Study of salivary arecoline in areca nut chewers

Abstract

Aims: Arecoline, a predominant alkaloid present in arecanut, has been implicated in the pathogenesis of several oral diseases because of its mutagenic and carcinogenic potential. The response of cultured cells to arecoline is highly dependent on its concentration; arecoline stimulates cultured cells above 0.1 μg/ml and is cytotoxic above 10 μg/ ml. Although this alkaloid seems important for areca nut induced oral diseases and carcinogenesis, little is known of the levels achieved before, during and after chewing. Also, it is prudent to understand its effects in arecanut chewers for a comprehensive understanding of its pathogenesis. Accordingly, the present study quantified the salivary arecoline levels in arecanut chewers.

Materials and methods: The study participants were divided into Study Group A & B and Control Group C; unstimulated whole saliva was collected by spitting method for a period of 5 min. Then, participants in Group A and C chewed 0.5 g of areca nut without any other additives while in Group B were asked to chew 0.5 g of inert rubber base impression material. Stimulated whole saliva from all three groups was collected into graduated tubes during chewing at time intervals of 1, 3, 5, 10, 15, 20 and 25 min. Then, all participants were asked to remove nut particles or inert rubber base material from the mouth, and saliva samples were collected further up to 20 min, changing tubes at 5 min interval. Salivary arecoline was quantitated by HPLC-MS. The tabulation and descriptive statistics of the study were carried out.

Results: In the present study, baseline levels of arecoline were zero in all three groups, whereas mean salivary arecoline levels during chewing were 76.93 ng/ml, 129.83 ng/ml and 64.83 ng/ml and after chewing were 196.17 ng/ml, 321.12 ng/ml and 43.75 ng/ml in Groups A, B and Control respectively, which were significantly higher than reported threshold levels.

Conclusions: The data from this study reveals that a significant amount of arecoline would be trapped in oral cavity, or being re-circulated between blood and saliva might have resulted in surprisingly high levels of arecoline even 10 mins after chewing in both groups after which the levels started declining. The higher levels of salivary arecoline achieved during and after chewing are enough to cause cytotoxic and genotoxic effects on oral tissues over a period of time in chronic chewers. The great differences in salivary arecoline levels achieved during chewing, may contribute to the variable response to areca nut seen in communities where this habit is widespread. Areca nut users have persistent background salivary arecoline levels long after chewing, whereas concentrations achieved are highly variable and consistent with a role in oral pre-malignancy and malignancy..

Keywords: Arecanut; HPLC-MS; OSCC; OSMF; SALIVA; arecoline.

Graph 1

Salivary arecoline level comparison during and after chewing at different time intervals

Graph 2

Mean salivary arecoline level comparison during and after chewing