CN1861098A - Calmophil, pharmaceutical composition and producing method - Google Patents
Calmophil, pharmaceutical composition and producing method Download PDFInfo
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Abstract
The invention relates to the pharmaceutical industry, namely to the technology of manufacturing pharmaceutical compositions from the tissues of the marine organisms and may be used for obtaining the drug intended for recovery of the malfunctioned surfactant system of the lungs. The technical task of the invention consisted in the elaboration of the method for manufacturing pharmaceutical compositions intended for recovery of the malfunctioned surfactant system based on the novel technology for processing the reproductive organs of the marine organisms allowing for the production of the natural surface-active complex comprising the complex of phospholipids with omega-3 polyunsaturated fatty acids, plasmalogenous phospholipids, the short-chain polypeptides, and amino acids (trade mark Calmophil).
Description
Technical field
The invention belongs to the pharmaceuticals industry field.Relate to a kind of technology of extracting medical substance in the marine organisms organ, this material can be used as a kind of medicine and is used for the treatment of diseases such as people's lung surface active systemic-function disorder.
Background technology
Modern study shows that pulmonary surfactant (SAS) is a kind of lipid and proteinic mixture, in alveolar fluid, can alleviate the surface tension of pulmonary by the secretion of II type alveolar epithelial cells.The liquid monolayer of two Palmic acid phosphatidylcholines (DPPC) of about 40%-50% that contained one deck enrichment in the alveolar liquid film, main effect is exactly the surface tension that reduces under the gas/liquid state of lung.DPPC one end contains two saturated acyl chains, and the unsaturated fatty acid ester dense arrangement of the other end is on monolayer, for keeping the stable extremely important of film.Pulmonary surfactant is the mixture of numerous chemical compounds, comprise that phospholipid substance is as two Palmic acid phosphatidylcholines (DPPC), PHOSPHATIDYL ETHANOLAMINE (PEA), Phosphatidylserine (PS), phosphatidyl glycerol ester (PG), sphingomyelins (SM), phosphinositides (PI), phosphatidic acid.The main component of the fatty acid in the liquid phospholipid is a Palmic acid.DPPC and some other phospholipid even the protein that some are special can both reduce the lung surface tension, and wherein DPPC is most important composition.In addition, DPPC, PEA, PG and PI also help the expansion of alveolar.
Human lung surface active system disorders disease is very common.Hyaline membrane disease of newborn (NRDS) claims respiratory distress syndrome (PHMD) again, is the early stage dead major reason of neonate.Primary disease for want of pulmonary surfactant causes that gestational age is 50% in the incidence rate in 26-28 week, and 30-31 week is 20-30%.Adult's adult respiratory distress syndrome (ARDS) is meant that body suffers in the serious disease process in the outer and lung of severe trauma, infection and various lung, cause a kind of be the acute respiratory failure syndrome of feature with respiratory distress of carrying out property and refractory hypoxemia.The case fatality rate of ARDS is 90% before the seventies in 20th century, and be about 60% the eighties, and be 30%～40% the nineties, still maintains this level at present.When ARDS took place, various paathogenic factors caused pulmonary surfactant (SAS) secondary deficiency and deactivation, and the unusual generation and the development that promotes ARDS of pulmonary surfactant.Recover SAS function, block the theoretical basis that this vicious cycle is pulmonary surfactant alternative medicine treatment ARDS.It is disorderly that oxygen is rare or too much in the tissue, bronchial infection can cause that all surfactant generates, correct the feasibility .1984 of lung surface active system disorders referring to Berezovsky V.A., 41,107-111 (Possibilities of correction of thesurfactant system state.Vrachebnoe delo, 1984,41,107-111 (Russia) .)
Experiment has at present confirmed that alternative medicine can reverse pulmonary surfactant disappearance (referring to SA 5032585,1991) in the body.Shown gratifying therapeutic effect from the exogenous surfactant of pulmonis Bovis seu Bubali and amniotic fluid acquisition in treatment child respiratory distress syndrome, and do not found that toxicity is (referring to U 2198670,2003; U 2066197,1996).This points out us, and it is necessary that the means by separation and purification obtain natural surfactant.
Can be used for alleviating dyspnea, delaying the course of disease owing to extracting the self-faced active substance that obtains,, be used for treating " congenital alveolar dysplasia " so there is research worker further to attempt to obtain the class surfactant with the way of chemosynthesis.But can reduce the capillary surfactant of lung fast and must be pulmonary surfactant in The Nomenclature Composition and Structure of Complexes and the body all similar mix lipid, comprising two Palmic acid phosphatidylcholines and some other low melting point lipid, this point is difficult to realize by the way of chemosynthesis, so research just seems important and urgent by the natural pulmonary surfactant medicine that extraction separation obtains to be applied to treat human surfactant disappearance disease.
Obviously, the composition of present DPPC preparation or the mixed phosphatide preparation be made up of DPPC and saccharide, aminoacid, ethanol and fatty acid etc. and content are incomplete same, and (EP 0286011,1988), so be not that all artificial surface active substances have all shown good clinical and pharmacological effect, existing preparation such as synthesising preparation Exosurf (U.S.) (USA 5110866,1992), Alek (moral); Semi-synthetic preparation Survanta (U.S.) and natural medicine Curosurf (U.S.) are referring to Grebennikov V.A. hyaline membrane disease of newborn. books .1995 (Grebennikov V.A.Respiratory distress-syndrome innewborns.M., 1995).Existing patent shows that this mixed phosphatide and neutral fat can be treated various respiratory distress syndrome (ivrds)s, after clinical use, can significantly improve the amount of body inner surface active substance, referring to DE 2900300 (Keitzing L.V.), JP58222022 (Teijin K), EP 110498, and USA 4312860.These medicines contain active component aquation soybean lecithin (phospholipon-90H) (RU 2213746 C1 10.10.2003), and this composition has surface activity, can be as the replacement therapy of Curosurf disease.In addition, short-term is taken ω-lipid can improve that fatty acid in the pulmonary macrophage is formed and the quantity of phospholipid, thereby reaches the purpose of rapid adjustment membrane phospholipid and polyunsaturated fatty acid, referring to Palombo J.D, and Lydon E-E, Chen-P1 etc.The changes of contents of lung fatty acid, macrophage and surface activity phospholipid behind the short-term oral administration n-3 lipid. lipid, 1994,29 (9): 643-649 (Palombo J.D, Lydon E-E, Chen-Pl et al.Fatty acid composition oflung, macrophage and surfactant phospolipids after short-term enteral feeding with n-3 lipids.Lipids.1994,29 (9): 643-649).
Also have one important to studies show that various clinically pulmonary disease or pulmonary surfactant disappearance disease patient often with the disappearance of polyunsaturated fatty acid and some crucial phospholipid, referring to Carrie I..Clement M., de Javel D, etc.Replenish phospholipid for because the behavior of n-3 unsaturated lipids disappearance mice and the reverse effect of biochemical indicator. lipid research magazine .2000,41 (3): 473-480 (Carrie I..Clement M., de Javel D et al.Phospholipid supplementation reversesbehavioural and biochemical alterations induced by n-3 polyunsaturated fatty acids deficiency inmice.J.Lipid res.2000,41 (3): 473-480), point out and exist close ties between the two.There is a kind of theory to think that a large amount of ω-3 is relevant with their low prevalence with omega 6 polyunsaturated fatty acid in Eskimos's food, might be that polyunsaturated fatty acid has changed 20 carbon fatty acids and transforms arachidonic biological approach, the leukotriene that therefore causes causing bronchoconstriction reduces.Animal experiment study shows that eicosapentaenoic acid (EPA) can increase animal lung qi pipe and also can improve thromboxance B-2 level to endotoxic resistance, and referring to Michael L Burr, the application of n-3 fatty acid inspires.Amer.J.Clin.Nutr.2000，71(1S)：397S-8S(Michael L Burr，Lessons from the story of n-3 fatty acids.Amer.J.Clin.Nutr.2000，71(1S)：397S-8S)。Also have data to show that eicosapentaenoic acid (EPA) can promote the leukotriene level in the synthetic of phospholipid and the reduction neutrophilic granulocyte.These have shown that all polyunsaturated fatty acid treatment pulmonary disease has foundation biology.The epidemiology result of the test also shows the protective effect of polyunsaturated fatty acid for childhood asthma and bronochitic.Omega-3 unsaturated fatty acid or the fish oil that contains omega-3 unsaturated fatty acid have been employed the multiple disease of treatment in recent years.The omega-3 unsaturated fatty acid instability, under the situation that does not have antioxidant to exist, in medicine or in vivo oxidized easily rotten, if but contain the phospholipid of certain content in the unsaturated fatty acid then can suppress this Oxidation (Song J.H., Jhoul Y., Miyazawa T., Biosci.Biotechnol.Biochem.1997,01, № 12).Omega-3 unsaturated fatty acid phospholipid in the food can enter into the immobilized artificial membrane at pulmonary alveolar macrophage very soon, and specific ionization fatty acid-eicosapentaenoic acid and docosahexenoic acid are more prone to.In this case, the increase of omega-3 unsaturated fatty acid has reduced the level of ω such as arachidonic acid-6 fatty acid in the immobilized artificial membrane.
Summary of the invention
At the deficiencies in the prior art, the present invention aims to provide a kind of medicine Ka Mofei (Calmofil) that is used for the treatment of lung surface active system disorders disease and preparation method thereof.The new method of use of the present invention is extracted natural surface activity medicine from halobiontic gonad.
A kind of medicine Ka Mofei that is used for the treatment of lung surface active system disorders disease, it is characterized in that a kind of is main component with omega-3 unsaturated fatty acid phospholipid, comprises the compositions of plasmalogen, small peptide and aminoacid etc.The feature of described medicine also is to have surface activity, can be used for the replacement therapy of the disease of lung surface active system disorders.The extract drugs method of invention has been guaranteed in this medication preparation process to be to the enrichment of surfactant and to reject antigenic substance as much as possible.
The present invention is a kind of medicine Ka Mofei that is used for replacement therapy lung surface active system disorders disease, it is characterized by, this medicine Ka Mofei is a kind of compositions that comprises omega-3 unsaturated fatty acid phospholipid, plasmalogen (ethanolamine plasmalogens and choline plasmalogen), cholesterol, fatty acid, small peptide, aminoacid, saccharide, and it consists of: omega-3 unsaturated fatty acid phospholipid accounts for 75-85%; Plasmalogen (ethanolamine plasmalogens and choline plasmalogen) accounts for 0.1-0.5%; T-CHOL accounts for 0.2-2.5%; Short-chain peptide accounts for 0-0.5%; Saccharide accounts for 0-0.5%; Remainder is a free fatty, and hydrocarbon derivative and/or aminoacid all are weight percentage.
Preferably, this medicine obtains from the marine organisms gonad.
Preferably, this medicine can also contain vitamin E and heparin, and concrete content is vitamin E 0.05～0.2%; Heparin 0.1-0.15% all is weight percentage.
The present invention also provides the described preparation method that is used for the medicine Ka Mofei of replacement therapy lung surface active system disorders disease, it is characterized in that: frozen or fresh halobiontic gonad is pulverized, the room temperature solvent extraction, extracting liquid filtering, filtrate decompression concentrates and boils off organic solvent, redissolves in low polar organic solvent, adds cleanser simultaneously and extracts processing, collect organic solvent layer, concentrating under reduced pressure obtains surfactant then; Add normal saline and heparin, last low-pressure refrigeration drying obtains lyophilized powder.
Said medicine also can be prepared in order to following method: frozen halobiontic gonad is pulverized, the room temperature solvent extraction, extracting liquid filtering, filtrate boils off organic solvent through concentrating under reduced pressure, redissolve in low polar organic solvent, add cleanser simultaneously and extract processing, collect organic solvent layer, the vitamin E of adding 0.05～0.2%, concentrating under reduced pressure obtains surfactant then.Add physiological solution and heparin, last low-pressure refrigeration drying obtains lyophilized powder.
Preferably, the raw material that uses is the gonad of marine fishes biology, and described marine fishes biology is squid, cuttlefish, morrhua or other ocean fishes; Described gonad is fish roe and/or roe.
Preferably, the extraction solvent that uses can be the mixed solvent of lower alcohols such as acetone, ethyl acetate, ethanol, methanol and/or these solvents and water, and use amount is a raw material: solvent=1: 5～20 (weight: volume).Preferably, described extraction solvent is an ethanol, and use amount is a raw material: solvent=1: 10 (weight: volume).
Preferably, the low polar organic solvent that uses can be normal hexane, cyclohexane extraction, petroleum ether etc.
Preferably, the cleanser of use can be 5～15% alcoholic solution, can contain 5～15mmol/L CaCl in this solution
2, the preferred use contains 10mmol/L CaCl
2Alcoholic solution.
Preferably, add heparin and normal saline, make surfactant: normal saline: heparin=1: 100: (0.1～0.15), (weight: volume: weight).Disperse 30-60min to obtain little fat body.
Preferably, concentrating under reduced pressure obtains before the surfactant, can add vitamin E, addition 0.05～0.2% (weight ratio).
Preferably, through the dry final products that obtain of low-pressure refrigeration.
Said medicine can be used as the treatment of the particularly neonatal surfactant disappearance of the replacement therapy disease of people's lung surface active system disorders disease.
Analysis result to the surfactant of gained of the present invention shows its composition and the surfactant close (the results are shown in Table 1 and 2) that extracts from animal lung tissue obtaining.
The qualitative, quantitative of table 1SAS composition is table as a result
| The composition title | Marine organisms SAS | Animal lung SAS |
| Content % weight | Content % weight | |
| Phospholipid | 75-85 | 68.6-90.7 |
| T-CHOL | 0.5-2.5 | 0-8.0 |
| Free fatty | 0.1-2.8 | 1.0-27.7 |
| Hydrocarbon derivative | 0.1-0.3 | - |
| The peptide class | 0.1-0.5 | - |
| Protein | - | 1.0-3.0 |
| Phosphatidylcholine | 72-87 | 58.4-85.0 |
| PHOSPHATIDYL ETHANOLAMINE | 7.0-12.8 | 2.6-3.5 |
| Sphingomyelins | 5.7-9.0 | 3.2-12.0 |
| Phosphatidyl glycerol fat | 0.4-8.0 | 0.1-12.0 |
| Phosphatidylinositols | 1.5-2.2 | 1.2-6.6 |
| Phosphatidylserine | 0.6-1.2 | 1.0-6.4 |
| Plasmalogen (ethanolamine plasmalogens and choline plasmalogen) | 0.1-0.5 | - |
| Saccharide | 0.1-0.5 | 0.1-2.0 |
| SAS: normal saline: heparin | 1∶100∶(0.1-0.15) | - |
Table 2 extracts the property list of the surface activity phospholipid that obtains from the squid gonad
| Project | Content of inorganic phosphorus (mg/ml) | Nitrogen content (mg/ml) | Total lipid content (mg/ml) | Amino acid content in the hydrolyzate (mg/ml) |
| Products obtained therefrom | 300-400 | 140-160 | 3.5-5.0 | 5-7 |
Method of the present invention relates to the method that obtains surfactant from seafood fish (squid, morrhua, cuttlefish etc.), and its characteristics are:
1, the leftover bits and pieces that the raw material that uses produces as aquatic products processing;
2, extract highly purified omega-3 unsaturated fatty acid phospholipid with the seafood fish gonad, wherein omega-3 unsaturated fatty acid and two Palmic acid phosphatidylcholines are its main components, also has plasmalogen (ethanolamine plasmalogens and choline plasmalogen) in addition, the arginine of high level, glycine, glutamic acid isoreactivity aminoacid and can improve surface-active short-chain peptide;
Add during 3, with normal hexane extraction and contain 10mM CaCl
2Alcohol-water mixture, can remove impurity such as cholesterol derivative, fatty acid and the carbohydrate content of some;
4, the advance of this law also is embodied in the compositions that products obtained therefrom is the omega-3 unsaturated fatty acid phosphide, mainly contain the two Palmic acid phosphatidylcholines of 35-50%, about 20% docosahexenoic acid, the eicosapentaenoic acid of 10-15%, all phospholipid of resulting composition all contains Palmic acid;
5, comprise adjusting peptide, aminoacid and denier protein in the product; Make the preparation protein content and in the 0.1-0.05% scope, do not find side effect, also not finding has antigenicity in the immunoreation of lung local cells, in the test of rat macrophage group, the macrophage number is at 1h, 1d after the administration, 3d, 7d compares no significant difference with matched group.
The inventive method has advantage and is embodied in:
1, extracts the Bio-surface active material in the albacore leftover bits and pieces (as the squid gonad), have higher productive rate (about 7%), have only 1.5% productive rate and from the mammal lung tissue, extract.
2, contain CaCl by in normal hexane, adding
2Alcohol-water solution extract, played the purpose of purified product, make when medicine is inhaled into, to have better therapeutic, simultaneously also help being adsorbed better by the lung surface, more help expanding the alveolar of depression.
3, obtain little fat body by method with suspended dispersed;
4, in the gained surface-active compounds come, add the low dispersed structure that heparin can improve blood microcirculation and stablize little fat body medicine;
5, the plasmalogen of the some that contains in the phospholipid that obtains from fish is produced in the ocean has greatly strengthened surface activity, simultaneously still the antioxidant in the lipid peroxidation and the antioxidant of low density lipoprotein, LDL;
6, vitamin E is lipid constituent on the one hand, can protect pulmonary surfactant protein to avoid oxidation on the other hand
7, pharmaceutical preparation is easy;
8, the omega-3 unsaturated fatty acid phosphide that obtains has protection cell membrane, immunoregulatory characteristic; can improve the disappearance of unsaturated fatty acid in the tissue (as docosahexenoic acid and eicosapentaenoic acid), regulate and the synthetic relevant arachidonic content of prostaglandin.
The present invention utilizes new technique to extract the surface activity medicine of preparation from the albacore gonad, has following effect:
1, increased the surface activity of lung by surface tension being dropped to normal value (48.5 and 11.7din/cm);
2, pulmonary alveolar macrophage quantity does not have obvious change (6.5-6.9 * 10 with the matched group ratio in process of the test
6/ L).
3, test data shows that medicine Ka Mofei of the present invention can also be further used for the lung surface active system disorders that acute poisoning causes.In intact animal's test, surface activity class material can improve the pulmonary function that causes of poisoning and descend, and reduces the tension force in the foam generative process, recovers the cell effect of pulmonary surfactant and slows down the toxic reaction process.
Short-term uses omega-3 unsaturated fatty acid phosphide preparation can improve the phospholipid cell membrane of lung tissue, pulmonary alveolar macrophage and facies pulmonalis cordis active system fast.It mainly is the phospholipid composition, in the phospholipid composition again based on omega-3 unsaturated fatty acid DPPC, the docosahexenoic acid that also has plasmalogen (ethanolamine plasmalogens and choline plasmalogen) and some in addition, these compositions can reduce surface tension and rock-steady structure effectively, and the surface activity of raising mixture of phospholipids, referring to M.Rudiger, Kolleck, Puiz at al. plasmalogen can reduce surface tension (the .Plasmalogens effectively reduce the surface tension of surfactant-like phospholipid mixtures.Amer.J.Physiol. of class surface activity mixture of phospholipids effectively, 1998, v.274,1, L.143-148).Product of the present invention contains the aminoacid of a certain amount of biologically active such as lysine, arginine, glycine, glutamic acid in addition, and vitamin E and heparin.
Feature of the present invention is that also the technology of using can remove undesirable lipid impurity, and products obtained therefrom can improve the disorder of tissue surface activity system, can be used for pulmonary disease, tissue surface active substance disappearance and neonate " respiratory distress syndrome ".
Essence of the present invention relates to separate from albacore first and obtains the compositions-omega-3 unsaturated fatty acid phosphide of biologically active, and is used for the particularly neonatal surfactant disappearance of the replacement therapy disease of pulmonary disease.
The specific embodiment
Below by specific embodiment the present invention is set forth, but do not limit the scope of the invention in any form.
Embodiment 1 squid gonad (roe and roe) 100 grams, freezing state is pulverized down, adds 10 times of volume of ethanol, stirs in the container and extracts 3 hours, then with extracting liquid filtering, boils off the ethanol in the filtrate under vacuum state.Remainder was by 1: 5 (weight: volume) add normal hexane and 5%10mMCaCl
2Alcoholic solution, 25-30 ℃ of down extraction left standstill, and discards lower floor, adds the vitamin E of 0.05% weight in the normal hexane layer, boils off solvent under the vacuum; Add 0.1% weight heparin, suspended dispersed is 30 minutes in the normal saline, obtains the fat part, lyophilization, promptly.Output 8 grams (yield 8%).
Embodiment 2: morrhua roe 100 grams, with pulverizing, add 20 times of volume of ethanol under the freezing state, and stirred 2 hours in the container, filter extracting solution, boil off ethanol under the vacuum, remainder was in 1: 10 (weight: volume) ratio adding petroleum ether and 10%10mMCaCl
2Alcoholic solution, 25-30 ℃ of down extraction left standstill, and discards lower floor, adds the vitamin E of 0.1% weight in the petroleum ether layer, boils off solvent under the vacuum; The heparin that adds 0.15% weight, suspended dispersed is 30 minutes in the normal saline, obtains the fat part.Lyophilization, promptly.Output 7.5 grams (7.5%).
Embodiment 3: cuttlefish roe 100 restrains, and freezing state is pulverized down, adds 10 times of volume of ethanol, stirs 1 hour in the container, filters extracting solution, boils off ethanol under the vacuum, and remainder was in 1: 5 (weight: volume) ratio adding normal hexane and 15%10mMCaCl
2Alcoholic solution, 25-30 ℃ of down extraction left standstill, and discards lower floor, adds the vitamin E of 0.15% weight in the normal hexane layer, boils off solvent under the vacuum; The heparin that adds 0.15% weight, suspended dispersed is 30 minutes in the normal saline, obtains the fat part, lyophilization, promptly.Output 7 grams (7%).
Embodiment 4: cuttlefish roe 100 grams, pulverize under the freezing state, and add the methanol of 18 times of volumes, stirred 1 hour in the container, filter extracting solution, boil off methanol under the vacuum, remainder was in 1: 5 (weight: volume) ratio adding cyclohexane extraction and 15%10mMCaCl
2Alcoholic solution, 25-30 ℃ of down extraction left standstill, and discards lower floor, suspended dispersed is 30 minutes in the normal saline, obtains the fat part, lyophilization, promptly.Output 7.3 grams (7.3%).
Embodiment 5: cuttlefish roe 100 grams, and freezing state is pulverized down, adds the ethyl acetate of 5 times of volumes, stirred 1 hour in the container, filter extracting solution, boil off ethyl acetate under the vacuum, remainder was in 1: 5 (weight: volume) ratio adding normal hexane and 15%10mMCaCl
2Alcoholic solution, 25-30 ℃ of down extraction left standstill, and discards lower floor, adds the heparin of 0.15% weight, suspended dispersed is 30 minutes in the physiological solution, obtains the fat part, lyophilization, promptly.Output 7.0 grams (7%).
Embodiment 6: cuttlefish fish roe 100 grams, pulverize under the freezing state, and add the acetone of 10 times of volumes, stirred 1 hour in the container, filter extracting solution, boil off acetone under the vacuum, remainder was in 1: 5 (weight: volume) ratio adding petroleum ether and 10%10mMCaCl
2Alcoholic solution, 25-30 ℃ of down extraction left standstill, and discards lower floor, adds the vitamin E of 0.15% weight in the normal hexane layer, boils off solvent under the vacuum; Suspended dispersed is 30 minutes in the normal saline, obtains the fat part, lyophilization, promptly.Output 6.9 grams (6.9%).
Embodiment 7: the drug efficacy study of surfactant medicine of the present invention
The follow procedure operation is pressed in experiment:
● use the poisonous substance modeling, cause lung surface active systemic-function disorders model, will not cause death if do not treat.
● laboratory animal is taked the mode of inhalation.
● observe animal state (dead animal quantity, lung surface active system status parameters)
● under toxic state, use medicine Ka Mofei preparation of the present invention, measure each parameter.
Measure following parameters in the experimentation: the cell effect of animal dead rate, body weight change, lung surface tension, BAM number, lung surface active system, lung foaming function, evaporation time.
1, medicine rat of the present invention sucks effect test.Medicine of the present invention is made concentration and is 1% solution, inhalation.Rat is divided into 7 groups, promptly uses 3 kinds of poisonous substance groups, 3 kinds of poisonous substances difference modelings administration group again, blank group separately, every group of 10 animals.Poisonous substance is 40%meprotane, 35% ammonia, 50% gasoline, and TD is respectively half toxic dose (IDT50) separately, and experimental observation is 14 days after the modeling.The medicine inhalation dose is no more than 0.25ml/min, sucking diameter of aspirin particle is 0.03-3.0 μ m, measure macrophage number (AM) in each treated animal alveolar surface tension (ST) and the alveolar, experimental result shows capillary the increasing (p＜0.001) that medicine of the present invention can significantly reduce the modeling poisonous substance and causes, and makes administration group survival of rats rate increase (table 3).
Table 3 rat sucks the effect test result
| The modeling poisonous substance | IDT50 | IDT50+ medicine of the present invention | ||
| AM(10 6/L) | ST *(mN/m) | AM(10 6/L) | ST(mN/m) | |
| Meprotan | 7.8±1.7 | 54.9±0.7 | 14.7±2.5 ▲ | 42.2±2.7 ** |
| Ammonia | 6.4±0.9 | 51.3±1.6 | 13.8±0.6 ▲ | 42.0±0.9 ** |
| Gasoline | 8.2±1.02 | 49.9±0.5 | 15.8±0.3 ▲ | 47.6±0.8 ** |
| Matched group | 15.2±0.66 | 40.1±0.3 | - | - |
*: compare before the surface tension after the administration and the administration and have significant difference (p＜0.001); ▲: the macrophage number is compared with matched group after the administration does not have significant difference.
2, the toxicity assessment of medicine of the present invention test.Totally 4 groups of laboratory animals, 10 every group.Between rat body weight 195.6 ± 4.5g, the mice body weight is between 23.4 ± 1.7g.The mass concentration of medicine inhalation of the present invention is controlled at 33333.0mg/m
3To 666mg/m
3, animal is in 1h, administration in first day, the 3rd day, the 7th day, and every day 1 time, rat is kept at every turn and sucks 4h, mice 2h.Tested animal dead number after the statistics administration 14 days.Drawing average lethasl concentration is rat 11200.0+606.4mg/m
3With mice 8000+542.0mg/m
3, drawing with international standard (DEST 12.1.007-76) contrast, the present invention sucks medicine and belongs to 3 grades of danger, promptly belongs to the acceptable harmful grade.(table 4).
The toxicity assessment table of table 4 surfactant of the present invention
| Route of administration | Animal | LD50 | Harmful grade |
| Irritate stomach | Mice | 3300±36.6mg/kg | Moderate risk |
| Rat | 4800±127.5mg/kg | - | |
| Percutaneous | Mice | About 2500mg/kg | Low dangerous |
| Rat | About 2500mg/kg | - | |
| Suck | Mice | 14700±433.6mg/kg | Moderate risk |
| Rat | 32500±2333.3mg/m 3 | - | |
| 3 grades of danger | |||
4, medical surfaces tension force of the present invention and surface activity test.Surface tension and surface activity test that " Ka Mofei " carried out with " Survant " and " Exosurf " in contrast in U.S. markon Fo Niya university, the luxuriant and rich with fragrance activity of display card is used for above-mentioned two preparations of surfactant replacement therapy near U.S. at present as a result.In the surface activity test of the premature labor rabbit that lacks surfactant, Ka Mofei and " Survanta ", " Exosurf " all can reduce the respiratory pressure level.In the animals survived test, to compare with matched group " Survanta ", " Exosurf ", Ka Mofei treated animal survival number increases.At CCl
4In inductive toxicity and the response to oxidative stress, in vivo test and in vitro tests all can reduce the toxic reaction that metabolic poison causes, increase the animals survived number, weaken lipid peroxidation, and improve CCl
4Signs of toxicity.(table 6).
The character of table 6 medicine Ka Mofei
| Medicine | Parameter | |||
| Surface tension (din/cm) | Poisoning animals survived number (only) | Active (A0*, LP, SOD**), % | Peak Inspiratory Pressure | |
| Ka Mofei | 11.7-48.5 | 1.5-2.0 | In the body: LP-153, SOD-72 is external: AO-50 | 24.9±1.1 |
| The animal surfactant | 10.3-42.3 | - | - | - |
| CCl 4 | - | - | AO-265 | - |
| SOD | - | - | AO-46 | - |
| “Survanta” | 4.1-37.5 | - | - | 21.2±1.1 |
| “Exosurf” | 21.0-67.5 | - | - | 21.2±1.1 |
| Vitamin E | - | - | AO-53 | - |
| Phospholipid in the lung tissue | - | - | AO-17 | - |
*The AO-antioxidant
*The SOD-superoxide dismutase
5, medicine of the present invention has been done complete pulmonary macrophage function assessment and morphologic test.Behind the rat inhalation, in the immunoreation of lung local cells, do not find any antigen-reactive.The test of macrophage group is in experiment 1h, observation in 1 day, 3 days, 7 days, and macrophage quantity is compared with matched group and do not seen difference after the administration.Antigen protein in the quantitative assay blood, the result shows content between 0.1-0.5%, within the scope of safety.
The invention provides the preparation method of the medicine Ka Mofei that extraction obtains in the marine organisms gonad, contain phospholipid, essential amino acids, vitamin E and heparin in the medicine.Resultant composition has surface activity and anti-oxidation characteristics.Make the dysfunction that medicine can replacement therapy people lung surface active system.
Claims (10)
1, a kind of medicine Ka Mofei, it is characterized by, this medicine is a kind of compositions that comprises omega-3 unsaturated fatty acid phospholipid, plasmalogen (ethanolamine plasmalogens and choline plasmalogen), cholesterol, fatty acid, small peptide, aminoacid, saccharide, and it consists of: omega-3 unsaturated fatty acid phospholipid accounts for 75-85%; Plasmalogen (ethanolamine plasmalogens and choline plasmalogen) accounts for 0.1-0.5%; T-CHOL accounts for 0.2-2.5%; Short-chain peptide accounts for 0-0.5%; Saccharide accounts for 0-0.5%; Remainder is free fatty, hydrocarbon derivative and/or aminoacid, all is weight percentage.
2, the described medicine Ka Mofei of claim 1 is characterized in that this medicine obtains from the marine organisms gonad.
3, medicine Ka Mofei as claimed in claim 1, its feature also is to contain vitamin E and heparin, and concrete content is vitamin E 0.05-0.2%; Heparin 0.1-0.15% all is weight percentage.
4, the preparation method of the described medicine Ka Mofei of claim 1, it is characterized in that: frozen halobiontic gonad is pulverized, the room temperature solvent extraction, extracting liquid filtering, filtrate decompression concentrates and boils off organic solvent, redissolves in low polar organic solvent, adds cleanser simultaneously and extracts processing, collect organic solvent layer, concentrating under reduced pressure obtains surfactant then; Add physiological solution and heparin, last low-pressure refrigeration drying obtains lyophilized powder.
5, the preparation method of medicine Ka Mofei as claimed in claim 4 is characterized in that the raw material that the uses gonad as the marine fishes biology, and described marine fishes biology is squid, cuttlefish, morrhua or other ocean fishes; Described gonad is fish roe and/or roe.
6, the preparation method of medicine Ka Mofei as claimed in claim 4, it is characterized in that the extraction solvent that uses can be the mixed solvent of acetone, ethyl acetate, ethanol, methanol, lower alcohols and/or these solvents and water, use amount is a raw material: solvent=1: 5～20, preferably, described extraction solvent is an ethanol, use amount is a raw material: solvent=1: 10 is w/v.
7, the preparation method of medicine Ka Mofei as claimed in claim 4, it is characterised in that the low polar organic solvent that uses can be normal hexane, cyclohexane extraction, petroleum ether etc.The cleanser that uses can be 5～15% alcoholic solution, can contain 5～15mmol/L CaCl in this solution
2, the preferred use contains 10mmol/L CaCl
2Alcoholic solution.
8, the preparation method of medicine Ka Mofei as claimed in claim 4, it is characterized in that adding heparin and normal saline, by weight: volume: weight makes surfactant: normal saline: heparin=1: 100: 0.1～0.15, disperse 30-60min to obtain little fat body.
9, the preparation method of medicine Ka Mofei as claimed in claim 4 is characterised in that: concentrating under reduced pressure obtains before the surfactant, can add vitamin E, addition 0.05-0.2% (weight ratio); Through the dry final products that obtain of low-pressure refrigeration.
10, the purposes of Ka Mofei as claimed in claim 1 is used for the medicine of replacement therapy lung surface active system disorders disease, the medicine of particularly neonatal surfactant disappearance disease.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| UAA200510611 | 2005-11-09 | ||
| UAA200510611A UA76074C2 (en) | 2005-11-09 | 2005-11-09 | Calmophil, pharmaceutical composition for substitution therapy aimed at recovering malfunctioned surfactant system of lungs |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1861098A true CN1861098A (en) | 2006-11-15 |
| CN100457123C CN100457123C (en) | 2009-02-04 |
Family
ID=37388684
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CNB2006100709297A Active CN100457123C (en) | 2005-11-09 | 2006-03-23 | Calmophil, pharmaceutical composition and producing method |
Country Status (2)
| Country | Link |
|---|---|
| CN (1) | CN100457123C (en) |
| UA (1) | UA76074C2 (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN115192528A (en) * | 2022-07-01 | 2022-10-18 | 国科温州研究院(温州生物材料与工程研究所) | Lung surface active composition containing plasmalogen and preparation method thereof |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| ITMI20021058A1 (en) * | 2002-05-17 | 2003-11-17 | Chiesi Farma Spa | MIXTURES OF SYNTHETIC LIPIDS OPTIMIZED FOR THE PREPARATION OF A RECONSTITUTED SURFACTANT |
-
2005
- 2005-11-09 UA UAA200510611A patent/UA76074C2/en unknown
-
2006
- 2006-03-23 CN CNB2006100709297A patent/CN100457123C/en active Active
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN115192528A (en) * | 2022-07-01 | 2022-10-18 | 国科温州研究院(温州生物材料与工程研究所) | Lung surface active composition containing plasmalogen and preparation method thereof |
| CN115192528B (en) * | 2022-07-01 | 2023-12-01 | 国科温州研究院(温州生物材料与工程研究所) | Lung surface active composition containing plasmalogens and preparation method thereof |
Also Published As
| Publication number | Publication date |
|---|---|
| UA76074C2 (en) | 2006-06-15 |
| CN100457123C (en) | 2009-02-04 |
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