CN105267187A - Medicinal composition for treating fatty liver and preparation method and application thereof - Google Patents
Medicinal composition for treating fatty liver and preparation method and application thereof Download PDFInfo
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- CN105267187A CN105267187A CN201410794496.4A CN201410794496A CN105267187A CN 105267187 A CN105267187 A CN 105267187A CN 201410794496 A CN201410794496 A CN 201410794496A CN 105267187 A CN105267187 A CN 105267187A
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- resveratrol
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Abstract
The invention relates to a medicinal composition for treating fatty liver and a preparation method and application thereof. Use of resveratrol and curcumin for the manufacture of a pharmaceutical composition for the treatment of a fatty liver patient, and when the pharmaceutical composition is administered to the fatty liver patient, the curcumin and the resveratrol provide a combined therapeutic effect. The invention also discloses a pharmaceutical composition containing the two and a preparation method thereof. By utilizing the application, the medical composition and the preparation method thereof, the curative effect on fatty liver patients can be improved.
Description
[technical field]
The present invention relates to a kind of medical composition, the preparation method of medical composition and the purposes of manufacture medical composition.
[background technology]
Along with socioeconomic lifting, sanitary improvement, with the progress of medical treatment, popular and the exquisite cuisines of west fast food culture prevailing, the healthy index of modern is made often to have blood pressure, high blood glucose and dyslipidemia etc., and this kind of many metabolic risk factors clustering phenomenon, be referred to as metabolism syndrome.Wherein, metabolism syndrome not only has closely related with chronic diseases such as general common cerebrovascular disease, heart disease, diabetes and hypertension, also can cause the disease of liver, such as fatty liver.
Fatty liver is wherein a kind of clinical symptom of metabolism syndrome, it refers to the excessive accumulation of fat in liver, when its reason formed is human body excess intake fatty, and the burden of hepatocyte digestion fat, make it cannot remove superfluous fat, and the lipid storage in hepatocyte is increased gradually.Or be, hepatocyte impaired (as medicine, be addicted to drink, viral hepatitis etc. causes) and the metabolic function of fat of can not bringing into normal play time, make the fatty abnormal stacking in liver.And when the fat content of liver exceedes 5% of liver gross weight, then can be referred to as fatty liver.On clinical medicine, loss of weight and motion are the fatty liver Therapeutic Method uniquely confirmed the validity at present, current rare use Drug therapy fatty liver.Although hypoglycemic drug, liver protect agent, antioxidant etc. all the progress studied on the rear liver index of display treatment or pathology, but still need more research to confirm its curative effect, be not identified effective fatty liver medicine at present.
The aldehydes matter that resveratrol (Resveratrol) is non-flavonoid.Natural resveratrol exists in a lot of plant, is the antitoxin type material of one that plant produces to resist pathogenic bacteria invasion.And the same with the polyphenol substance of most of flavonoid, resveratrol also has very strong anti-oxidation function, and cholesterol can be stoped in the deposition of blood vessel, and then prevents the symptom of the metabolism syndromes such as cardiovascular disease.
On the other hand, curcumin (Curcumin) is by the material extracted in the rhizome of Rhizoma Curcumae Longae (Turmeric).And the extract of Rhizoma Curcumae Longae mainly comprises three kinds of compositions, major part is curcumin (Curcumin), comparatively small part is demethoxycurcumin (DemethoxyCurcumin) and 1,7-bis(3,4-dimethoxyphenyl)-1,6-heptadiene-3,5-dione (BisdemethoxyCurcumin), and demethoxycurcumin and 1,7-bis(3,4-dimethoxyphenyl)-1,6-heptadiene-3,5-dione also can be described as the derivant of curcumin, curcumin of the present invention also comprises its derivant.
Curcumin is used in food industry as a kind of natural pigment very early; and on medical domain; more find that curcumin belongs to the curcumin of polyphenol compound, has pharmacological action widely, the effects such as such as anti-inflammatory, antioxidation, scavenging free radicals, cardiovascular protection.Especially the resistance to oxidation of curcumin is 10 times of vitamin E, therefore is considered to have potentiality, can be developed to the medicine of the metabolism syndrome symptom such as diabetes, hyperlipidemia, heart disease.In addition, curcumin also can increase the secretion of bile, promotes the decomposition of fat.
Conclude, confirm that resveratrol or curcumin have stronger anti-oxidation function at present, therefore it has the curative effect of mitigation to a certain degree respectively for metabolism syndrome, in other words, using resveratrol or curcumin as in the clinical practice of medicine at present, major part or get individually effect of its antioxidation and anti-inflammatory, to be applied to aging resistance or to slow down the deterioration etc. of metabolism syndrome.But, for there is in metabolism syndrome the patient of fatty liver, be not identified effective fatty liver medicine at present.
[summary of the invention]
Owing to not being identified effective fatty liver medicine at present, though and have and confirmed that it has the curative effect of mitigation to a certain degree respectively for metabolism syndrome compared with the resveratrol of Strong oxdiative function and curcumin, but, whether resveratrol and curcumin can be applicable to treat the patient in metabolism syndrome with fatty liver symptom, and the curative effect had to a certain degree, for the technical field of the invention institute is for the person of knowing.Therefore, the present invention proposes a kind of by resveratrol and curcumin merging medical composition being used in Patients with Fatty Liver and preparation method thereof and purposes according to this, effectively promotes the therapeutic effect for fatty liver.
Specifically, the invention provides a kind of medical composition for the treatment of Patients with Fatty Liver, it comprises the pharmaceutically acceptable carrier of resveratrol and curcumin and at least one, diluent or excipient.Wherein, medical composition bestows Patients with Fatty Liver, and resveratrol and curcumin provide the effect of combined treatment.
The present invention separately provides a kind of method preparing the medical composition for the treatment of Patients with Fatty Liver, comprises following steps: weigh the resveratrol of a unit dose and the curcumin of a unit dose; The pharmaceutically acceptable carrier of emulsifying resveratrol, curcumin and at least one, diluent or excipient form the first mixture; And homogenization first mixture form the second mixture.Wherein, medical composition bestows Patients with Fatty Liver, and resveratrol and curcumin provide the effect of combined treatment.
The present invention provides again a kind of resveratrol and curcumin for the manufacture of the purposes of the medical composition for the treatment of Patients with Fatty Liver, and when medical composition is cast Patients with Fatty Liver, resveratrol and curcumin provide the effect of combined treatment.
Can know present technical characteristic of the present invention for making it rear content, below intend first defining specific term, then further illustrate content of the present invention.In the present invention, resveratrol and curcumin merge use, and be used for the treatment of the patient with fatty liver, and the effect of combined treatment is provided, and " combined treatment " refers to use resveratrol and these two kinds of compositions of curcumin simultaneously, and occupation mode can make one " medical composition ", it comprises resveratrol and curcumin two kinds of compositions simultaneously.And " medical composition " of the present invention also can comprise and make a kind of combination drug or composition of medicine, it comprises two kinds of medical things simultaneously, and its one has resveratrol ingredient, and another kind has curcumin composition, and uses this two kinds of medical things simultaneously.Preferably, medical composition has the resveratrol of effective dose and the curcumin of effective dose respectively, and wherein, described effective dose refers to that both can produce the dosage of synergism and combined treatment.In the present embodiment, resveratrol and curcumin are mixed and made into medical composition, and preserve in the mode of capsule, and all the other minutias describe in detail hereinafter.
When the fat content that used in this manual " Patients with Fatty Liver " word means the liver of patient exceedes 5% of its liver gross weight, be the patient with fatty liver.Generally speaking, fatty liver refers to there is athero in hepatocyte, and be commonly called as " powder liver " or " liver bag oil ", fatty liver is the disease that a kind of multi-pathogenesis but has similar pathological change, can be divided into alcoholic fatty liver and non-alcoholic fatty liver disease.And " Patients with Fatty Liver " of the present invention comprises alcoholic fatty liver and non-alcoholic fatty liver disease simultaneously.Wherein, non-alcoholic fatty liver disease is formed and is mainly adipose metabolism extremely, and main cause is that the accumulation of lipid is unbalance with discharge, and lipid accumulation becomes main cause with the absorption and the intercrescence of increase lipid that increase lipid; To discharge the factor reduced be then synthesis for reducing lipoprotein with secretion with reduce the oxidation of lipid.Generally speaking, metabolic disease is normal relevant with abnormal lipid synthesis speed, therefore when liver damage, fatty acid synthetase (FattyAcidSynthase) performance amount in liver cell is greater than the fatty acid synthetase in normally liver cell, and then causing the situation of lipid accumulation, severe patient then can cause the formation of fatty liver.And in experimental example of the present invention, can learn that resveratrol and curcumin regulate and control fatty acid synthetase jointly, namely suppress the performance of fatty acid synthetase, to treat Patients with Fatty Liver.
In addition, used in this manual " synergism " means when merging gives two or more material, its effect produced is equal to or greater than (comprise the response to treatment producing same nature, or reduce untoward reaction) summation of the effect that each one matter produces alone.In experimental example of the present invention, known resveratrol and curcumin merging gives Patients with Fatty Liver, its therapeutic effect produced, more single resveratrol or the therapeutic effect of curcumin better.Used in this manual " treatment " one word means to alleviate, releive, change, improve, improve or to affect cancer and symptom thereof, for example, suppresses or pain that Therapeutic cancer causes or hemorrhage or delay or tumor proliferative speed of reversing.
Used in this manual " resveratrol " one word is except can referring to resveratrol, also comprise basic resveratrol structure modify through chemistry or biology or replace the derivant produced, and still possess the character of former basic resveratrol structure or there is the character similar to former basic resveratrol structure.Certainly, " resveratrol " one word also comprise the pharmaceutically acceptable salt of resveratrol, it is formed according to the acid in resveratrol chemical constitution or base part.
Same, " curcumin " as used in this specification one word except can referring to curcumin, also comprise basic curcumin structure modify through chemistry or biology or replace the derivant produced, and still possess the character of former basic curcumin structure or there is the character similar to former basic curcumin structure, such as aforesaid demethoxycurcumin and 1,7-bis(3,4-dimethoxyphenyl)-1,6-heptadiene-3,5-dione.Certainly, " curcumin " one word also comprise the pharmaceutically acceptable salt of curcumin, it is formed according to the acid in curcumin chemical constitution or base part.
In addition, have in this manual and mention and the compound relevant to the technology of the present invention feature essence or material, all can contain any pharmaceutically acceptable form of alleged compound or material and its.Wherein, pharmaceutically acceptable form can such as but not limited to containing the various isomers (isomer) of diastereomer (diastereomer) and enantiomer (enantiomer), salt, free form, solvent, prodrug (prodrug), polymorph (polymorph) and racemic mixture (racemicmixture).
In the present invention, when resveratrol and curcumin merge use, the dosage of curcumin is better between 100 to 1000 milligrams, and the dosage of resveratrol is better between 100 to 1000 milligrams.Again in the present invention, the weight percent that resveratrol and curcumin merge when using is better between 1:1 to 1:5, in the present embodiment, for the percentage by weight of resveratrol and curcumin for 1:1.3 illustrates.Certainly, above-mentioned effective dose and percentage by weight also can be the component ratio in medical composition, or medical cover group actual use time modulation ratio.
In sum, according to purposes of the present invention, medical composition and preparation method thereof, it merges resveratrol and curcumin uses, and for improvement or enhancement treatment Patients with Fatty Liver, provides significant effect.
[accompanying drawing explanation]
Fig. 1 is a kind of method flow diagram preparing the medical composition for the treatment of Patients with Fatty Liver of one embodiment of the invention.
Fig. 2 A and Fig. 2 B be human hepatocarcinoma cells's strain HepG2 in experimental example two born of the same parents in the result figure of triglyceride content.
Fig. 3 is the result figure of the fatty acid synthetase content of human hepatocarcinoma cells's strain HepG2 in experimental example three.
[embodiment]
Below will coordinate accompanying drawing that embodiments of the invention and experimental example are described, only relevant context explanation can refer to aforementioned, does not repeat them here.
According to a kind of resveratrol of the present invention and the curcumin purposes for the manufacture of the medical composition for combined treatment Patients with Fatty Liver, in one embodiment, can by both be prepared into a medical composition jointly, then give patient and take to reach.Wherein, the extracting process of resveratrol and curcumin is person with usual knowledge in their respective areas of the present invention institute energy understanding person, does not repeat them here.Above-mentioned medical composition can be dosage form that is solid-state, liquid or semi liquid state, preferably, is the dosage form of liquid state, and can prepare described medical composition according to a kind of method preparing the medical composition for the treatment of Patients with Fatty Liver of the present embodiment.
As shown in Figure 1, Fig. 1 is a kind of method flow diagram preparing the medical composition for the treatment of Patients with Fatty Liver of one embodiment of the invention, and described preparation method comprises following steps: weigh the resveratrol of a unit dose and the curcumin (S10) of a unit dose; The pharmaceutically acceptable carrier of emulsifying resveratrol, curcumin and at least one, diluent or excipient form the first mixture (S20); And homogenization first mixture form the second mixture (S30); And the second mixture is suppressed in soft capsule (S40).Wherein, medical composition bestows Patients with Fatty Liver, and resveratrol and curcumin provide the effect of combined treatment.
Generally speaking, exist in the mode of powder after resveratrol and curcumin extraction, the unit dose of resveratrol is between 100mg to 1000mg, and the effective dose of curcumin is between 100mg to 1000mg.In step slo, weigh the resveratrol of a unit dose and the curcumin of a unit dose, refer to the medical composition that can contain a dosage unit in each formula, in other words, a formula, namely containing the dosage that can produce therapeutic effect completely to Patients with Fatty Liver, directly casts to facilitate.In the present embodiment, the resveratrol of a unit dose is 100mg, and the curcumin of a unit dose is 130mg is example, therefore weighs the resveratrol extract of the purity 98% of 102mg, and weighs the curcumin extract of purity 95% of 138mg.In force, once can prepare the medical composition of multiple dosage unit, for example, if for the medical composition once preparing 100 formula, then can weigh the curcumin of 100 unit dose and the resveratrol of 100 unit dose in step S10, to carry out the preparation process continued, the present embodiment for a unit dose, but not as limit.
The person of connecing, in step S20, the pharmaceutically acceptable carrier of emulsifying resveratrol, curcumin and at least one, diluent or excipient form the first mixture.In the present embodiment, pharmaceutically acceptable carrier, diluent or excipient comprise hydrophobic liquid, namely the material of oils is commonly referred to as, and preferably, the oily substance of hydrophobic liquid for being made up of polybasic unsaturated fatty acid, at the present embodiment for Semen Lini oil, certainly, in other embodiments, can be also fish oil, Oleum sesami or Folium Perillae wet goods, the present invention be as limit.In addition, carrier, diluent or excipient more comprise other waterborne liquid, such as pure water, or can be used as low-carbon (LC) alcohols or the gelatin solution of surfactant.Therefore, hydrophobic liquid is added pulverous resveratrol, curcumin by step S20, preferably, by stirring hydrophobic liquid and pulverous resveratrol and curcumin emulsifying to form the first mixture.In other words, the first mixture is the mixture after resveratrol and curcumin and hydrophobic liquid emulsifying.Preferably, the viscosity of the first mixture is less than 100dPa.s (decipascal-second).
Preferably, hydrophobic liquid is more than or equal to 60% for the percentage by weight of resveratrol and curcumin, in other words, if the percentage by weight of hydrophobic liquid and resveratrol and curcumin mixture is considered as 100%, percentage by weight then shared by hydrophobic liquid is more than or equal to 60%, and the percentage by weight of resveratrol and curcumin mixture is less than or equal to 40%, and in the present embodiment, the percentage by weight of hydrophobic liquid is 60%, and the percentage by weight of resveratrol and curcumin mixture is 40% for example.
Because resveratrol and curcumin are Powdered at the present embodiment, and there is the two-phase stuff and other stuff of multiple size inequality, i.e. the two-phase stuff and other stuff of liquid (hydrophobic liquid) coated solid-state (resveratrol and curcumin powder) with the first mixture that (step S20) after hydrophobic liquid emulsifying is formed.Therefore preferably, between step S10 and step S20, first resveratrol and curcumin can be carried out suitable combination process, to guarantee, in each two-phase stuff and other stuff in the first mixture, there is more impartial resveratrol and curcumin.Certainly, in other embodiments, be not limited with resveratrol and the modulation of curcumin two material Homogeneous phase mixing, in other words, in the medical composition of same one dosage type low temperature, resveratrol can have mixing of other degree with curcumin, also can even not mix, for example, medical composition can be a kind of soft capsule, wherein part is resveratrol, and another part is curcumin, carries out step S20 and the step S30 be described in detail below and step S40 respectively by resveratrol and curcumin.
And after step S20 emulsifying the first mixture of gained, the size of its two-phase stuff and other stuff is comparatively large, and by making made medical composition solution be absorbed by human body, must reduce the size of two-phase stuff and other stuff.Therefore in step s 30, homogenization first mixture also forms one second mixture, namely by the two-phase stuff and other stuff of mode refinement first mixture of homogenization, the second mixture formed then has the two-phase stuff and other stuff of reduced size.And in the mode of operation " homogenization ", general common on the market homogenizer can be used, or there is the emulsifying homogeneous machine of emulsifying or homogenizing function simultaneously, and pass through rotating speed or the pressure of adjustment homogenizer or emulsifying homogeneous machine, to form the two-phase stuff and other stuff that particle diameter is greater than 100 nanometers.And described herein " homogenization " refer to the size of the two-phase stuff and other stuff of the second mixture to control between 100 to 1000 nanometers, between the particle size size namely after emulsifying and after homogenizing.Generally speaking, the particle after emulsifying compares the two-phase stuff and other stuff of second mixture of the present embodiment, and the particle diameter of the particle after emulsifying is comparatively large, therefore the speed be absorbed by the body is slower; And the particle after homogenizing compares the two-phase stuff and other stuff of second mixture of the present embodiment, its homogenize after the particle diameter of particle less, though infiltration rate is very fast, but also easily discharged by body metabolism, therefore the particle diameter of the two-phase stuff and other stuff of second mixture of the present embodiment is between emulsifying and homogenizing, the medical composition that preparation is completed is absorbed by the body and the speed of metabolism can maintain preferably in scope, comparatively complete to help absorption of human body.
Preferably, the method for the medical composition of preparation treatment Patients with Fatty Liver can comprise step S40 further, is suppressed in soft capsule by the second mixture.Specifically, the carrier of the present embodiment, diluent or excipient comprise gelatin solution further, and gelatin solution is except can be used as surfactant, can be used as the composition forming soft capsule shell simultaneously.In force, second mixture of step S30 gained is inserted vacuum (or negative pressure) epoxy glue device, and carry out the program of colloidal sol, namely in vacuum (or negative pressure) and temperature is under the environment of 60 ~ 80 DEG C, continue to mix, and mixture is now called colloidal sol, and after the viscosity of colloidal sol is 13000 to 15000CPS, then colloidal sol is inserted in mould compressing.In force, according to each institute for the soft capsule number of compacting, resveratrol and the curcumin of corresponding unit dose can be weighed in step S10, to form the medical composition that namely each soft capsule contains a dosage unit.Certainly, in other embodiments, a dosage unit also can be made up of the multiple medical compositions be scattered in several dosage units or secondary packaging, such as, to be scattered in two to three soft capsules but to be packaged in same blister plastic packaging (blisterpack).
As previously mentioned, when resveratrol and curcumin merging are used in treatment Patients with Fatty Liver, the dosage range of resveratrol controls about between 100mg to 1000mg, and the dosage range of curcumin control about between 100mg to 1000mg for good, and the percentage by weight of resveratrol and curcumin being good between 1:1 to 1:5.In above-mentioned scope, the therapeutic effect produced between each dosage may have drop, following experimental example with the percentage by weight of resveratrol and curcumin for about 1:1.3 treats.Certainly, resveratrol or curcumin dosage may with coordinate treatment on active ingredient, cast path or individuality in need and physiological situation thereof difference and change to some extent.General, oral way needs higher dosage, reviews the dosage that the treatment initial stage uses then relatively low.
Further illustrate, according to purposes of the present invention, after medical composition casts Patients with Fatty Liver, wherein can there is synergism in resveratrol and curcumin, to improve or to promote the effect being used alone resveratrol or curcumin and treat fatty liver, and then reach elimination, suppression, improvement, alleviation, prevent fatty liver and symptom thereof or reach the medical effect similar to above-mentioned purpose.Specifically, resveratrol and curcumin, by jointly regulating and controlling fatty acid synthetase, suppress the performance of fatty acid synthetase, to treat Patients with Fatty Liver.
The present invention provides again a kind of medical composition and manufacture method thereof for the treatment of Patients with Fatty Liver, and it comprises the pharmaceutically acceptable carrier of resveratrol and curcumin and at least one, diluent or excipient.But, described medical composition and manufacture method roughly the same with the purposes of the described medical composition of aforementioned explanation, with reference to aforementioned, can not repeat them here.Only be noted that, though resveratrol and curcumin can be formed in different soft capsules respectively in medical composition, but resveratrol and curcumin are still and will be limited to merge use, then, alleged merging uses one of them can cast patient in advance at both, before not ineffective effect, cast both another kinds separately; Certainly preferably both cast patient without time difference simultaneously, or one of them cast after cast alternative mode immediately and implement.
From the above, according to purposes of the present invention, medical composition and manufacture method thereof, it merges resveratrol and curcumin uses, and for improvement or enhancement treatment Patients with Fatty Liver, provides significant effect.
Experimental example one: prepare medical composition
As shown in Figure 1, the resveratrol of 100 milligrams and the curcumin (S10) of 130 milligrams can be weighed; The pharmaceutically acceptable carrier of emulsifying resveratrol, curcumin and at least one, diluent or excipient form the first mixture (S20); And homogenization first mixture form the second mixture (S30).Wherein, medical composition bestows Patients with Fatty Liver, and resveratrol and curcumin provide the effect of combined treatment.
In step slo, resveratrol used in the present invention is purchased from Li Tong biotech inc, and curcumin is purchased from Li Tong biotech inc.After obtaining, at room temperature weigh purity 98% resveratrol of about 102 milligrams and purity 95% curcumin of 138 milligrams respectively, be about resveratrol and the curcumin of 1:1.3 ratio mixing powdery according to percentage by weight, be a unit dose.Then, in step S20, the pharmaceutically acceptable carrier of at least one, diluent or excipient comprise Semen Lini oil (hydrophobic liquid), and use 1 ~ 2% fatty glyceride for emulsifying agent, and resveratrol, curcumin and Semen Lini oil emulsifying are formed the first mixture.Then, in step s 30, aforesaid first mixture is inserted in high-speed vacuum emulsifying mixer and carries out homogenization, to obtain second mixture of size between 100 to 1000 nanometers, and the second mixture is packaged as the form (step S40) of soft capsule, under being stored in room temperature.
Experimental example two: resveratrol and curcumin medical composition have synergistic inhibition for cytolipin synthetic reaction
Utilize tissue culture technique by human liver cancer (Hepatocellularcarcinoma, Human) Cell Line HepG2 is cultured to right quantity in the cell culture fluid DMEM of the hyclone (BovineSerum, FBS) containing concentration 10%.Then, get containing quantity 2 × 10
4the suspension of HepG2 cell, and be seeded in 96 holes (96-wells) culture dish, cultivate after 16 hours, again original culture fluid is taken out, to be replaced as the cell culture fluid DMEM containing 30mM glucose, add the resveratrol (100 to 50 μMs) of variable concentrations, curcumin (160 to 40 μMs) respectively, and the compositions of resveratrol and curcumin.
In this experimental example, be divided into four kinds of variable concentrations (in this experimental example, referring to ultimate density) and test.Wherein, the molecular weight of resveratrol is about 228g/mol, and the molecular weight of curcumin is about 368g/mol, and the percentage by weight being converted into resveratrol and curcumin is 1:1.3 to 1:2.6.The component dose of each group arranges as shown in the table:
After Cell Line HepG2 adds above-claimed cpd respectively, then the incubator of the culture dish of group 1,2 and group 3,4 being inserted 37 DEG C continues cultivation 48 hours, carries out lipid synthesis test afterwards.This experimental example is to detect the content of triglyceride (HepG2intracellularTriglyceride) in Cell Line HepG2 born of the same parents.
Specifically, after terminating aforementioned cultivation, sequentially the culture fluid in each hole and the compound added are absorbed, separately with normal saline solution cleaning, preferably clean secondary, then add the lipid extraction liquid of 100 μ L in each hole, whole culture dish dish is placed in 95 DEG C and treats 30 minutes, lipid extraction liquid is dissolved.After aforementioned 30 minutes, take out culture dish and rock one minute, take out the lipid extraction liquid 50 μ l that each hole is dissolved again respectively, the lipid extraction liquid dissolved in each hole of culture dish is made to be similarly 50 μ l, again by triglyceride reaction mixture (purchased from Abnova-KA0902, AdipogenesisAssaykit) 50 μ L add in each hole of culture dish, after mix homogeneously, put into the incubator of 37 DEG C and lucifuge reacts 30 minutes, with ferment immunoassay plate reading with wavelength 570nm for benchmark, detect the light absorption value in each hole in culture dish.
Fig. 2 A and Fig. 2 B be human hepatocarcinoma cells's strain HepG2 in experimental example two born of the same parents in the result figure of triglyceride content.From Fig. 2 A, the content of triglyceride in Cell Line HepG2 born of the same parents, when merging use 50 μMs of resveratrols and 40 μMs of curcumins, the Rhizoma Curcumae Longae being obviously comparatively used alone 50 μMs of resveratrols or being used alone 40 μMs usually lower.Same, when merging the curcumin of use 100 μMs of resveratrols and 80 μMs, it generates the inhibition of triglyceride for Cell Line HepG2, the Rhizoma Curcumae Longae being obviously comparatively used alone 100 μMs of resveratrols again or being used alone 80 μMs usually better.And from Fig. 2 B, the content of triglyceride in Cell Line HepG2 born of the same parents, when merging the curcumin of use 50 μMs of resveratrols and 80 μMs, the Rhizoma Curcumae Longae being obviously comparatively used alone 50 μMs of resveratrols or being used alone 80 μMs usually lower.Same, when merging the curcumin of use 100 μMs of resveratrols and 160 μMs, it generates the inhibition of triglyceride for Cell Line HepG2, the Rhizoma Curcumae Longae being obviously comparatively used alone 100 μMs of resveratrols again or being used alone 160 μMs usually better.Therefore, the merging of resveratrol and curcumin uses for cytolipin synthetic reaction, as the generation of triglyceride, has the effect of synergistic inhibition.
Experimental example three: resveratrol and curcumin regulate and control fatty acid synthetase jointly to treat fatty liver
According to the step roughly the same with experimental example two and condition, cultivate HepG2 cell with tissue culture technique, and get containing quantity 1.5 × 10
6the suspension of HepG2 cell, and be seeded in the culture dish of 6 centimeters, cultivate after 16 hours, again original culture fluid is taken out, to be replaced as the cell culture fluid DMEM of hyclone containing 5mM glucose and 10%, add 50 μMs of resveratrols, 40 μMs of curcumins respectively, and the compositions of 50 μMs of resveratrols and 40 μMs of curcumins (percentage by weight being converted into resveratrol and curcumin is 1:1.3), then the incubator of culture dish being inserted 37 DEG C continues cultivation 48 hours.In addition, after original culture fluid is taken out, be replaced as containing 5mM glucose but containing 10% hyclone and not containing the cell culture fluid DMEM of sodium glutamate (glutamate), using as hungry (starvation) matched group; The another cell culture fluid DMEM (containing the hyclone of 10%, but not containing sodium glutamate) be replaced as by culture fluid containing 30mM glucose, as nutrition again (re-nutrition) matched group.
Then, by the broth out in culture dish, rinse after twice with normal saline solution, often dish adds the protein extraction liquid of 100 μ l, scrape with burning-in knife and take out protein extraction liquid, namely obtain the mixture of HepG2 cell and protein extraction liquid simultaneously, and 4 DEG C of reactions after 30 minutes, again by high speed centrifuge with rotating speed 12000rpm centrifugal 30 minutes, and get its supernatant observes the protein concentration of fatty acid synthetase performance with Western blotting method of testing.First, carry out the program of colloid electrophoresis, in this experimental example, by the supernatant of aforementioned each sample, get each hole that its 100 μ g injects the upper strata glue of SDS-PAGE, and bestow voltage 80 volts, continue after 30 minutes, then with voltage 120 volts, continue 120 minutes.Then, colloid after taking-up electrophoresis completes carries out the program of protein transfer printing, be transferred on PVDF (polyvinylidene fluoride, poly (vinylidenefluoride)) film (transfer printing condition is 400mA, 30 minutes) by the protein on colloid.After protein on colloid is transferred to pvdf membrane, the pvdf membrane after transfer printing is placed in the buffer solution with 5% bovine serum albumin and rocks more than 15 minutes.Then, remove the buffer solution of 5% bovine serum albumin, and after cleaning pvdf membrane with the phosphate buffered solution with surfactant Tween-20 (PBST solution), then pvdf membrane is placed in has that anti-fatty acid synthetase (anti-Fas) antibody-solutions (first antibody) is mixed rocks 2 hours.Wherein, anti-Fas antibody is dissolved in PBST solution with the ratio of 1:100.Then, 5 minutes are being cleaned again with TBST solution, and in triplicate after, hyperphoric in the sub-IgG-HRP antibody-solutions (second antibody) of anti-rabbit and mixing rock 1 hour, afterwards with chemiluminescent enhancement reagent dyeing one minute, make egative film photosensitive, in order to detect the content of the fatty acid synthetase (FattyAcidSynthase, FAS) in Cell Line HepG2.
Fig. 3 is the result figure of the fatty acid synthetase content of human hepatocarcinoma cells's strain HepG2 in experimental example three.From the result of Fig. 3, Cell Line HepG2 merges and uses resveratrol and curcumin (percentage by weight is 1:1.3) for the fatty acid synthetase of T suppression cell strain HepG2 compared to being used alone resveratrol or curcumin, has preferably inhibition.Therefore, resveratrol and curcumin jointly can regulate and control and suppress the generation of fatty acid synthetase, and then can be used for treating fatty liver.
The foregoing is only illustrative, but not be restricted.Anyly do not depart from the spirit and scope of the present invention, and to its equivalent modifications of carrying out or change, all should be contained in appending claims.
Claims (15)
1. treat a medical composition for Patients with Fatty Liver, it comprises:
Resveratrol and curcumin; And
The pharmaceutically acceptable carrier of at least one, diluent or excipient,
Wherein, described medical composition bestows described Patients with Fatty Liver, and described curcumin and described resveratrol provide the effect of combined treatment.
2. medical composition as claimed in claim 1, the unit dose of wherein said resveratrol is between 100 to 1000 milligrams, and the unit dose of described curcumin is between 100 to 1000 milligrams.
3. medical composition as claimed in claim 1, the percentage by weight of wherein said resveratrol and described curcumin is between 1:1 to 1:5.
4. medical composition as claimed in claim 1, wherein said pharmaceutically acceptable carrier, diluent or excipient comprise hydrophobic liquid.
5. medical composition as claimed in claim 1, it is soft gelatin capsules.
6. prepare a method for the medical composition for the treatment of Patients with Fatty Liver, comprise following steps:
Weigh the resveratrol of a unit dose and the curcumin of a unit dose;
Described in emulsifying, resveratrol, described curcumin and the pharmaceutically acceptable carrier of at least one, diluent or excipient form the first mixture; And
First mixture described in homogenization also forms the second mixture,
Wherein, described medical composition bestows described Patients with Fatty Liver, and described resveratrol and described curcumin provide the effect of combined treatment.
7. method as claimed in claim 6, comprises further:
Described second mixture is suppressed in soft capsule.
8. method as claimed in claim 6, the percentage by weight of wherein said resveratrol and described curcumin is between 1:1 to 1:5.
9. method as claimed in claim 6, wherein said pharmaceutically acceptable carrier, diluent or excipient comprise hydrophobic liquid.
10. method as claimed in claim 9, wherein said first mixture and described second mixture are formed by multiple two-phase stuff and other stuff, and the particle diameter of the two-phase stuff and other stuff of wherein said second mixture is greater than 100 nanometers.
11. 1 kinds of resveratrols and curcumin are for the manufacture of the purposes of the medical composition for the treatment of Patients with Fatty Liver, and when described medical composition is cast described Patients with Fatty Liver, described resveratrol and described curcumin provide the effect of combined treatment.
12. purposes as claimed in claim 11, wherein said resveratrol and described curcumin synergism are to treat described Patients with Fatty Liver.
13. purposes as claimed in claim 11, wherein said resveratrol and described curcumin regulate and control fatty acid synthetase jointly to treat described Patients with Fatty Liver.
14. purposes as claimed in claim 11, the dosage of wherein said resveratrol is between 100 to 1000 milligrams, and the dosage of described curcumin is between 100 to 1000 milligrams.
15. purposes as claimed in claim 11, the percentage by weight of wherein said resveratrol and described curcumin is between 1:1 to 1:5.
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|---|---|
| TW201600084A (en) | 2016-01-01 |
| TWI542342B (en) | 2016-07-21 |
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