Effects of centrifugation and sonification on whole saliva composition

Objectives: Due to differences in whole saliva (WS) viscosity centrifugation and/or sonification may often be used to obtain homogenous material for laboratory handling. However, little is known about the effects of both on WS. Aim: To test the effect of centrifugation and sonification on WS. Methods: Unstimulated WS was collected from 30 healthy test persons giving rise to 10 pooled samples of 50 ml WS containing saliva from 5 test persons in a random order each. Each pooled sample was divided into 3 aliquots and handled as 1) an untreated WS fraction before centrifugation, 2) a supernatant and 3) a pellet fraction after centrifugation at 16000 g in 10 min. Each of the 3 fractions was further subdivided in A) a fraction that received sonification and B) a fraction that did not. For all six fractions multiple inorganic and protein parameters were determined. For non-sonificated fractions numbers of Streptococcus Mutans, Lactobacilli and Candida were determined by cultivation subsequent with Gram staining of the supernatant and pellet fractions. Results: No effects of centrifugation were obtained for WS concentrations of sodium, potassium, chloride, and phosphorous, as well as amylase activity, while centrifugation halved total calcium in the supernatant (p<0.05) and increased total calcium and protein in the pellet. SDS-PAGE revealed heavily stained proline rich protein bands in the pellet compared to the supernatant. No inorganic or protein effects were obtained by sonification. Significantly higher counts for Lactobacilli and Candida were obtained in the pellet than in the supernatant, although, Gram staining showed micro organisms in both. Gram staining also revealed numerous desquamated epithelial cells that were present in the pellet only. Conclusion: The main effect of centrifugation on WS is the removal of epithelial cells. However, compared to sonification caution must be taken as centrifugation also affects WS inorganic and protein composition.