Development and validation of an HPLC method for the determination of benzodiazepines and tricyclic antidepressants in biological fluids after sequential SPE.
A novel method for the simultaneous determination of six benzodiazepines (BZDs) and four tricyclic antidepressants (TCAs) in biological fluids by HPLC with UV detection at 240 nm has been developed. After a deproteinization step biological fluids were analyzed by direct injection. SPE on Nexus cartridges was also applied. Since two compounds, namely imipramine and diazepam, were coeluting, a sequential SPE protocol has been developed. BZDs were eluted by a mixture of methanol/ACN(1:1), followed by the elution of TCAs with methanol. Separation was performed on a Kromasil C8 column (250 x 64 mm(2) id, 5 microm) using a mobile phase of 0.05 MCH3COONH4/ACN/methanol (initial composition 55:15:30 v/v/v) at a flow rate of 1.0 mL/min delivered by a gradient program within 15 min. Colchicine was used as the internal standard (4 ng/microL). The method was linear for all analytes up to 20 ng/lL, with coefficients of regression between 0.996 and 0.99996. LODs and LOQs were 0.08-1.17 and 0.28-3.91 ng/lL, respectively. Recovery was in the range of 92.8-108.7% for within-day and 91.9-109.9% for between-day assays, with RSD values lower than 10.0% for all matrices.
Uddin
Mohammad Nasir
MN
Department of Chemistry, Laboratory of Analytical Chemistry, Aristotle University of Thessaloniki, Thessaloniki, Greece.
Samanidou
Victoria F
VF
Papadoyannis
Ioannis N
IN
D014151:Anti-Anxiety Agents / Q000032:analysis / Q000737:chemistry / Q000592:standards; D000929:Antidepressive Agents, Tricyclic / Q000032:analysis* / Q000737:chemistry / Q000592:standards; D001569:Benzodiazepines / Q000032:analysis* / Q000737:chemistry / Q000592:standards; D001826:Body Fluids / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation / Q000379:methods* / Q000706:statistics & numerical data; D006801:Humans; D012015:Reference Standards; D012680:Sensitivity and Specificity; D052616:Solid Phase Extraction / Q000379:methods
D014151:Anti-Anxiety Agents; D000929:Antidepressive Agents, Tricyclic; D001569:Benzodiazepines