CN102218057B - Application of 3,4-dichloroisocoumarin in preparation coccidiosis-resistant medicaments - Google Patents
Application of 3,4-dichloroisocoumarin in preparation coccidiosis-resistant medicaments Download PDFInfo
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- CN102218057B CN102218057B CN2011101023666A CN201110102366A CN102218057B CN 102218057 B CN102218057 B CN 102218057B CN 2011101023666 A CN2011101023666 A CN 2011101023666A CN 201110102366 A CN201110102366 A CN 201110102366A CN 102218057 B CN102218057 B CN 102218057B
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- coccidiosis
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- dichloroisocoumarin
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Abstract
The invention discloses application of 3,4-dichloroisocoumarin in preparation coccidiosis-resistant medicaments. By utilizing the inhibiting action of 3,4-dichloroisocoumarin on cell invasion of sporozoit, the 3,4-dichloroisocoumarin is applied to the field of verminosis, and the yield of the filial generation oocysts of coccidium is controlled to prepare coccidiosis-resistant medicaments which are used for preventing and treating coccidiosis.
Description
Technical field
The invention discloses 3, the purposes of 4-dichloro isocoumarin in preparation coccidiosis medicine can suppress the intrusion of Eimeria tenella to host cell, is used for the prevention and the treatment of coccidiosis, belongs to medical science pharmaceutical technology field.
Background technology
Coccidiosis be always raise chickens produce in one of disease, the especially intensification of major control and treatment raise chickens that the large tracts of land of mode is promoted and application after, more become common multiple disease, wherein serious with Eimeria tenella.Coccidiostat can be described as one of medicine that must add in the feedstuff, but in recent years because drug-fast problem reaches problems such as drug residue, makes the research of coccidiostat get into the stage of a bottleneck.Apical organ door parasitic protozoa life cycle is complicated, therefore it is invaded Study on Mechanism and is perplexing researcher always.The cell membrane of Eimeria tenella contains a kind of serine protease, i.e. Rhomboid albumen, and the performance of its enzymatic activity is the key that polypide is invaded host cell.3,4-dichloro isocoumarin is a kind of serpin, and whether it can suppress Eimeria tenella Rhomboid protein active, does not see that report is arranged thereby hinder polypide intrusion host cell.
Summary of the invention
The invention provides 3, the purposes of 4-dichloro isocoumarin in preparation coccidiosis medicine for a kind of new medical application, is used for the coccidiosis prevention and treatment of diseases.
Technical scheme of the present invention is achieved in that
In-vitro separation Testis et penis Gallus domesticus cell carries out primitive cell culture to it.With 3,4-dichloro isocoumarin is diluted to the culture fluid of different final concentrations simultaneously.Extract the Eimeria tenella zygoblast then, each concentration solution be divided into two parts, a part before adding Tissue Culture Plate with the zygoblast effect, insert cell behind the flush away medicine then; Another part and zygoblast together insert cell, and behind the cultivation certain hour, with Hank's liquid flush away medicine, the methanol fixed and stained is counted at microscopically.Observe 3 of variable concentrations, 4-dichloro isocoumarin is to the influence of intrusion, schizont and the egg capsule formation of zygoblast.
Following pharmacological evaluation proves therapeutic effect of the present invention:
External carry out the Testis et penis Gallus domesticus cell former be commissioned to train foster; Extract the Eimeria tenella zygoblast then; With 3; 4-dichloro isocoumarin is mixed with the solution of 100 μ M through DMSO (dimethyl sulfoxide) dissolving, in Tissue Culture Plate, is adding 3; During 4-dichloro isocoumarin, being diluted to final concentration respectively is the culture fluid of 1 μ M, 10 μ M, 20 μ M, 30 μ M, 40 μ M, 50 μ M.
Each concentration solution is divided into two parts, a part before adding Tissue Culture Plate with the zygoblast effect, insert the Testis et penis Gallus domesticus cell then behind the flush away medicine; Another part and zygoblast together insert the Testis et penis Gallus domesticus cell.After cultivating certain hour, count at first generation schizont, second filial generation schizont and the filial generation egg capsule of microscopically antithetical phrase Sporulation, concrete criterion is following: the toxicity of inhibitor pair cell is standard with the cell detachment.
Employing takes into account the principle of first generation schizont quantity, second filial generation schizont quantity and egg capsule quantity, is first index with first generation schizont quantity promptly, judges 3, and 4-dichloro isocoumarin is invaded the influence of cell to zygoblast;
With second filial generation schizont quantity is second index; Judge 3,4-dichloro isocoumarin is right
E. tenellaTherapeutic effect (second filial generation fragmentation stage is the Main Stage that causes chicken dead);
With egg capsule quantity is the 3rd index, judges 3, and 4-dichloro isocoumarin is to the effect (egg capsule of just discharging with the trouble chicken manure is the source of infection of healthy chicken flock) of cause of disease control.
First generation schizont number: zygoblast inoculation back 48 h, observe and calculate the respectively second filial generation schizont number of group, 20 visuals field of every set of calculated under 100 times of inverted microscopes;
Second filial generation schizont number: zygoblast inoculation back 108 h, observe and calculate the respectively second filial generation schizont number of group, 20 visuals field of every set of calculated under 100 times of inverted microscopes.
Egg capsule forms number: zygoblast inoculation back 8 d, same method statistic filial generation egg capsule quantity.
1, first generation schizont quantity
Will
E.tenellaAfter zygoblast is inoculated in Testis et penis Gallus domesticus cell 48h; Through inverted microscope first generation schizont quantity is added up, as shown in Figure 1, along with serpin 3; The increase of 4-dichloro isocoumarin concentration; The first generation schizont quantity of culture group is the trend that reduces gradually altogether, and then amplitude of variation is little for this group of advanced processing, and trend is not obvious.Compare with blank blank group, altogether culture group is 3, and 4-dichloro isocoumarin concentration 20 μ M and 30 μ M time difference heteropoles are (P < 0.01) significantly, significant difference when 40 μ M and 50 μ M (P < 0.05), the pre-treatment group then with blank blank group no significant difference.When 3, certain obscission appearred in cell when 4-dichloro isocoumarin concentration reached 40 μ M and 50 μ M, it is thus clear that 3,4-dichloro isocoumarin has suppressed the growth and the metabolism of cell when 40 μ M, thereby has caused the death of cell.
The result shows, 3 of 20 μ M, and 4-dichloro isocoumarin concentration can effectively suppress the intrusion of external coccidiosis to the Testis et penis Gallus domesticus cell.
2, second filial generation schizont quantity
Behind the zygoblast inoculation Testis et penis Gallus domesticus cell 108h; Through inverted microscope second filial generation schizont quantity is added up; As shown in Figure 2; Altogether the second filial generation schizont quantity of culture group is along with serpin 3, the increase of 4-dichloro isocoumarin concentration and reducing gradually, and this group of advanced processing does not have significant change.Compare with the blank group, altogether culture group is 3, and significant difference during 4-dichloro isocoumarin concentration 20 μ M (P < 0.05), 30 μ M, 40 μ M, 5 μ M time difference heteropoles be (P < 0.01) significantly, and the pre-treatment group is then similar with blank blank group quantity.When cell culture after 4 days, 3,4-dichloro isocoumarin concentration is when 40 μ M and 50 μ M, and the cell detachment phenomenon is obvious, the quantity showed increased of cell death.
The result shows, 3 of 30 μ M, and 4-dichloro isocoumarin concentration can obviously be controlled the second filial generation schizogony of coccidiosis, and chicken coccidiosis is had certain potential therapeutical effect.
3, egg capsule quantity
Behind the zygoblast inoculation Testis et penis Gallus domesticus cell 8d; Through inverted microscope the filial generation egg sac number is added up; As shown in Figure 3; Altogether the filial generation egg sac number of culture group is along with serpin 3, and the increase of 4-dichloro isocoumarin concentration appears and reduces trend gradually, and this group of advanced processing does not have significant change.Compare with the blank group; Altogether culture group is 3, significant difference during 4-dichloro isocoumarin concentration 20 μ M (P < 0.05), difference not significantly (P>0.05) during 30 μ M; 40 μ M, 50 μ M time difference heteropoles are (P < 0.01) significantly, the pre-treatment group then with blank group quantity no significant difference.When cell culture after 8 days, 3,4-dichloro isocoumarin concentration is when 40 μ M and 50 μ M, and large stretch of obscission has appearred in cell, visible finite concentration 3,4-dichloro isocoumarin has the chronic inhibition splitted effect of growing for cell.
The result shows; The Eimeria tenella zygoblast has been accomplished whole life cycle (like Fig. 4) in the Testis et penis Gallus domesticus cell of In vitro culture; 3 of 20 μ M, 4-dichloro isocoumarin concentration just can effectively be controlled coccidiosis filial generation egg capsule quantum of output, for the propagation of cause of disease potential control action is arranged.
Comprehensive above-mentioned experimental result; Through 3; 3 of the external inhibition description of test 20 μ M to coccidiosis zygoblast infected cell of 4-dichloro isocoumarin, 4-dichloro isocoumarin can effectively suppress the outer intrusion to the Testis et penis Gallus domesticus cell of exotospore, can control the quantum of output of coccidiosis filial generation egg capsule simultaneously; Coccidiosis there is certain therapeutical effect, can becomes the main component of coccidiostat.
Good effect of the present invention is:Utilize 3,4-dichloro isocoumarin is invaded the cell inhibiting effect to zygoblast, and with 3,4-dichloro isocoumarin is applied to the parasitic disease field, the quantum of output of control coccidiosis filial generation egg capsule, and the medicine of preparation treatment coccidiosis is used for the coccidiosis prevention and treatment of diseases.
Description of drawings
The number change of Fig. 1, Eimeria tenella first generation schizont;
The number change of Fig. 2, Eimeria tenella second filial generation schizont;
Fig. 3, Eimeria tenella filial generation be spore egg capsule number change not;
The microscopy of Fig. 4, Eimeria tenella In vitro culture is observed picture;
Band * person among the figure, expression is compared significant difference with contrast, and it is extremely remarkable that band * * representes to compare with contrast difference.
The specific embodiment
Through following examples the present invention is described for example further; And do not limit the present invention in any way; Under the prerequisite that does not deviate from technical solution of the present invention, any change or change that those of ordinary skills that the present invention did are realized easily all will fall within the claim scope of the present invention.
With 3,4-dichloro isocoumarin can be made into any pharmaceutical dosage form on the materia medica as effective ingredient, comprising: granule, powder, tablet; Also can pass through dmso solution, be mixed with the solution of 100 μ M, become 20 μ M to get final product solution dilution during use.
Claims (1)
1. 3, the purposes of 4-dichloro isocoumarin in preparation coccidiosis medicine.
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| CN2011101023666A CN102218057B (en) | 2011-04-24 | 2011-04-24 | Application of 3,4-dichloroisocoumarin in preparation coccidiosis-resistant medicaments |
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| CN2011101023666A CN102218057B (en) | 2011-04-24 | 2011-04-24 | Application of 3,4-dichloroisocoumarin in preparation coccidiosis-resistant medicaments |
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| CN102218057B true CN102218057B (en) | 2012-08-08 |
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO1998022098A2 (en) * | 1996-11-20 | 1998-05-28 | Qlt Phototherapeutics, Inc. | Cpp32 inhibitors for regulating apoptosis |
| WO2011050188A1 (en) * | 2009-10-22 | 2011-04-28 | Genentech, Inc. | Anti-hepsin antibodies and methods using same |
Family Cites Families (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20060194769A1 (en) * | 2005-01-25 | 2006-08-31 | University Of Vermont And State Agricultural College | Small molecules that reduce fungal growth |
| CN1785993B (en) * | 2005-12-14 | 2012-03-14 | 南开大学 | Preparation method of non natural isocoumarin and its 3,4-dihydro derivative and use thereof |
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO1998022098A2 (en) * | 1996-11-20 | 1998-05-28 | Qlt Phototherapeutics, Inc. | Cpp32 inhibitors for regulating apoptosis |
| WO2011050188A1 (en) * | 2009-10-22 | 2011-04-28 | Genentech, Inc. | Anti-hepsin antibodies and methods using same |
Non-Patent Citations (2)
| Title |
|---|
| 毕韵梅等.源自微生物次生代谢产物的3 4-二氢异香豆素.《云南化工》.2005 |
| 毕韵梅等.源自微生物次生代谢产物的3,4-二氢异香豆素.《云南化工》.2005,第32卷(第3期),47-50. * |
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