Three positional isomers of sulphobromophthalein glutathione monoconjugate (BSP-mGSH) were detected using a paired-ion HPLC method that employs triethylamine phosphate (TEA-H3PO4) as a pairing agent. To confirm that these compounds were glutathione (GSH) conjugates, sulphobromophthalein (BSP) was incubated with a four-fold volume of GSH under alkaline ammonium hydroxide. At least 6 metabolites (3 di-GSH conjugates and 3 isomers of mono-GSH conjugates) were produced under these conditions. The three mono-GSH conjugates were each purified and identified as compounds with a molecular weight of 1,020 according to FAB mass spectrometry results. Positional isomers of BSP-GSH were provisionally distinguished via the addition of the symbols alpha, beta and delta to the end of each abbreviation, to reflect the amount of isomers present. Thus, the isomer present in the largest quantity was termed BSP-mGSH(alpha), the second most abundant isomer was termed BSP-mGSH(beta) and the third was termed BSP-mGSH(delta). Interestingly, a species difference was recognized in that rat cytosol GSH S-transferase (GST) primarily produced BSP-mGSH(alpha), whereas guinea-pig cytosol generated BSP-mGSH(delta), BSP-mGSH(alpha) and BSP-mGSH(beta) equally and rabbit cytosol mainly produced BSP-mGSH(beta).