Simultaneous Determination of 1,4-Benzodiazepines and Tricyclic Antidepressants in Saliva after Sequential SPE Elution by the Same HPLC Conditions

A new method has been described to determine both benzodiazepines (six) and tricyclic antidepressants (four) simultaneously in saliva by HPLC with a UV detector set at 240 nm using cholchicine as the internal standard. A careful specific sequential solid-phase elution was optimized and performed to elute benzodiazepines using a mixture of methanol-acetonitrile (1:1 v/v) followed by the elution of tricyclic antidepressants with methanol. Separation of the compounds was performed on a Kromasil column (250 x 4 mm, 5 mu m) by a gradient eluents consisting of 0.05 M CH3COONH4-acetonitrile-methanol (55:15:30 v/v/v). The results were linear for both benzodiazepines and tricyclic antidepressants up to 20 ng mu L-1 with the correlation coefficients greater than 0.998. The sensitivity limits, LOD and LOQ were 0.08-0.34 ng mu L-1 and 0.28-1.13 ng mu L-1, respectively. The method is simple, fast and reliable with good specificity and sensitivity, will be suitable for use in a clinical setting, where there is a concomitant use of 1,4-benzodiazepines and tricyclic antidepressants.