Determination of cotinine in urine using glass capillary gas chromatography and selective detection, with special reference to the biological monitoring of passive smoking

Abstract

A capillary gas chromatographic (GC) method using selected-ion monitoring (SIM) was developed for the analysis of cotinine (C.A.S. No. 486-56-6) in human urine. The method is based on basic extraction of cotinine from 2 ml of urine into dichloromethane. After evaporation of the dichloromethane solution to dryness, 100 μl of toluene were added, prior to GC—mass spectrometric (MS) analysis. Trideuterated cotinine (C.A.S. No. 97664-65-8) was used as the internal standard. More than 1000 automatic chromatographic analyses were made without column degradation. Molecular ions (M) of cotinine and trideuterated cotinine, ( m/ e = 176 and 179), were monitored in the electron impact (EI) mode and m/ e = 177 (M + 1) and m/ e = 180 (M + 1) in the chemical ionization (CI) mode with isobutane. The correlation coefficient with SIM and EI was 0.998 (5–20 ng/ml) and with CI was (0.2–2 ng/ml). For thermionic specific detection the correlation coefficient was 0.998 (10–510 ng/ml). Only capillary columns with an apolar bonded stationary phase film thickness of 1 μm showed sufficient inertness for cotinine analysis at the sub ng/ml level. The relative standard deviations for 5 and 20 ng/ml were 5.2 and 3.5% respectively ( n = 12) using EI. Spiked urine samples from six non-smokers (5 ng/ml) showed a relative standard deviation of 5%. The overall recovery (25 ng/ml) was 100 ± 4%. The minimum detectable concentration, using SIM, was ca. 2 ng/ml in the EI mode and ca. 0.2 ng/ml in the CI mode. The half-time for cotinine was ca. 18 h for both active smokers and non-smokers.