| Objective To optimmize the vehicle properties and study the effect of developing new medicine carbomycin (CBMC) which composed of activated carbon nanoparticals (ACNP) absorbing mitomycin C (MMC). Materials and Methods ACNP was prepared by ball milling and deposition in distilled water. Then the morphology and size of ACNP were examined through transmission electron microscopy (TEM) and atomic force microscopy (AFM). Mixed suspension of ACNP and MMC was prepared and adsorption was allowed under ultrasonic condition and the concentration of MMC in the fluid was smeasured. The time at which the arc tangent line of the adsorption curve crosses with the horizontal line was selected as the adsorption equilibrium time. A series of different proportion mixture of ACNP and MMC were prepared and unabsorbed MMC was measured to learn the absorption equation. Then the equation to ensure the best proportion of ACNP for MMC. The release of CBMC was examined in PBS in a homoeothermic swing bed with dialytic equipment. The effect of CBMC to kill four kinds of cells including BGC-823, EC-10, HCT-8 and SMMC-7701 was studied. This experiment involved control group, CBMC groups (0.2, 1, 5, 25, 50, 100, 200, 400, 800μg/ml), MMC groups with the same concentrations as that of CBMC groups. The concentrations of CBMC groups were caculated by the MMC in the system. The effect of CBMC to restrain the S180 solid tumor mice model and to extend the life of H22 liver cancer mice model was studied. BGC-823-Luc cell lines which could stably expressed luciferase was built and then injected into the nude mice to study the effect of CBMC to cure the BGC-823-luc tumor mice model with the IVIS system and then evaluate whether the CBMC have better efficacy than free MMC. The experimental series for abdomen plant tumors included control groups, MMC groups, 0.33 mg/kg, 1 mg/kg and 3 mg/kg CBMC groups. And that for tissue injury studies included 0.02,0.04,0.08,0.10,0.16,0.20,0.40mg dose groups, in which the drugs were subcutaneously injected.Results The ACNP has a similar size, a regular spherical shape and a smooth surface. The average size is 60±20nm; In the concentration range of 0.1-20mg/L, the absorption of MMC (A) had direct proportion with the concentration (C). The equation was A=0.00421+0.06591C(r=0.99987,P<0.0001). Under 25℃, ACNP adsorption for MMC got to stable state 25 minutes later with a the equal temperature equation was C/X= -0.0123+0.00506C(r=0.99916,P<0.0001). 1g ACNP could maximum adsorption with a value of 199.63mg MMC. The optimal ratio for adsoption was MMC: ACNP=1:5. In the release experiment of CBMC, MMC groups released 98% within 8h but CBMC groups only released 70.15% within 28 day. The sustained-release is obvious. To the four kinds of cancer cell lines cultured in vitro, CBMC groups behaved a greater killing ability than MMC groups and the results were statistically significant. Besides, ACNP groups did not have a greater killing effect than control groups. In S180 solid tumor mice model, 1mg/kg MMC group and three different doses of CBMC groups were all behaved a greater killing effect than control groups. With statistical significance (P<0.001) but the ACNP groups have no killing effect. 1mg/kg and 3mg/kg doses of CBMC groups had a more significant effect to control the growth of tumor than MMC groups (P<0.001). In the H22 mice liver cancer model, MMC groups and three CBMC groups all significantly extended the life of mice than control groups (P<0.001). Besides, 1mg/kg and 3mg/kg CBMC groups significantly extended the life of mice more than MMC groups (P<0.01 and P<0.001). In the experiment of viviperception, BGC-823-luc we built highly expressed fluorescence both in vitro and in vivo and the intensity was relate to the cell numbers. By intraperitoneal and subcutaneous injection of BGC-823-luc cells, we found that there were obvious cancer niches through IVIS system after seven days. Seven days later, MMC group and three CBMC groups had significant fewer photons than control group (P<0.05). 3mg/kg CBMC group had significantly fewer photons than MMC group (P<0.05). Fourteen and twenty-one days later, MMC group and three CBMC groups all had more significantly fewer photons than control group (P<0.01). 1mg/kg CBMC group and 3mg/kg CBMC group all had more significantly fewer photons than MMC group (P<0.01). The results of subcutaneous injection models were similar to that of intraperitoneal injection models. In the experiment of local tissue injury study, the results showed that CBMC groups had a significantly smaller tissue injury areas than MMC groups (P<0.01). The injury area of CBMC groups recovered quicker than MMC groups and re-grew hair twenty-eight days later. Conclusions ACNP prepared by ball mill and deposited in the distilled water had a smaller size, a better quality than that sold in the market and it can be produced with a big scale. The best proportion of MMC and ACNP was 1/5 and the best time was twenty-five minutes for adsorption. CBMC had an excellent sustained release ability and abrupt release was not obvious. Containing the same dose of MMC, CBMC groups had a more significant killing effect on BGC-823, EC-10, HCT-8 and SMMC-7701 cell lines than MMC groups. ACNP itself could not kill the cells but it could boost up the effect of MMC. CBMC could significantly prohibit the growth of S180 solid tumor, extend the life of H22 liver cancer, prohibit the growth of BGC-823-luc cancer and prevent the metastasis and recurrence. ACNP absorbing MMC could significantly reduce the toxicity of MMC to the tissue. |
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