ORTHO-TYROSINE AND META-TYROSINE FORMATION FROM PHENYLALANINE IN HUMAN SALIVA AS A MARKER OF HYDROXYL RADICAL GENERATION DURING BETEL QUID CHEWING

The habit of betel quid chewing, common in South-East Asia and the Sou th Pacific islands, is causally associated with an increased risk of o ral cancer. Reactive oxygen species formed from polyphenolic betel qui d ingredients and lime at alkaline pH have been implicated as the agen ts responsible for DNA and tissue damage. To determine whether hydroxy l radical (HO.) is generated in the human oral cavity during chewing o f betel quid, the formation of o- and m-tyrosine from L-phenylalanine was measured. Both o- and m-tyrosine were formed in vitro in the prese nce of extracts of areca nut and/or catechu, transition metal ions suc h as Cu2+ and Fe2+ and lime or sodium carbonate (alkaline pH). Omissio n of any of these ingredients from the reaction mixture significantly reduced the yield of tyrosines. Hydroxyl radical scavengers such as et hanol, D-mannitol and dimethylsulfoxide inhibited the phenylalanine ox idation in a dose-dependent fashion. Five volunteers chewed betel quid consisting of betel leaf, areca nut, catechu and slaked lime (without tobacco). Their saliva, collected after chewing betel quid, contained high concentrations of p-tyrosine, but no appreciable amounts of o- o r m-tyrosine. Saliva samples from the same subjects after chewing bete l quid to which 20 mg phenylalanine had been added contained o- and m- tyrosine at concentrations ranging from 1010 to 3000 nM and from 1110 to 3140 nM respectively. These levels were significantly higher (P < 0 .005) than those of subjects who kept phenylalanine in the oral cavity without betel quid, which ranged from 14 to 70 nM for o-tyrosine and from 10 to 35 nM for m-tyrosine. These studies clearly demonstrate tha t the HO. radical is formed in the human oral cavity during betel quid chewing and is probably implicated in the genetic damage that has bee n observed in oral epithelial cells of chewers.