clinical use, has expanded its range of application.
Accompanying drawing explanation
Fig. 1 is the apoptotic index figure after the effect of flow cytometer detection of drugs.
The specific embodiment
Below with reference to specific embodiment, the present invention is further elaborated, these examples are only for illustration purpose, and are not used in the restriction scope of the invention.Laboratory animal of the present invention, purchased from Test Animal Centre, Academy of Military Medical Sciences, P.L.A (production licence number: SCXK (army) 2007-004), selects the clinical estradiol valerate that has good therapeutic effect to do positive control.Oxidized resveratrol, press document (Fu great Xu, Chen Lei, Hou Aijun, Yao Qing, Zhang Wenyu. the chemical constitution study of black Mulberry. Chinese herbal medicine, 2005,36 (9): 1296-1299) methods Laboratory self-control, purity >=98% (HPLC), the water-soluble administration concentration that is mixed with.
The proliferation function (mtt assay) of [embodiment mono-] oxidized resveratrol in vitro gonad granulocyte
Get 22-27 age in days female rats, after the conventional 48h of raising, cervical vertebra dislocation method is put to death, the aseptic bilateral ovaries of winning rat, granular cell is discharged ovary fragmentation, in DMEM-F12 culture fluid with 2mL containing penicillin and streptomycin, dilute, and piping and druming gently, make it be dispersed into individual cells.Centrifugal filtration collecting granules cell.By finite concentration cell suspension inoculation in 24 orifice plates, every hole 80 μ l, every hole add again 120 μ l culture fluid adjust cell density to 200 μ l/ holes, establish 6 parallel holes for every group.Be placed on 5%CO
2in incubator, cultivate 24h.Get CO
2in incubator, cultivated the granular cell suspension of 24h, every hole successively adds modeling concentration Caddy (Cleary) and variable concentrations oxidized resveratrol sample (1.0mgmL
-1, 0.1mgmL
-1, 0.01mgmL
-1), parallel setting Normal group (normal saline) and model group (Caddy (Cleary) and normal saline).After cellar culture 20h, in every hole, add MTT solution (5mgmL
-1) 20 μ L continuation cultivation 4h, every hole adds three liquid of 80 μ L, and incubated overnight is selected 570nm wavelength, measures each hole absorbance on enzyme-linked immunosorbent assay instrument, the results are shown in Table 1.After result shows positive drug, the effect of various dose oxidized resveratrol, in vitro gonad granulocyte cultivation effect all has significant difference (all p < 0.01) with model group.
The proliferation function of table 1 oxidized resveratrol in vitro gonad granulocyte
*p < 0.05,
*p < 0.01vs model group.
Δp < 0.05,
Δ Δp < 0.01vs Normal group
The estrogen and progestogen secretory action (radioimmunology) of [embodiment bis-] oxidized resveratrol in vitro gonad granulocyte
Get 22-27 age in days female rats, after the conventional 48h of raising, cervical vertebra dislocation method is put to death, the aseptic bilateral ovaries of winning rat, granular cell is discharged ovary fragmentation, in DMEM-F12 culture fluid with 2ml containing penicillin and streptomycin, dilute, and piping and druming gently, make it be dispersed into individual cells.Centrifugal filtration collecting granules cell.By finite concentration cell suspension inoculation in 24 orifice plates, every hole 80 μ l, every hole add again 120 μ l culture fluid adjust cell density to 200 μ l/ holes, establish 6 parallel holes for every group.Be placed on 5%CO
2in incubator, cultivate 24h.Get CO
2in incubator, cultivated the granular cell suspension of 24h, every hole successively adds modeling concentration Caddy (Cleary) and variable concentrations oxidized resveratrol sample, parallel setting Normal group (normal saline) and model group (Caddy (Cleary) and normal saline).Be placed on 5%CO
2in incubator, cultivate 24h.Its cell conditioned medium liquid of centrifuging and taking, measures E with radioimmunological kit
2, P content, the results are shown in Table 2.Result display model group and Normal group all have significant difference (p < 0.01), after positive drug, the effect of various dose oxidized resveratrol, in vitro gonad granulocyte estrogen secretion level and model group have significant difference (p < 0.01), and after the effect of various dose oxidized resveratrol, in vitro gonad granulocyte Progesterone Secretion level and model group do not have significant difference.
The estrogen and progestogen secretory action of table 2 oxidized resveratrol in vitro gonad granulocyte
*p < 0.05,
*p < 0.01vs model group.
Δp < 0.05,
Δ Δp < 0.01vs Normal group
The impact (radioimmunology) of [embodiment tri-] oxidized resveratrol on the secretion of naturally-aged rat estrogen and progestogen
Adopt the senilism model of naturally-aged.Choose 14 month female SD rats, body weight (270 ± 30) g, Animal adaptability was fed after one week, make colpocytology smear, 4 oestrous cycles of Continuous Observation, with the colpocytology performance oestrous cycle extend, Constant Estrus, pseudopregnancy cell phase time repeatedly afterwards, as female aged model success.Naturally-aged rat is divided into model group, oxidized resveratrol (3mg/kg, 6mg/kg, 12mg/kg) group and positive drug group at random, every group each 10, separately getting 11 of 4 month female rats is Normal group, every day gavage 1 time, Normal group and model group give isometric(al) normal saline, after successive administration 30 days, before administration and after last administration, get blood from rat eye socket venous plexus, centrifugal, get serum, the content of measuring respectively estrogen and progesterone in serum with radioimmunology, the results are shown in Table 3.Result display model group and Normal group all have significant difference (p < 0.01), gavage positive drug after 30 days estrogen and progestogen all have significant difference (p < 0.01) with model group, and oxidized resveratrol (high, normal, basic dosage) promotes estrogen secretion after 30 days and model group has significant difference (p < 0.01), though Progesterone Secretion ability is improved, with model group there was no significant difference.
The comparison of table 3 oxidized resveratrol group and matched group estrogen and progestogen level
*p < 0.05,
*p < 0.01vs model group.
Δp < 0.05,
Δ Δp < 0.01vs Normal group
The impact (flow cytometry) of [embodiment tetra-] oxidized resveratrol on naturally-aged Cultured Granulosa apoptosis
Adopt the first old model of naturally-aged.Choose 14 month female SD rats, body weight (270 ± 30) g, Animal adaptability was fed after one week, make colpocytology smear, 4 oestrous cycles of Continuous Observation, with the colpocytology performance oestrous cycle extend, Constant Estrus, pseudopregnancy cell phase time repeatedly afterwards, as female aged model success.Naturally-aged rat is divided into model group at random, oxidized resveratrol (3mg/kg, 6mg/kg, 12mg/kg) group and positive drug group, every group each 10, separately getting 11 of 4 month female rats is Normal group, every day gavage 1 time, Normal group and model group give isometric(al) normal saline, after successive administration 30 days, after last administration, collect gonad granulocyte, carry out flow cytometer detection (seeing Fig. 1), analyze the relation with apoptosis of administration and gonad granulocyte, result shows: in apoptosis figure, UL represents dead cell, UR represents apoptosis in late period, LL represents living cells, LR represents early apoptosis, UR+LR is apoptosis total value.Figure 1A (Normal group) apoptosis value is 25.70, Figure 1B (model group) apoptosis value is 86.65, Fig. 1 C (positive controls) apoptosis value is 47.04, Fig. 1 D (oxidized resveratrol high dose group) apoptosis value is 29.78, Fig. 1 E (dosage group in oxidized resveratrol) apoptosis value is 46.05, Fig. 1 F (oxidized resveratrol low dose group) apoptosis value is 55.43, according to the visible positive drug of numerical value, oxidized resveratrol (high, normal, basic dosage) after 30 days the anti-apoptotic effect to gonad granulocyte obvious.
The impact of [embodiment five] oxidized resveratrol on Model of Polycystic Ovarian Syndrome rat
24d age teenage SD female rats, body weight 30-40g, 0.5mg18-methylnorethindron day 1 subcutaneous injection, in the time of 27d age, add with HCG1.5IU day 2 subcutaneous injections, inject altogether 21d.After modeling success, be divided at random model group, sample (high, normal, basic dosage) group and positive drug group (metformin 0.018g/d), every group each 10, separately getting 10 of 45d SD in age female rats is Normal group, every day gavage 1 time, Normal group and model group give isometric(al) normal saline, successive administration is after 15 days, after last administration, get blood examination and survey FSH, LH, T concentration in serum, the results are shown in Table 4.Result display model group and Normal group all have significant difference (p < 0.01), gavage positive drug, oxidized resveratrol (high, normal, basic dosage) after 15 days FSH, LH, T level all have the difference of different significances from model group.
The comparison of FSH, LH, T concentration in table 4 oxidized resveratrol group and matched group serum
*p < 0.05,
*p < 0.01vs model group.
Δp < 0.05,
Δ Δp < 0.01vs Normal group
The impact of [embodiment six] oxidized resveratrol on premature ovarian failure sex hormone level in rats
Adopt 9d SD in age female rats, 14 days cyclophosphamide 8mg/ (kgd) of lumbar injection modeling continuously.After modeling success, be divided at random model group, sample (high, normal, basic dosage) group and positive drug group (premarin 0.075mg/kg), every group each 10, separately getting 10 of 23d SD in age female rats is Normal group, every day gavage 1 time, Normal group and model group give isometric(al) normal saline, and successive administration, after 35 days, is got blood examination and surveyed FSH, E in serum after last administration
2concentration, the results are shown in Table 5.Result display model group and Normal group all have significant difference (p < 0.01), gavage positive drug, oxidized resveratrol (senior middle school's dosage) FSH, E after 35 days
2level all has the difference of different significances from model group.
FSH, E in table 5 oxidized resveratrol group and matched group serum
2the comparison of concentration
*p < 0.05,
*p < 0.01vs model group.
Δp < 0.05,
Δ Δp < 0.01vs Normal group
[embodiment seven] oxidized resveratrol capsule
Oxidized resveratrol 100g, starch 80g, carboxymethyl starch sodium 24g, dextrin 17g, 70% appropriate amount of ethanol, makes 1000 (every containing oxidized resveratrol 100mg).
[embodiment eight] oxidized resveratrol capsule
Oxidized resveratrol 15g, starch 10g, carboxymethyl starch sodium 4g, dextrin 12g, 70% appropriate amount of ethanol, makes 1000 (every containing oxidized resveratrol 15mg).
[embodiment nine] oxidized resveratrol sheet
Oxidized resveratrol 200g, lactose 55g, amylum pregelatinisatum 15g, hyprolose 10g, magnesium stearate 5g, 70% appropriate amount of ethanol, makes 1000 (every containing oxidized resveratrol 200mg).
[embodiment ten] oxidized resveratrol granule
Oxidized resveratrol 6g, lactose powder 800g, magnesium stearate 194g, 70% appropriate amount of ethanol, makes 1000 grams of granules (containing oxidized resveratrol 6mg in every g granule system).
[embodiment 11] oxidized resveratrol soft capsule
Oxidized resveratrol 5g, soybean oil 10g, gelatin 5g, glycerol 5g, distilled water 7g, makes 1000 (every containing oxidized resveratrol 5mg).
[embodiment 12] oxidized resveratrol emulsifiable paste
Oxidized resveratrol 0.5g, stearic acid 100g, hexadecanol 20g, glyceryl monostearate 10g, white oil 10g, methyl hydroxybenzoate 0.8g, butyl hydroxybenzoate 0.2g, glycerol 140g, potassium hydroxide 5g, ethanol 10g, distilled water adds to 1000g.
[embodiment 13] oxidized resveratrol emulsifiable paste
Oxidized resveratrol 0.5g, stearic acid 100g, hexadecanol 20g, glyceryl monostearate 10g, white oil 10g, methyl hydroxybenzoate 0.8g, butyl hydroxybenzoate 0.2g, glycerol 140g, potassium hydroxide 5g, ethanol 10g, distilled water adds to 1000g.
[embodiment 14] oxidized resveratrol gel
Oxidized resveratrol 0.5g, PEG400450g, hexanediol 80g, Tween 80 20g, carbomer 30g, ethanolamine 6ml, benzyl alcohol 10g, distilled water adds to 1000g.
[embodiment 15] oxidized resveratrol injection
Oxidized resveratrol 25.0g, is dissolved in 1000ml water for injection, fully stirring and dissolving, benefit injects water to 2000ml, add pin activated carbon 2.5g, be heated to 60 ℃ and stir 30 minutes, carbon-point filters, filtrate is through 0.25 μ m filtering with microporous membrane degerming, be sub-packed in 1000 cillin bottles, loading amount 2.0ml/ props up, sealing, sterilizing, to obtain final product.
Claims (5)
1. the application of oxidized resveratrol in the old and feeble medicine of the anti-ovary of preparation.
2. according to the application of claim 1, it is characterized in that described ovary senile symptom is polycystic ovarian syndrome, premature ovarian failure or menopausal syndrome.
3. application according to claim 1, is characterized in that oxidized resveratrol and pharmaceutical carrier that described medicine contains 5-200mg.
4. application according to claim 1, the dosage form that it is characterized in that described medicine is any dosage form of pharmaceutically approving.
5. application according to claim 4, is characterized in that the dosage form of described medicine comprises granule, tablet, electuary, soft capsule, soft capsule, drop pill, ointment or injection.
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