Artificial saliva

Screen-printed electrodes (SPE) were purchased from Zensor R&D Co. Chitosan (a low molecular weight compound with 75% deacetylation), d(+)-glucose (alpha), ethanol (≥99.8%), lactic acid (LA), potassium chloride (KCl), acetic acid, artificial saliva (used in medical and dental research), GO_x (from Aspergillus niger, 235 U mg⁻¹), and uric acid (UA) were all purchased from Sigma-Aldrich. Potassium ferricyanide (K₃Fe(CN)₆), ascorbic acid (AA), and ferrocenecarboxaldehyde were purchased from Duksan Pure, Honeywell Fluka, and Thermo Fisher Scientific, respectively. All chemicals were of analytical grade and used as received.

Prior to extraction and analysis of saliva samples collected from the IPV patients, the extraction procedure was optimized using artificial saliva (Sigma–Aldrich). The stock solutions of the analytes of interest [the EPA priority PAHs mixture, B(a)P parent compound, and B(a)P metabolites such as the B(a)P 4,5-diol, B(a)P 3,6-dione, and 3(OH) B(a)P] were dissolved in methanol at the concentration of 100 μg/ml and stored at 4 °C. The working solutions prepared from the stock solutions at the level of 0, 10, 25 and 50 μg/ml were used to spike the artificial saliva. To check the percent recovery of analytes of interest, the samples were subjected to the extraction protocol detailed below.