CN1332687C - Fatty acid synthetic enzyme inhibitor and its production method and application - Google Patents
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Abstract
The present invention discloses a fatty acid synthetic enzyme inhibitor, a production method thereof and an application thereof, which relates to an enzyme inhibitor, a production method thereof and an application thereof. The present invention aims to provide the fatty acid synthetic enzyme inhibitor which can effectively inhibit the activity of a fatty acid synthetic enzyme. The fatty acid synthetic enzyme inhibitor of the present invention is made by extracting keemun black tea with an organic solvent and/or water, wherein the organic solvent is a polar organic solvent with favorable effect and is preferably a water solution of ethanol. The production method of the fatty acid synthetic enzyme inhibitor comprises the following procedures: (1) placing the pulverized keemun black tea in a container and adding the organic solvent or water or a mixture of the organic solvent and water; (2) extracting the solution for 1 to 5 hours at 20 DEG C to 40 DEG C; (3) centrifuging or filtering the solution to collect the supernatant fluid. Medicines and additives with the fatty acid synthetic enzyme inhibitor of the present invention as an active ingredient can be widely used.
Description
Technical field
The present invention relates to inhibitor and the manufacture method and the application of enzyme, particularly relate to a kind of fatty acid synthetase inhibitor and manufacture method thereof and application.
Background technology
Studies show that the body fat of higher mammal is on the one hand by directly taking in, more by self synthesize in the body (Lenninger, A.L., 1982.Principles of Biochemistry, Chapter 13, p333-360 and Chapter21, p593-614; Wakil, S.J.et al., 1983.Ann.Rev.Biochem.52, p537-579).Fatty acid synthetase (FAS) is a most important enzyme in the interior synthctic fat process of biological cell.Tian Weixi etc. discover the poultry fat acid enzyme, the fat level in poultry abdominal cavity and fatty acid synthetase activity have very high positive correlation, and proposition control fatty acid synthetase activity is the theory (Tian Weixi of the effective ways of control animal body lipid level, 1994. body lipid level of animal and the active regulation and control of fatty acid synthetase. the chemistry of life, 14, (1): 184; Tian Weixi etc., body lipid level of 1996. different growing stages laying hens and the active relation of liver fatty acid synthetase. journal of biological chemistry, 12 (2): 234-236; Mei Li et al., 1999.Factors influencing the levels of fattyacid synthase complex activity in fowl.Biochemistry and Molecular BiologyInternational, 47 (1): 63-69).Recently, the Loftus of John Hopkins University etc. prove the result of study of mice fatty acid synthetase, suppress the accumulation that fatty acid synthetase can cause malonyl coenzyme A, and the latter suppresses the expression with the relevant hypothalamic neuropeptide Y that takes food, appetite is descended, and body weight reduces.Loftus etc. point out that fatty acid synthetase can be used as the potential treatment target position of controlling body weight (Loftus.T.M.et al., 2000.Science, 288 (5475): 2379-2381).Above result shows, suppresses the activity of fatty acid synthetase, interior fatty synthesizing not only capable of inhibiting cell, and can also suppress the appetite of animal, and then limit the heat of directly taking in.
In addition, the confidential relation that has that also there are some researches show fatty acid synthetase and cancer, thundering high expressed fatty acid synthetase is all arranged in many cancerous cell, as high level expression and high activity (the Kuhajda FP that just presents fatty acid synthetase in the human breast cancer, Jenner K, Wood FD, et al.Fatty acid synthesis:a potential selective target for antineoplastic therapy.Proc Natl Acad SciUSA, 1994,91:6379-6383).The pernicious kind of apoplexy due to endogenous wind that studies show that these cancers of breast carcinoma, carcinoma of prostate, uterus carcinoma and thyroid carcinoma etc. existed the preferential expression of fatty acid synthetase.The relation that exists between the expression of fatty acid synthetase and the malignant tumor shows that also the synthetic activity of fatty acid plays an important role in tumor is kept its malignant phenotype's process.In fact, except breast and endometrium, fatty acid synthetase all is subjected to the negative adjusting of fat in the diet in most normal structures, and it is opposite, fatty acid synthetase but high expressed normally in cancerous cell, this tissue distribution specificity makes fatty acid synthetase become a noticeable novel targets of treatment cancer.And existing experimental results show that the activity that suppresses fatty acid synthetase has in vivo shown really to the human cancer cell and optionally kill and wound virulence (Francis P.Kuhajda, MD, 2000.Fatty-Acid Synthase and humancancer:new perspectives on its role in tumor bology.Nutrion 16:202-208).
Fatty acid synthetase inhibitor is human cancer cell's a selecting cell toxin, and the relation between malignant tumor and FAS express shows that the expression of fatty acid synthetase and the activity of himself may be all most important to human cancer cell's existence and growth.Experiment in vitro proof FAS inhibitor cerulenin (Cerulenin) is human cancer cell's a selecting cell toxin, the cerulenin of 5g/ml reached ID50 to the growth inhibited of human NIH:OVCAR-3 ovarian cancer in 72 hours, but normal skin fiber blast cell there is not growth inhibited (Pizer ES, Wood FD, Heine HS, et al.Inhibition of fatty acid synthesis delays disease progression in a xenograftmodel of ovarian cancer.Cancer Res 1996; 56:1189).It has resistance of wide spectrum to many human cancer cell lines, comprise mammary gland (MCF-7, ZR-75-1, SKBR3, MDA435) (Kuhajda FP, Jenner K, wood FD, et al.Fatty acid synthesis:a potential selective target forantineoplastic therapy.Proc Natl Acad Sci USA, 1994; 91:6379), ovary (OVCAR-3) (Pizer ES, Wood FD, Heine HS, et al.Inhibition of fatty acid synthesisdelays disease progression in a xenograft model of ovarian cancer.Cancer Res1996; 56:1189), colon (HCT116, RKO), leukocyte (HL60) (Pizer ES, Wood FD, PasternackGR, et al.Fatty acid synthase (FAS): a target for cytotoxic antimetabolitiesin HL60 promyelocytic leukemia cells.Cancer Res1996,56 (4): 745-751), prostate (TSU-prl).E.Pizer etc. find the toxicity and fatty acid synthetic relevant (RashidA of cerulenin to cancerous cell, Pizer ES, Moga M, et al.Elevated expression of fatty acid synthase andfatty acid synthetic activity in colorectal neoplasia.Am J Pathol, 1997,150 (1): 201-208).Inhibition to fatty acid synthetase causes apoptosis to show that the survival of cancerous cell depends on the fatty acid route of synthesis.
In vivo test also obtains similar results, human OVCAR-3 is transplanted in the nude mouse, handle with cerulenin after 24 hours, can improve survival rate and stop it to malignant development (Pizer ES, Wood FD, Heine HS, et al.Inhibition of fatty acid synthesis delays disease progression in a xenograftmodel of ovarian cancer.Cancer Res1996,56 (6): 1189-1193).This processing has the greatest impact to ascites tumor, and malignant development is reduced to 21% from 65% of contrast, and is smaller relatively to the growth metabolism influence of solid tumor.Pizer etc. studies show that, what growth was the fastest in the human body organizes endometrium at proliferative phase chrotoplast expression thereon significant quantities of fat acid enzyme (Pizer ES, Kurman RJ, Pasternack GR, et al.Expressionof fatty acid synthase is closely linked to proliferation and stromaldecidualization in cycling endometrium.Int J Gynecol Pathol 1996,16:45-50).With flow cytometer the HL-60 cell is analyzed demonstration, when suppressed fatty acid when synthetic cell aggregation can not pass through cell cycle in the G1 phase.Find also that in human colon's cancerous cell line DNA is synthetic in 90 minutes behind the inhibition FAS is suppressed, apoptosis begins after 6 hours.These find all to show that the synthetic hypertrophy with tissue of fatty acid exists certain to get in touch.
Another kind of inhibitor---acetyl-CoA carboxylase (ACC) inhibitor, as TOFA (Halvorson DL.Inhibition of fatty acid synthesis in isolated adipocytes by5-(tetradecyloxy)-2-furoic acid.Lipids, 1981,19:851-856), also can suppress the synthetic of fatty acid in the cell, but it can not priming cancer cell apoptosis, and the ACC inhibitor can also make cancerous cell avoid the murder by poisoning of FAS inhibitor.The catalytic reaction of ACC is positioned at the upstream of the catalytic reaction of FAS, and the former makes acetyl-coa carboxylase become malonyl coenzyme A, and the latter then continues S-acetyl-coenzyme-A and malonyl coenzyme A are condensed into fatty acid.
Further two kinds of inhibitor of research are to the toxic difference of cancerous cell, finds that two kinds of inhibitor are handled cells after, different variations take place in the contents level of malonyl coenzyme A in the cell.Utilize HPLC to detect the content of malonyl coenzyme A in the cell of handling with above-mentioned two kinds of enzyme inhibitors, after finding TOFA processing with 20 μ g/ml, the content comparison of malonyl coenzyme A is according to low 60%, and in the cell of handling with the fatty acid synthetase inhibitor C75 of the cerulenin of 10 μ g/ml and another chemosynthesis respectively, the content of the malonyl coenzyme A 930% and 370% (PizerES that raises respectively, Thupari J, Han WF, et al.Malonyl-coenzyme-A is a potential mediatorof cytotoxicity induced by fatty-acid synthase inhibition in human breastcancer cells and xenografts.Cancer Res, 2000,60 (2): 213-218).Obviously, high-caliber malonyl coenzyme A accumulation is a distinguishing feature of only being brought by fatty acid synthetase inhibitor.The fatty acid synthetase inhibitor priming cancer cell death may be to be produced by the malonyl coenzyme A mediation.Malonyl coenzyme A is crucial regulatory molecule in the cellular energy metabolism, and it can suppress the activity of Carnitine palmitoyltransferase I on the mitochondrial outer membrane (CPT I).CPT I can be attached to another kind of cytokine Bcl-2 and go up and make it further to block apoptosis.If suppressed CPT I, Bcl-2 may just not have an effect, and apoptosis takes place cell thereupon.
Because fatty acid synthetase very likely becomes the target spot of obesity and cancer, its inhibitor is very likely developed the medicine of brand-new cancer and obesity, therefore, the inhibitor of development and exploitation fatty acid synthetase will be of great immediate significance and potentiality to be exploited.
Summary of the invention
The purpose of this invention is to provide and a kind ofly can effectively suppress the active fatty acid synthetase inhibitor of fatty acid synthetase.
Fatty acid synthetase inhibitor of the present invention be with organic solvent or/and the mixture of water, the material that from keemun black tea, extracts.
Described organic solvent effect is polar organic solvent preferably, wherein, and preferably ethanol, the most preferably ethanol of 30%-70%, the more preferably ethanol of 40-50%.
Another object of the present invention provides a kind of method of making above-mentioned fatty acid synthetase inhibitor.
Produce the method for above-mentioned fatty acid synthetase inhibitor, may further comprise the steps:
(1) keemun black tea after will pulverizing places container, adds organic solvent or/and the mixture of water;
(2) under 20 ℃ of-40 ℃ of conditions, extracted 1-5 hour;
(3) supernatant is collected in centrifugal or filtration.
In aforementioned production method, the organic solvent of described adding or/and the 5-50 that the weight of the mixture of water can be described keemun black tea weight doubly, wherein preferably 10-20 is doubly; Described organic solvent is a polar organic solvent, wherein, preferred alcohol, most preferably concentration is the ethanol of 30%-70%, more preferably concentration is the ethanol of 40%-50%.
If extraction is being stirred, carried out under the normal temperature condition, then extract and got final product in 2 hours; If centrifugal is to carry out under 8000 rev/mins condition, then carried out 15 minutes.
With fatty acid synthetase inhibitor of the present invention is the medicine that active component is produced, and especially for fat-reducing be used for the treatment of the medicine of cancer, is the main application fields of fatty acid synthetase inhibitor of the present invention.When needing, in these medicines, can also add one or more pharmaceutically acceptable carriers.Described carrier comprises diluent, excipient, filler, binding agent, wetting agent, disintegrating agent, absorption enhancer, surfactant, absorption carrier, lubricant of pharmaceutical field routine etc., can also add flavouring agent, sweeting agent etc. in case of necessity.
Medicine of the present invention can be made various ways such as injection, tablet, powder, granule, capsule, oral liquid, unguentum, cream.The medicine of various dosage forms all can be according to the conventional method preparation of pharmaceutical field.
Fatty acid synthetase inhibitor of the present invention can also be used widely as food additive, household chemicals (particularly cosmetics) additive and products and health products additive agent etc.
Keemun black tea is called for short " keemun ", usually lists one of China's ten famous tea in.The main place of production is Qimen, Anhui Province, to the east of, Guichi, Shitai County, Yixian County etc.The title of tea is gained the name in Qimen County.Keemun black tea leaf bar rope is tight thin elegant long, and the soup color is red gorgeous bright, and particularly its fragrance exactly likes fruital, is with Cymbidium ensifolium (L.) Sw. perfume (or spice) again, and is clear bright and lasting.Both can steep drink separately, and also can add milk and transfer drink.
The present invention dexterously with the broken end of keemun black tea with organic solvent or/and the mixture of water, particularly the mixture of second alcohol and water extracts, and creatively discloses, the material that is extracted is effective fatty acid synthetase inhibitor.Because keemun black tea does not have toxicity as existing more than the 100 year history of beverage, thereby provide wide prospect for the further development and use of this fatty acid synthetase inhibitor.Experiment shows: fatty acid synthetase inhibitor can effectively suppress the activity of fatty acid synthetase, and the interior synthctic fat of biological cell what the active power of fatty acid synthetase directly has influence on.That is to say, the activity that has suppressed fatty acid synthetase, the process of synthctic fat can weaken in the biological cell, fat content promptly can be controlled, even reduce, the more important thing is that fatty acid synthetase inhibitor can also be by biting the appetite of the expression of signal peptide Y with the control animal before suppressing in the brain, therefore fatty acid synthetase inhibitor of the present invention can be used as the active component of slimming medicine, produces the slimming medicine of various dosage forms such as oral liquid, electuary, capsule, tablet, injection.Therefore fatty acid synthetase inhibitor has the selective killing effect to cancerous cell, is the medicine of treatment cancer of the various dosage forms of active component with fatty acid synthetase inhibitor of the present invention, owing to be free from side effects, will obtain using very widely.Fatty acid synthetase inhibitor of the present invention can also be as the additive of health product, food, such as, can be used as the additive of phitosite, make the people of happiness food phitosite both satisfy gourmet's luck, do not worry increasing fat again.Fatty acid synthetase inhibitor of the present invention can also as making slimming cream, be fixed a point to smear to the position of needs fat-reducing as the additive of beauty treatment household chemicals.Simultaneously, because the very big wide material sources of keemun black tea output, technology maturation is for the suitability for industrialized production of fatty acid synthetase inhibitor provides assurance.
The invention will be further described below in conjunction with specific embodiment.
The specific embodiment
In following embodiment, used fatty acid synthetase is with conventional method (W.X.Tian et al., 1985.J.Biol.Chem., 260 (20): 11375-11387; Tian Weixi etc., body lipid level of 1996. different growing stages laying hens and the active relation of liver fatty acid synthetase. journal of biological chemistry, 12 (2): 234-236) in fresh duck liver, separate to purify, and detect through sds polyacrylamide gel electrophoresis and to be single band.
Active S-acetyl-coenzyme-A, malonyl coenzyme A, the NADPH of adopting of fatty acid synthetase is standard measuring method for activity mensuration (W.X.Tian et al., 1985.J.Biol.Chem., 260 (20): 11375-11387 of substrate; Tian Weixi etc., body lipid level of 1996. different growing stages laying hens and the active relation of liver fatty acid synthetase. journal of biological chemistry, 12 (2): 234-236).
The degree of fatty acid synthetase (FAS) activity inhibited represents that with I% (inhibition degree) its computational methods are: add the experimental group that is of fatty acid synthetase inhibitor extracting solution in the substrate, recording the active numerical value of its fatty acid synthetase is A.With what only add corresponding extraction solvent in the substrate is contrast, measures its fatty acid synthetase activity value, this value A
0Expression, and to set it be 100%;
RA% (residual activity)=A/A
0* 100%
I% (inhibition degree)=100%-RA%
Embodiment 1: extract fatty acid synthetase inhibitor with ethanol from keemun black tea
(1) gets keemun black tea, pulverize and be placed in the container 50% ethanol of 10 times of weight of adding, sealing after fully stirring;
(2) under room temperature, stirring condition, extracted 2 hours;
(3) with above-mentioned extract under 8000 rev/mins condition centrifugal 15 minutes, get supernatant, be the extracting solution of fatty acids synthetase inhibitors, use in order to following experiment.
Can use the method identical, use ethanol (as 10%, 30%, 50%, 70%, 90%, 100%), the water of variable concentrations gradient to extract fatty acid synthetase inhibitor with this embodiment.If under non-stirring condition, extract, then should stir half an hour once at least, extracted 3-4 hour.In addition, can also wait with butanols, the acetone of low pole to extract, but experimental results show that dissolubility is not high, insoluble in non-polar solven.Method with this embodiment extracts fatty acid synthetase inhibitor in keemun black tea, under the stirring condition, as long as the extraction time was not less than 1 hour, temperature all can extract effective ingredient between 20-100 ℃.In actual production, the general pressure filter that adopts in extraction back is collected supernatant.
1 gram keemun black tea after extracting with 10ml 50% alcoholic solution removes the volatilization of the ethanol in the extract, dries 2 hours for 120 ℃, can obtain the dry (standard deviation (SD) is 8.8mg) of 221mg.
Embodiment 2: the fatty acid synthetase inhibitor that adds varying number in the identical experimental system solution is to the active influence of FAS (surveying the enzyme addition of living is every milliliter of experimental system solution 2.72 micrograms).
Listed the treatment dosage and the experimental result that in identical experimental system solution, add the fatty acid synthetase inhibitor of varying number in the table 1:
| The microlitre number that adds embodiment 1 extracting solution in every milliliter of system's solution | 0.05 | 0.10 | 0.15 | 0.20 | 0.25 |
| Be converted to the milligram number that adds keemun black tea | 0.01 | 0.02 | 0.03 | 0.04 | 0.05 |
| Be converted to the milligram number that every milligram of enzyme adds keemun black tea | 3.67 | 7.34 | 11.0 | 14.7 | 18.3 |
| The repressed degree I% of FAS enzymatic activity | 35 | 60 | 80 | 84 | 92 |
By above experimental result as can be seen, along with adding the increase of extracting liquid measure, the FAS enzymatic activity is suppressed degree I% and raises, and when adding the extracting solution of 0.15 microlitre in every milliliter of experimental system solution, I% is up to 80%.
Embodiment 3: extractant is to the active influence of fatty acid synthetase
Get the experimental system solution identical with embodiment 2, the ethanol that in every ml soln, adds 0.10,0.2,0.3,0.4,0.5 microlitre 50% respectively, the enzyme amount that adds is also identical with example 2, the repressed degree I% of detection computations FAS enzymatic activity, the result is that I% is 0, show that the extractant ethanol of institute's dosage can not suppress the activity of fatty acid synthetase, suppressing fatty acid synthetase active is the material that extracts from keemun black tea.
Embodiment 4: the 503nhibiting concentration of fatty acid synthetase inhibitor
Extracting solution with embodiment 1 gained experimentizes, the result shows, reach I%=50%, need to add the extracting solution of 0.081 microlitre in every ml soln, being converted to a milligram number that adds keemun black tea is 0.016, and being converted to the milligram number that the activity that suppresses every milligram of enzyme need add keemun black tea is 5.9, and 503nhibiting concentration is very low, illustrate that fatty acid synthetase inhibitor of the present invention has had strong inhibitory effects to the activity of fatty acid synthetase, and required effective dose is very low.
Embodiment 5: different extractants are to the active influence of the fatty acid synthetase inhibitor that extracts
Use aqueous solvent respectively, 50% ethanol, ethanol, acetone, butanols, ether, petroleum ether extracts fatty acid synthetase inhibitor according to the method for example 1, respectively gets 1 microlitre enzyme to same amount in identical experimental system solution then and suppresses active detection, and the result is as shown in table 2:
Table 2:
| Solvent | Water | 50% ethanol | Ethanol | Acetone | Butanols | Ether | Petroleum ether |
| I％ | 24 | 97 | 3 | 6 | 5 | 6 | 5 |
From above experimental result as can be seen, fatty acid synthetase inhibitor of the present invention can be water-soluble, and in the higher solvent of water and alcoholic acid mixture isopolarity, and dissolubility is not high in low solvent of polarity and non-polar solven.
Embodiment 6: the water of different proportion and alcoholic acid mixture are to the active influence of extraction fatty acid synthetase inhibitor
The difference water, 10%, 30%, to be solvent extract fatty acid synthetase inhibitor according to the method for embodiment 1 with 50%, 70%, 90% ethanol, detect them then to the active influence of FAS. (containing enzyme 10 micrograms in every milliliter of experimental system solution), the result is as shown in table 3:
Table 3:
| Extractant | Water | 10％ | 30％ | 50％ | 70％ | 90％ | 100％ |
| The microlitre number that adds extracting solution in every milliliter of system's solution | 0.5 | 0.5 | 0.5 | 0.5 | 0.5 | 0.5 | 0.5 |
| Be converted to the milligram number that adds the Chinese medicine keemun black tea | 0.05 | 0.05 | 0.05 | 0.05 | 0.05 | 0.05 | 0.05 |
| Be converted to the milligram number that every milligram of enzyme adds the Chinese medicine keemun black tea | 5 | 5 | 5 | 5 | 5 | 5 | 5 |
| The repressed degree I% of FAS enzymatic activity | 24.4 | 21.4 | 78.6 | 97.0 | 36.0 | 4.17 | 2.98 |
As can be seen from the above results, concentration of ethanol has remarkable influence to the content of active ingredient in the extract, comparative optimization be that concentration is the ethanol of 30%-70%, most preferably concentration is the ethanol of 40-50%.
Embodiment 7: extract is to the judgement of the irreversible inhibition of fatty acid synthetase
The extract of a certain amount of embodiment 1 gained is mixed with enzyme, and the residual activity that makes enzyme was placed 80 minutes near 0%, crossed the centrifugal post of Sephadex G25 then.If inhibitor wherein and enzyme are reversible bonded words, behind the post, micromolecule can be stayed on the post excessively, and the activity of enzyme will be recovered, if irreversible fixation will pass through gel column together with enzyme, the activity of enzyme can not be recovered.For preventing that post itself from causing inactivation to produce irreversible false picture to enzyme, do a contrast with the enzyme that does not add extract simultaneously.Experimental result finds that activity still remained 70% after the enzyme of matched group was crossed post, and did not recover with the activity of the blended enzyme of extract, illustrated that extract has irreversible inhibition to fatty acid synthetase except that fatty acid synthetase being had very strong fast combination.
Embodiment 8, extract are to the antiobesity action of rat
(1) handle extract, the influence of removing organic solvent: the extracting solution among the embodiment 1 is dry at normal temperatures, and reuse is removed precipitation with the water dissolution of dosage;
(2) 30 standard test rats are divided into 3 groups at random, 10 one group, are respectively negative control group, positive controls, administration group.2ml water is fed in every filling every day of negative control group rat; 2ml 0.1mg/ml fenfluramine is fed in every filling every day of positive controls rat, 2ml step (1) treating water liquid is fed in every filling every day of administration group rat.
(3) measure body weight, the food ration of every rat every other day, Therapy lasted 20 days is divided into and experiment mid-term, full phase calculates body weight, the food ration meansigma methods of 3 groups of rats and carry out the t check analysis.Experiment disconnected cervical vertebra last day is put to death rat and is got determination of serum blood glucose by standard method, triglyceride, cholesterol and low density lipoprotein, LDL take out the liver of every group of rat and distinguish homogenate, its supernatant fatty acid synthetase activity is measured in centrifugal back, and the result is respectively shown in table 4, table 5, table 6:
Table 4, rat body weight change list
| 10 days | 20 days | |||||
| Negative control group | Positive controls | The administration group | Negative control group | Positive controls | The administration group | |
| Meansigma methods (g) | 2.65 | -13.8 | -18.7 | 7.55 | -8.05 | -15.8 |
| Standard deviation (g) | 9.9 | 13 | 12. | 13 | 24 | 15 |
| T verification P value | 0.00289 | 0.000455 | 0.0433 | 0.00181 | ||
Table 5, rat food ration change list
| 10 days | 20 days | |||||
| Negative control group | Positive controls | The administration group | Negative control group | Positive controls | The administration group | |
| Meansigma methods (g) | 22.5 | 19.9 | 19.4 | 23.6 | 21.6 | 20.4 |
| Standard deviation (g) | 1.9 | 1.9 | 1.4 | 1.8 | 1.5 | 1.9 |
| T verification P value | 0.0036 | 0.0009 | 0.0164 | 0.00121 | ||
Table 6, rat blood serum regulating liver-QI fatty acid synthetase data
| The mensuration project | Negative control group | Positive controls | The administration group |
| Blood glucose | 5.44±0.78 | 4.65±0.43 | 5.13±0.21 |
| Cholesterol | 1.27±0.13 | 1.63±0.33 | 1.26±0.40 |
| Triglyceride | 0.29±0.05 | 0.32±0.05 | 0.22 ± 0.05 (t verification P value=0.039) |
| Low density lipoprotein, LDL | 0.53±0.06 | 0.71±0.2 | 0.56±0.2 |
| The vigor of relative negative control group liver fatty acid synzyme | 1 | 0.73±0.06 | 0.59±0.1 |
The t check results shows that fatty acid synthetase inhibitor of the present invention loses weight and the effect of appetite-suppressing by oral can playing than positive control is more significant, find this inhibitor content of triglyceride reducing significantly by every index of measuring in the serum, and can significantly reduce the activity of fatty acid synthetase in the liver.Under test dose, administration group rat does not show any untoward reaction, and is movable normal, do not observe overt toxicity.
Embodiment 9, extract are to the effect of cancerous cell
(1) handle extract, the influence of removing organic solvent: the extracting solution among the embodiment 1 is dry at normal temperatures, and reuse is removed precipitation with the water dissolution of dosage;
(2) under the standard culture condition, human breast cancer cell MCF-7 and fibroblast are cultivated in RPMI culture medium 1640 (containing 10%FBS), cell is divided in 96 orifice plates, 8000 cells in every hole, overnight incubation, add the extracting solution (from 5 μ l-50 μ l) after the not commensurability processing then respectively, use solvent (water) to do negative control simultaneously, do positive control with 5 fluorouracil and cisplatin respectively with dosage.After adding sample, cell is incubated 72 hours under 37 ℃ of degree, with MTT dyeing, 37 ℃ of degree are incubated 4 hours down, add the DMSO cessation reaction, measure absorption value under 570nm, obtain its 503nhibiting concentration (IC50) by linear regression then.
The result shows that the growth that adds the MCF-7 cell of inhibitor is subjected to obvious inhibition; The matched group that adds solvent (water) does not have the growth inhibited phenomenon.The above results shows that fatty acid synthetase inhibitor of the present invention is to the certain selective lethal effect of cancerous cell, and IC50 is 0.698 ± 0.624mg Folium Camelliae sinensis/ml.The IC50 of positive control 5 fluorouracil, cisplatin is respectively 9.43 ± 4.88 * 10
-3Mg/ml and 4.295 ± 1.415 * 10
-3Mg/ml.
Claims (4)
1, the application of keemun black tea extract in the active medicine of preparation inhibition fatty acid synthetase, described keemun black tea extract is with polar organic solvent or/and the material that water extracts from keemun black tea.
2, application according to claim 1 is characterized in that: described polar organic solvent is an ethanol water.
3, application according to claim 2 is characterized in that: described concentration of ethanol is 30%-70%.
4, application according to claim 3 is characterized in that: described concentration of ethanol is 40%-50%.
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| CN1079106A (en) * | 1992-05-29 | 1993-12-08 | 宇斌 | The production technology of caffein |
| CN1161321A (en) * | 1996-09-28 | 1997-10-08 | 浙江大学 | Method for extracting tea polyphenol and by-products from tea leaves |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108524678A (en) * | 2018-07-13 | 2018-09-14 | 河南中医药大学 | A kind of tea seed cake is used as inhibitor for squalene synthetic enzyme and the purposes for lipid-loweringing |
Also Published As
| Publication number | Publication date |
|---|---|
| CN1475249A (en) | 2004-02-18 |
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