Soil contamination with non-metabolized antibiotics is an emerging environmental concern, especially on agricultural croplands that receive animal manure as fertilizer. In vitro conjugation reactions showed that glutathione s-transferases (GST) induced in maize catalyzed the conjugation of glutathione (GSH) with chlortetracycline (CTC), producing stable conjugates that were structurally characterized using liquid chromatography (LC) with ion-trap mass spectrometry (IT-MS). The chromatogram of the GST-catalyzed reactions displayed two characteristic features of the putative CTC-GSH conjugate: 1) the conjugate eluted sooner than CTC and 2) the conjugate eluted in multiple peaks. The removal of chlorine and the addition of a glutathione moeity result in a highly water soluble CTC-GSH conjugate, thus a much shorter retention time relative to the more hydrophobic CTC is expected. Several fragmentation patterns in the mass spectrum of the CTC-GSH conjugate can be used to verify the identity of the enzyme reaction product. The expected molecular ion [M+H]+ of CTC-GSH conjugate (m/z 751) with chlorine removal was not observed in the positive electrospray ionization. Instead, a base peak of m/z 677, corresponding to the loss of glycine (MW=75 Da) was observed. When m/z 677 was isolated in the ion trap and subjected to further fragmentation, characteristic peaks corresponding to the loss of glutamic acid (m/z=129) and water (m/z 18) were observed in the MS/MS spectrum. Other pharmaceutical-GSH conjugates were characterized by LC/MS and results will be included in the presentation.