Stepwise injection determination of isoniazid in human urine samples coupled with generalized calibration method

• Stepwise injection analysis coupled with generalized calibration method
• Eliminating the matrix effect of sample without its manual predilution
• Fully automated method for isoniazid determination in urine samples
• Automation of spectrophotometric analysis of complex sample matrices

Novel simple and fully automated procedure – the stepwise injection analysis (SWIA) coupled with the generalized calibration method (GCM) – has been suggested for the automation of spectrophotometric analysis of complex sample matrices. The performance of the suggested approach is demonstrated by the SWIA spectrophotometric determination of isoniazid (INH) in human urine samples. The method is based on the determination of complex between ammonium metavanadate (AMV) and INH in the presence of citrate ions which was excreted with the patient's urine in acidic medium with subsequent UV–VIS spectrophotometric detection. The conditions of complex formation have been studied for flow procedure. In order to detect, reduce matrix interferences and exclude manual sample predilution the GCM was applied. The linear ranges were from 1 to 100 mM of INH in human urine samples, and the detection limit, calculated as 3s of a blank test (n = 5), was found to be 0.3 mM. The relative standard deviation of 5 mM of INH varied in the range 1.5–3.5% (n = 10). The sample throughput was 5 h− 1. The proposed method was successfully applied for determination of N-acetyltransferase activity in human hepatocytes. The results of SWIA determination of INH were compared with those obtained by the reference method.