DETERMINATION OF TOTAL AND FREE URINARY THOC AND COT Total and free THOC and COT determinations in urine were performed using the corresponding deuterated compounds as internal standards.
Urine calibrators were prepared by drying known concentrations of THOC and COT methanol solution to dryness and redissolving them in negative urine obtained from a nonsmoker.
Absolute extraction recovery was determined by comparing GC-MS peak areas from extracted urine samples to which known THOC and COT concentrations had been added with peak areas from nonextracted THOC and COT in methanol.
Lower limits of detection were determined as three times the background noise at the appropriate retention times, converting the peak areas to THOC and COT concentrations.
DETERMINATION OF URINARY TOTAL AND FREE THOC AND COT IN TRANSDERMAL SUBJECTS
To determine total urinary THOC and COT (glucuronides and free), the samples were first adjusted to pH 6.
Calibrators for NIC, COT, THOC, and COT-G were prepared at several concentrations in Tris buffer (pH 7.
Stock solutions of NIC, COT, and THOC were prepared in methanol, stored at <0 [degrees]C, and used with appropriate dilutions for chromatographic analyses.
In addition, we added known quantities of COT and THOC to blank urines obtained from nonsmokers who had not been exposed to environmental tobacco smoke.
Further, we observed no cross-reactivity of THOC and COT-G at increasing concentrations with anti-NIC antiserum, nor of COT-G with anti-COT antiserum.
by adjusting urinary COT for the urinary creatinine concentration  or by using a more sensitive and specific method of detection such as isotope dilution liquid chromatography-tandem mass spectrometry  or a monoclonal antibody immunoassay tested for cross-reactivity with THOC.
Indeed, although cross-reactivity of THOC for COT antiserum might be useful from an epidemiological point of view (by indirectly increasing the sensitivity of the COT RIA), it represents an analytical bias, which should be indicated in reports of the characteristics of the assay.