Assessment of enamel discoloration in vitro following exposure to cigarette smoke and emissions from novel vapor and tobacco heating products.

Purpose: To evaluate in vitro enamel sample discoloration following exposure to a scientific reference cigarette (3R4F) or emissions from next generation tobacco and nicotine products (NGPs) such as electronic cigarettes (EC) and tobacco heating products (THP). Methods: Bovine enamel blocks (6.5 x 6.5 mm) were prepared and pre-incubated with human or artificial saliva, to form a pellicle layer before exposure to either particulate matter (PM) or whole aerosols. PM was prepared by capturing 3R4F cigarette smoke (CS), a commercial THP (THP1.0) or a novel vapor product (NVP)/next generation e-cigarette aerosols on Cambridge filter pads followed by elution with dimethyl sulfoxide (DMSO). Ten enamel samples were exposed to each PM for 14 days. For aerosol exposure, 12 enamel samples were exposed (200 puffs per day, for 5 consecutive days) to 3R4F CS or THP1.0 and NVP aerosols. Control samples were incubated with DMSO (PM study) or phosphate buffered saline (PBS, aerosol study). Individual enamel sample color readings (L*, a*, b*) were measured at baseline and on each exposure day. Mean Delta L*, Delta a*, Delta b* and Delta E values were calculated for each product or control. A one-way ANOVA was used to assess the differences between the products and controls. The Tukey procedure for pairwise comparisons was also used. Results: At all timepoints, 3R4F PM and CS induced enamel discoloration that was statistically significant (P<0.0001) when compared to THP1.0 or NVP. After 14-day PM exposure, mean Delta E values were 29.4 +/- 3.6, 10.5 +/- 2.3, 10.7 +/- 2.6 and 12.6 +/- 2.0 for 3R4F, THP1.0, NVP and DMSO control respectively. After 5 day CS or aerosol exposure, mean Delta E values were 26.2 +/- 3.2, 3.6 +/- 1.9, 3.4 +/- 1.3, 5.3 +/- 0.8 for 3R4F CS, THP1.0, NVP or PBS control, respectively. Both exposure methods demonstrated that THP1.0 and NVP induced minimal staining, mean Delta L*, Delta a*, Delta b* and Delta E values were comparable to DMSO or PBS controls.