Method for the determination of tobacco specific nitrosamine related DNA adducts as their pentafluorobenzoyl ester in pancreatic tissue of sudden death victims

Abstract

Cancer of the pancreas is the 13th most common type of cancer worldwide. Smoking causes a 70% increased risk for this cancer. The tobacco specific nitrosamine 4-(methylnitrosamino)1-(3-pyridyl)-1-butanone (NNK) is a strong carcinogen inducing pancreatic tumours in rodents. 4-(Methylnitrosamino)-1-(3pyridyl)-1-butanol (NNAL), a major NNKmetabolite, proved to be a more potent pancreatic carcinogen than NNK. Metabolic activation ofNNKandNNN lead to the formation of pyridyloxobutyl (POB) and pyridylhydroxybutyl (PHB)DNA adducts releasing 4-hydroxy-1-(3pyridyl)-1-butanone (HPB) and4-hydroxy-1-(3-pyridyl)-1-butanol (diol) after acid hydrolysis, respectively. NNK and NNAL were detected in human pancreatic juice, but metabolic activation leading to DNA adducts as necessary initial step in carcinogenesis has not yet been proven in pancreatic tissue. For analysis of pancreatic tissue, the established method for detection of HPB released after acid hydrolysiswas adopted for the detection of diol-releasing DNA adducts. After hydrolysis the diol was transferred to organic solvent and derivatized with pentafluorobenzoyl chloride (PFBC). The resulting ester was quantified by gas chromatography highresolution mass spectrometry negative chemical ionization. The derivatization product was qualified through specific mass fragmentation, showing the fragments for themolecular ion (m/z 361), the [M-HF]− ion (m/z 341) and the attached pentafluorobenzoate (m/z 167). For quantification in the single ionmode themass traces of PFBCesters fromdiol, HPBand the internal standard [3,3,4,4-D4]HPB, m/z 341, m/z 359, and m/z 363 were recorded. This method allowed for the first time the detection POB and PHB adducts in human pancreatic tissue of smoking and non-smoking sudden death victims.