Analysis of Acrylamide and Glycidamide in Dried Blood Spot of Smokers Using Ultra-High-Performance Liquid Chromatography–Tandem Mass Spectrometry

Abstract

Introduction:Acrylamide is a genotoxic substance that can be found in cigarette smoke. Acrylamide is metabolized by the CYP2E1 enzyme in the body to form glycidamides, an epoxide that is reactive to DNA and can form carcinogenic adducts. Therefore, exposure to acrylamide can potentially cause cancer. This study aims to analyze the levels of acrylamide and glycidamide in dried blood spot samples of smokers using propanamide as an internal standard and non-smokers as the control subjects. Methods: Dried blood spot samples were extracted using the protein precipitation method and then analyzed by liquid chromatography-tandem mass spectrometry. Mass detection was performed using positive type electro spray ionization and multiple reaction monitoring type with m/z 72.0>55.02 for acrylamide, 88.1>45.0 for glycidamide, and 74.0>57.1 for propanamide as the internal standard. Results: Acrylamide and glycidamide levels in the dried blood spot sample of smokers ranged between 3.91 -10.25 µg/mL and 1.006-3.58 µg/mL, respectively. Data of the non-smokers on acrylamide and glycidamide levels were 0.75-3.16 µg/mL and 0-0.91 µg/mL. Discussion: The significant value of acrylamide and glycidamide between smokers and non-smokers was p < 0.05, which showed that there is a significant difference between acrylamide and glycidamide concentration in smokers and non-smoker subjects. The results of this study suggest that dried blood spots can be used to determine acrylamide and glycidamide levels in humans. Theoretically, acrylamide and glycidamide concentration should correlate to each other; however in reality, there are other factors (such as CYP2E1 polymorphism, dietary intake, etc) that can cause variation in their respective concentration.

Development and Validation of HPLC-Uv Method for Simultaneous Analysis of Acrylamide and Glycidamide in Volumetric Absorptive Microsampling

“…However, acetonitrile 100% as extracting solvent provided good selectivity, cleaner chromatogram, and less interference appeared at the retention time of analytes and internal standard. In addition, acetonitrile 100% had a benefit as compared to methanol-water (1:1) used in the reference sample preparation (8).…”

“…The dried sample was reconstituted in 100 µl of distilled water and then sonicated for 15 min, vortexed for 30 seconds, and centrifugated for 5 min at 3000 rpm. Finally, 20 µl of aliquot was injected into the HPLC system [8].…”

“…To assess the risk of ACR and GLY exposure in humans, it is important to measure the ACR and GLY levels in the blood. Determination of ACR and GLY has been done in several previous studies, using different biosampling techniques which are venipuncture [7] and dried blood spot (DBS) [8]. Collecting blood by venipuncture is more invasive, requires the phlebotomist, and the plasma sample obtained needs a refrigerator to keep the sample stable.…”