Identification and Quantification of Nicotine Biomarkers in Human Oral Fluid from Individuals Receiving Low-Dose Transdermal Nicotine: A Preliminary Study

Abstract

The objective of this preliminary study was to identify and quantify potential nicotine (NIC) biomarkers in post-exposure oral fluid samples collected from 10 NIC-abstinent human participants administered 7 mg transdermal NIC using liquid chromatography-tandem mass spectrometry (LC-MS-MS). Oral fluid samples were collected prior to NIC patch application and at 0.5 and 0.75 h after patch removal using the Quantisal() oral fluid collection device. The validated LC-MS-MS analyte panel included nicotine-Nbeta-D-glucuronide, cotinine-N-oxide, trans-3-hydroxycotinine, norcotinine, trans-nicotine-1'-N-oxide, cotinine (COT), nornicotine, NIC, anatabine, anabasine, and cotinine-N-beta-D-glucuronide. Analytes and corresponding deuterated internal standards were extracted by solid-phase extraction. NIC and COT concentrations were quantifiable in oral fluid samples collected from 6 of the 10 participants 0.5 h after patch removal and in oral fluid samples collected from 7 of the 10 participants 0.75 h after patch removal. Based on the mean NIC and COT concentrations in oral fluid and plasma for the participants with both quantifiable NIC and COT at the 0.5 and 0.75 h collection times, the oral fluid-plasma ratio was 6.4 for NIC and 3.3 for COT. An ELISA procedure was also validated and successfully applied as a screening tool for these oral fluid samples in conjunction with LC-MS-MS confirmation. An ELISA cut-off concentration of 5.0 ng/mL provided excellent sensitivity for discrimination of COT-positive post-exposure oral fluid samples collected after low-level transdermal NIC exposure and oral fluid samples collected prior to patch application.

The Chemical Components of Electronic Cigarette Cartridges and Refill Fluids: Review of Analytical Methods

“…A number of reports [35][36][37][38][39][40][41] on determination of nicotine and related alkaloids by LC-MS/MS in different matrices (especially in biological fluids, like urine, plasma, saliva) have been published. However, these methods can be also suitable for e-liquids and aerosol from ECs, but their applicability to these matrices has to be verified.…”

Determination of biomarkers of tobacco smoke exposure in oral fluid using solid-phase extraction and gas chromatography–tandem mass spectrometry

“…Concheiro et al [36] have obtained similar values (90-94%), while Miller et al [19] report recovery values of 93-96% for COT. The higher recovery values for this sample are reported by Shakleya and Huestis [6] (98-113%), while Jacob et al [11] have obtained 53-65%.…”

“…Shakleya and Huestis [6] were able to detect 0.1 and 0.3 ng/mL of OH-COT and COT, respectively, with LOQs of 0.5 and 0.2 ng/mL for those compounds, yet using twice the sample volume. Using 0.5 mL of oral fluid, Miller et al [19] report LOQs of 1 ng/mL for all analytes. Therefore, our LLOQs are lower, particularly if one takes into account the lower sample volume used.…”

“…Several methods have been reported for the determination of NIC, COT and related alkaloids in various biological fluids, and those include immunoassays [18][19][20][21], gas chromatography (GC) coupled to either flame ionisation (FID) [22] or mass spectrometric (MS) detection [1,12,[23][24][25][26], high-performance liquid chromatography (LC) coupled to UV detector [27,28] or MS [7,16,[29][30][31][32][33][34][35][36]. Hydrophilic interaction chromatography-tandem mass spectrometry (HILIC-MS/MS) methods have been also reported [33].…”

“…Therefore, OF is the preferred biological specimen for many studies [9]. A few papers have dealt with the determination of NIC and metabolites in OF samples, using GC-MS [1] or LC-MS/MS [19,36]; these highlight the fact that, while adequate results can be obtained using less sensitive and less selective detectors such as UV or FID, MS-based methods represent the state of the art in this respect. In fact, those MS methods allow both the unequivocal identification of the analytes and the detection of the compounds even at very low concentrations, as often occurs in OF.…”