A New Automated Method and Sample Data Flow for Analysis of Volatile Nitrosamines in Human Urine.

James A Hodgson,Tiffany H Seyler,Ernest Mcgahee,Lanqing Wang,Stephen Arnstein

doi:10.4236/ajac.2016.72014

Abstract

Volatile nitrosamines (VNAs) are a group of compounds classified as probable (group 2A) and possible (group 2B) carcinogens in humans. Along with certain foods and contaminated drinking water, VNAs are detected at high levels in tobacco products and in both mainstream and sidestream smoke. Our laboratory monitors six urinary VNAs—N-nitrosodimethylamine (NDMA), N-nitrosomethylethylamine (NMEA), N-nitrosodiethylamine (NDEA), N-nitrosopiperidine (NPIP), N-nitrosopyrrolidine (NPYR), and N-nitrosomorpholine (NMOR)—using isotope dilution GC-MS/MS (QQQ) for large population studies such as the National Health and Nutrition Examination Survey (NHANES). In this paper, we report for the first time a new automated sample preparation method to more efficiently quantitate these VNAs. Automation is done using Hamilton STAR™ and Caliper Staccato™ workstations. This new automated method reduces sample preparation time from 4 hours to 2.5 hours while maintaining precision (inter-run CV < 10%) and accuracy (85% - 111%). More importantly this method increases sample throughput while maintaining a low limit of detection (<10 pg/mL) for all analytes. A streamlined sample data flow was created in parallel to the automated method, in which samples can be tracked from receiving to final LIMs output with minimal human intervention, further minimizing human error in the sample preparation process. This new automated method and the sample data flow are currently applied in bio-monitoring of VNAs in the US non-institutionalized population NHANES 2013-2014 cycle.

Highlights

Volatile nitrosamines (VNAs) are a class of nitrosated secondary and tertiary amines (Figure 1)
The formation of volatile nitrosamines occurs through the nitrosation of secondary and tertiary amines via interaction with nitrite, which itself is a product of nitrate reduction [3]-[5] [7] [8] [19] [20]
NDMA contamination has been detected when DCM comes into contact with any plastic consumable used during the sample preparation process, such as the 48-well plates and pipette tips

Summary

Introduction

Volatile nitrosamines (VNAs) are a class of nitrosated secondary and tertiary amines (Figure 1). VNAs are known carcinogens and teratogens in animals and are classified as group 2A and 2B carcinogens in humans [1][9]. The formation of volatile nitrosamines occurs through the nitrosation of secondary and tertiary amines via interaction with nitrite, which itself is a product of nitrate reduction [3]-[5] [7] [8] [19] [20] For this reason, VNAs can be formed from many items containing nitrates and nitrites, such as cured meats, fish products, cosmetics, certain types of beers, and tobacco products (as part of the curing process and during product assembly), as well as in both mainstream and sidestream tobacco smoke [2] [4] [7] [21]-[23]. Another significant source of VNA exposure can be drinking water: VNAs, NDMA, can form as byproducts during disinfection via chlorination and chloramination [2] [9] [24]-[28]

Methods

Results

Discussion

Conclusion