ABSTRACT
A method was developed for simultaneous determination of Clozapine, Quetiapine and Risperidone in human serum by fully automated online solid phase extraction ( SPE )-high performance liquid chromatography. With Capcell MF Ph-1 column as SPE cartridge and Acclaim C18 as analytical column, high selectivity could be achieved by this method according to the selective complementarity of the two columns. In the experiment, ACN-H2 O was used as the SPE mobile phase with a flow rate of 1 mL/min and ACN-10 mmol/L NH4 Ac was used as the analytical mobile phase with a flow rate of 1 mL/min. Serum samples were injected directly into the SPE column and the biological matrix was washed out with the loading solvent. By rotation of the switching valve, Clozapine, Quetiapine and Risperidone were eluted from the SPE cartridge in the back-flush mode and transferred to the analytical column by the chromatographic mobile phase. The whole time of the online SPE purification and chromatographic separation of the analytes was 18 min. Calibration curve of Clozapine with good linearity ( r=0 . 9996 ) was obtained in the range of 10-1800 μg/L in human serum, and the recoveries at low, medium and high concentration levels were 118. 4%, 105. 0% and 105 . 4%. Calibration curve of Quetiapine with good linearity ( r=0 . 9997 ) was obtained in the range of 3 . 6-640 μg/L in human serum, and the recoveries at low, medium and high concentration levels were 112. 8%, 101 . 1% and 101 . 5%. Calibration curve of Risperidone with good linearity ( r=0 . 9995 ) was obtained in the range of 0. 71-128 μg/L in human serum, and the recoveries at low, medium and high concentration levels were 100. 7%, 97. 2% and 98. 8%. In conclusion, the established automated online SPE-HPLC-UV method demonstrated good performance in terms of linearity, specificity, limits of quantification, and was successfully utilized to quantify Clozapine, Quetiapine and Risperidone in human serum.
ABSTRACT
To develop the methodology to purify and culture expand mesenchymal stem cells (MSC) from human bone marrow and investigate the optimal system for MSC differentiation into chondrocytes mononuclear cells were harvested by gradient centrifugation on Percoll at density of 1 073g/ml and seeded in low glucose DMEM containing 10% fetal calf serum. The purity of cells was evaluated by flow cytometric technique. MSC of 2 passages thereafter were absorbed into a biomaterial of gelatin sponge and induced to differentiate in to chondrocytes for one week under the influence of TGF ? and other inductive agents. The feature of chondrocytes in engineered tissues was identified by toluidine blue staining at various time points. The results showed that human mesenchymal stem cells culture expanded were positive for CD166, CD29, and CD44, but negative for CD34, CD45, and HLA DR. Furthermore, when treated cells absorbed into the biomaterial were implanted subcutaneously into BALB c nude mice, they formed cartilage like tissues after 4 weeks.Our conclusions is that cultured marrow MSC have unique biological features and the capacity to differentiate into chondrocytes in vivo , so they are useful for cartilage engineering by serving as the seed cells.