Generation of indole/skatole during malodor formation in the salivary sediment model system and initial examination of the oral bacteria involved

Abstract

Gram negative anaerobic microbial degradation of proteins, peptides and amino acids in saliva leads to production of oral malodor. Volatile sulfur compounds (VSCs) from cysteine and indole/skatole (I/S) from tryptophan are two major components of breath odor. In this study, salivary mixed bacteria and oral pure cultures were compared for their ability to produce odor from cysteine, tryptophan and salivary supernatant. The VSC malodor inhibitor, zinc, was used to help identify the malodor processes involved. Salivary sediment and salivary supernatant were both obtained from wax-stimulated whole saliva. Gram positive (13) and Gram negative (12) oral pure cultures of common salivary bacteria were each grown in appropriate media and harvested. Incubation mixtures were prepared with salivary sediment at 16.7% (v/v) or bacterial pure cultures at 8.3% (v/v). Cysteine or tryptophan at 6 mM or salivary supernatant at 33.3% (v/v) was added. When included, zinc as ZnCl2 was added at 6 mM. All incubations were carried out for 24 h at 37 °C. Odor generation was assessed organoleptically (0–4 severity scale); VSC was determined with a Halimeter (Interscan Corp., Chatsworth, CA) and I/S was determined using Kovac's colorimetric method. Organoleptic odor levels produced by salivary sediment mixed bacteria with the three substrates tested were comparable, though tryptophan and cysteine generated distinct and characteristic odors. With cysteine and supernatant, rapid and substantial VSC was generated; with tryptophan no such VSC formation was observed. Zinc inhibited VSC generation from cysteine and from salivary supernatant but had no effect on I/S from tryptophan. Surprisingly though, zinc which had no effect on I/S production from free tryptophan, reduced I/S production from supernatant. Of the 12 Gram negative bacteria, 75% produced I/S (mean 3.1 ± 1.5 SEM mg ml−1); most was generated by Porphyromonas intermedia, Porphyromonas gingivalis and Fusobacterium nucleatum. Gram positive pure cultures produced none. Comparable levels of organoleptic odor were produced by Porphyromonas gingivalis from tryptophan and cysteine. Malodor produced orally is complex because multiple microbial types, substrates, metabolic pathways and hence end products are involved. Zinc clearly inhibited I/S production in salivary supernatant, possibly by interfering with the release of tryptophan from peptides/proteins.