CROCHETED ETFE-REACTOR FOR ONLINE POSTCOLUMN PHOTODERIVATIZATION OF DICLOFENAC IN HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY

A sensitive and selective bioanalytical method for diclofenac using re versed-phase HPLC and fluorescence detection is described. Diclofenac was detected as its fluorescent derivative after on-line post-column p hotoderivatization. Irradiation with UV light of diclofenac in aqueous solutions leads to the sequential loss of both chlorine substituents and ring closure. The major product, carbazole-1-acetic acid, was dete cted by a fluorescence detector using an excitation wavelength of 286 nm and an emission wavelength of 360 nm. The self-made reactor was a c rocheted ethylene and tetrafluoroethylene (ETFE, named TEFZEL) capilla ry, 20 m in length, wound directly around a 253.7 nm UV lamp. The capi llary was crocheted in order to overcome peak widening. Chromatographi c separation was achieved by using a Regis SPS 100 RP-8 column (5 mu m ; 150 mm x 4.6 mm i.d.) and a LiChrospher 100 RP-18 (5 mu m) guard col umn from E. Merck. The detection limit was 1 ng ml(-1) at an injection volume of 20 mu l. Daily relative standard deviations (RSD) were 5.5% (73 ng diclofenac/ml, n=9), and 5.1% (405 ng diclofenac/ml, n=6), res pectively. Chromatograms of human aqueous humor and human serum contai ning diclofenac, and figures showing the time dependent increase/decre ase of the photoderivatization product, are shown. (C) 1997 Elsevier S cience B.V.