Separation of glucuronides from urine by coupled-column separation using underivatized silica as precolumn

Glucuronides are separated from urine by coupled-column separations (CCSs). The fraction containing the glucuronide(s) is transferred on-line from a silica precolumn to the analytical column (octadecyl derivatized silica), enriched, and separated by ion-pair chromatography. The retention and selectivity on the precolumn are controlled by pH, buffer components, organic modifier, and ion-pair agent. After the injection of filtered urine samples, glucuronides with different chemical properties can be separated. The total analysis of morphine-3-glucuronide and morphine-6-glucuronide is accomplished in less than six minutes, with UV detection at 210 nm.