The Primary Study On The RIP From Caesalpinia Minax Hance: Isolation, Purification And Its Anti-tumor Effect

Ribosome-inactivating protein - RIP is a group of toxic proteins which exist extensively in the nature.It can destroy the structure of ribosome so that it can inhibit the biological synthesis of the intracellular proteins.Researches show that RIP has a good biological activity in terms of anti-virus,anti-tumor,anti-fertility,immune-regulation and pest-resistance,and it has boasted an attractive perspective in the field of medicine and agriculture.This research takes the seeds of Caesalpinia minax Hance grown in Dehong,Yunnan as the material.The Caesalpinia minax Hance protein of a relatively higher concentration is obtained by the extracting and purifying steps of protein precipitation with ammonium sulfate,followed by C-25 cationic exchanging chromatography,and Sephades-G75 gel filtration.Through SDS-PAGE electrophoresis,it was shown that the protein obtained was mono-component,with molecular weight was 29.64KD.It proved that this kind of protein has RNA N-glycosidase activity and anti-tumor activities.Therefore,it can be concluded that we got a new kind of RIP from the seeds of Caesalpinia rninax Hance.For testing RIP's anti-tumor activity,this research studied on mouse's melanoma cells(B16 cells)and cervical cancer cells(Hela cells)for the biological activity in vitro experiment.It was found that the RIP of Caesalpinia minax Hance has a perfect inhibition effect on the proliferation of B16 cells and Hela cells,which presented an obvious quantity-effect relationship and time-effect relationship.(1)The quantity-effect experiment,in which RIP's inhibition activity on the proliferation of cells was tested,showed that with the higher the concentration of RIP,higer inhibition effect on the proliferation of cells would be gotten.Within 48 hours,Rip had an IC₅₀about 145μg/ml on Hela cells,and an IC₅₀about 136/ag/ml on B16 cells.(2)The time-effect experiment,in which RIP's inhibition activity on the proliferation of cells was tested,showed that with the longer the time,the better inhibition effect on the proliferation of cells would be reached.Under the concentration of 150μg/ml,the cells scarcely proliferated.Within 72 hours,the inhibition ratio on Hela cells was 85.61%,while on B16 cells was 88.91%.Comparatively speaking,the ratio on the proliferation inhibition in vitro of B16 cells is a little higher than that of Hela cells.(3)This research had a primary study on the RIP of Caesalpinia minax Hance's influence on B16 cells' adhesion effect.It was found that on B16 cells' adhesion to the components of ECM,RIP had little effect.It had little effect in the error range except that it could reduce B16 cells' adhesion to Collagenâ… (inhibition ratio was about 20%)under the concentration circumstance of 150μg/ml.