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 o f 5-fluorouracil (5-FU) and 5-fluoro-2'-deoxyuridine-5'-monophosphate (FdUM P) in pancreatic cancer cell line (PANC-1), culture medium, plasma and panc reatic tissue. The assay is based upon protein precipitation with acetonitr ile followed by a 9-min CZE analysis of the supernatant in an uncoated fuse d-silica capillary employing a berate buffer and on-column absorbance detec tion at 265 nm. Using 50 mul of sample, 5-FU levels between 4.12 and 132 mu g/ml (31.7-1000 muM) were found to provide linear calibration graphs. Intra -day and inter-clay 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 auld 5.29 mug/ml, respectively. As application, the accumu lation of 5-FU by PANC-1 cells over a 4-h time period was investigated. Hav ing a culture medium concentration of 100 mug/ml, the 5-FU cell content was estimated to become equal to that of the surrounding medium (i.e., 100 mug /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 hi uncoated capillaries sire compared to t hose measured in a fused-silica capillary whose inner surface was coated wi th linear polyacrylamide (about 10-fold reduction of electroosmosis). The l atter capillary format was found to be useless for simultaneous analysis Of 5-FU and FdUMP in pancreatic cells but could be potentially useful for ana lysis of these compounds in plasma. (C) 2001 Elsevier Science B.V. Ail righ ts reserved.