| Objective:Toxicity effect will be shown as materia medica is made into nanoparticles. Thus, it is critical to ensure biological safety of nanoparticles as a part of formulation assessment. In this study, characters of albendazole (ABZ) raw material powder and nanoparticles were compared, and hepatotoxicity of these two formulations was also assessed. Methods:1. The morphology, size, zeta potential and polydispersity index of the two formulations were detected by transmission electron microscopy (TEM), scanning electron microscopy (SEM) and Malvern laser particle scattering;2. ABZ nanoparticles were dispersed in ultrapure water, RMPI1640complete medium, RMPI1640culture medium, phosphate buffer, and0.9%sodium chloride, respectively. Morphology and size were observed by TEM and Malvern laser particle scattering and better media was selected. The suitable size of nanoparticles was selected according to the ultrasound method. And a suitble ultrasound time was selected during5～60min.3. Normal human hepatic cell line, HL-7702, were used for the hepatotoxicity assessment between ABZ raw material and nanoparticles using MTT method. Contents of TP, GSH-px, SOD, MDA in cell culture supernatant were detected by ELISA assay. Results:1. Morphology of ABZ raw material was irregular slices (mostly lamellar irregular).The size distributions is not uniform, while the nanoparticles become irregular multiple-unit, homogeneous and well distributed. The average particle size of ABZ raw material powder was1710±7.1nm, the average potential was5.0±2.20mV, and PDI was0.641±0.199. The average particle size of ABZ nanoparticles was320±2.13nm, the average potential was-18.0±3.25mV, and PDI was0.355±0.127. There were significant differences in characterization between raw powder and nanoparticles.2. Different degree agglomeration of ABZ nanoparticles was occurred in five media mentioned above, and it was less obvious in ultrapure water, phosphate buffer and0.9%sodium chloride. Consideration of the requirements of cell growth, phosphate buffer (pH=7) was a good media for nanoparticles dispersing. The method of combination of ultrasound method and ultrasonic cell disruption method was the optimal dispersing method for nanoparticles and there was no significant increase of the size. Meanwhile there was no significant differences in ultrasound time between30min and45min(P<0.05), and time of30min was selected.3. In cytotoxicity assay, incubations of24h and48h, the RCG of ABZ raw materials was84%and101%, respectively; ABZ nanoparticles was80%and101%, respectively. Cytotoxicity of both kinds of powder was Grade1. Incubation of72h, RCG of ABZ raw materials was between77%and93%, and ABZ nanoparticles was between75%and90%. For ABZ raw materials, the cytotoxicity at concentration of0.190mmol/L in vitro was Grade2, other concentrations were Grade1. For nanoparticles, the cytotoxicity at concentration of0.380mmol/L and0.190mmol/L in vitro were Grade2, other ones were Grade1. The toxic effects of ABZ raw materials and nanoparticles were at the same level.4. Incubation of ABZ for72h, there was no significant difference in total protein content between raw materials and nanoparticles. And there were no significant differences in contents of GSH-px, SOD and MDA in ABZ raw materials groups at different doses(P>0.05). The contents of GSH-px, SOD and MDA in nanoparticles groups at dose of0.380mmol/L and0.190mmol/L, respectively, were changed significantly compared with the sham group (P<0.05). Conclusion:1. ABZ nanoparticles obtained by the optimized micronization method had a better dispersion.2. Phosphate buffer was beneficial for the dispersion of ABZ nanoparticles. ABZ nanoparticles.obtained by the method of combination of ultrasound and ultrasonic cell disruption were suitable for cytotoxicity study.3. There were no cytotoxicity effects on HL-7702cells in both kinds of formulations. and thier cytotoxicity effects were similar.4. Cytotoxicity assay suggested that ABZ nanoparticles at a high dosage could produce a certain oxidative damage on HL-7702cells. |
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