Determination of 5-fluorouracil and 5-fluoro-2'-deoxyuridine-5'-monophosphate in pancreatic cancer cell line and other biological materials using capillary electrophoresis.
Capillary zone electrophoresis (CZE) was used for the rapid determination of 5-fluorouracil (5-FU) and 5-fluoro-2'-deoxyuridine-5'-monophosphate (FdUMP) in pancreatic cancer cell line (PANC-1), culture medium, plasma and pancreatic tissue. The assay is based upon protein precipitation with acetonitrile followed by a 9-min CZE analysis of the supernatant in an uncoated fused-silica capillary employing a borate buffer and on-column absorbance detection at 265 nm. Using 50 microl of sample, 5-FU levels between 4.12 and 132 microg/ml (31.7-1000 microM) were found to provide linear calibration graphs. Intra-day and inter-day RSD values evaluated from peak height ratios (n=5) were <7.6 and <8.8%, respectively. Corresponding RSD values of detection times (n=7) were <1 and <1.5%, respectively. The limits of detection for 5-FU and FdUMP were 1.72 and 5.29 microg/ml, respectively. As application, the accumulation of 5-FU by PANC-1 cells over a 4-h time period was investigated. Having a culture medium concentration of 100 microg/ml, the 5-FU cell content was estimated to become equal to that of the surrounding medium (i.e., 100 microg/ml or 3.61 fmol per cell with a volume of 4.7 pl) within that time period. The sensitivity of the assay was sufficient for the determination of 5-FU in all cell samples. FdUMP, however, could not be detected in these samples. Furthermore, the data obtained in uncoated capillaries are compared to those measured in a fused-silica capillary whose inner surface was coated with linear polyacrylamide (about 10-fold reduction of electroosmosis). The latter capillary format was found to be useless for simultaneous analysis of 5-FU and FdUMP in pancreatic cells but could be potentially useful for analysis of these compounds in plasma.
Procházková
A
A
Department of Clinical Pharmacology, University of Berne, Switzerland.
Liu
S
S
Friess
H
H
Aebi
S
S
Thormann
W
W
D000964:Antimetabolites, Antineoplastic / Q000032:analysis* / Q000097:blood; D002021:Buffers; D003470:Culture Media; D019075:Electrophoresis, Capillary / Q000379:methods*; D005468:Fluorodeoxyuridylate / Q000032:analysis* / Q000097:blood; D005472:Fluorouracil / Q000032:analysis* / Q000097:blood; D006801:Humans; D010190:Pancreatic Neoplasms / Q000737:chemistry* / Q000473:pathology; D012680:Sensitivity and Specificity; D014407:Tumor Cells, Cultured
D000964:Antimetabolites, Antineoplastic; D002021:Buffers; D003470:Culture Media; D005468:Fluorodeoxyuridylate; D005472:Fluorouracil