STUDY OF SALIVARY FILMS ON HYDROXYAPATITE 171 � 400 S 350 0 bb 300 'jg' 250 (a) ·a � 200 O 0. 1 0. 2 0. 3 0. 4 Concentration of metal ions(mol/1) 150 r-----------------,,-� (b ) 50..._ ________ ___._ ____ "----� O 0.1 0. 2 0. 3 0.4 Concentration of metal ions(mol/1) Figure 6. The effect of metal ions on the amount of adsorbed albumin (a) and salivary proteins (b). The reaction time is 90 minutes. 0: Ca 6: Mg D: Na ◊: Ca and Mg. electrode covered with the salivary materials was rinsed and transferred into additional freshly distilled water, and then PEI or PSS was injected. The injection of the PEI ions caused a mass increase for the submandibular saliva-coated surface and no mass change for the others. PSS ions, however, behaved oppositely to PEI. Those results suggest that the surface products from parotid and submandibular saliva are cationic and anionic, respectively, and that the surface covered with whole saliva is cationic as a whole, despite a mixture of a number of components. The above-mentioned findings suggest the possibility of alternate adsorption of salivary proteins in the mouth. We then attempted to assemble the pellicle-like structure by alternate adsorption. The results are shown in Figure 8. The mucin-coated gold electrode was immersed alternately in the parotid and the submandibular saliva for 20 minutes, with an intermediate rinse with water. The mass increase demonstrates that the pellicle-like multilayers were constructed from two kinds of salivas. When the electrode was immersed solely in the submandibular, parotid, and whole saliva, however, the mass remained at almost a constant value after the first injection cycle. DISCUSSION The present in situ and in vitro work has confirmed the preferential adsorption of proteins on the HAP surface. These observations have mostly coincided with those of other research-

172 ct! � 00 00 ct! 0 JOURNAL OF COSMETIC SCIENCE llOOng 5 _10 15 0 Time(minutes) (a) salivas (b) PEI 5 0 5 (c) PSS Figure 7. Time courses of mass changes on the mucin-coated QCM electrode, responding to the injection of salivas (5 ml) into distilled water (500 ml), then rinsed with water followed by the addition of PEI ion or PSS ion. The arrows indicate the time of injection. Figure 8. Changes of adsorption amount on mucin-coated QCM electrodes caused by saliva exchanges. 6: submandibular D: parotid e: whole. ers (10, 16,30-34). Several studies have shown that the pellicle formation reaches equi­ librium within 90 minutes (10,17) or 45 minutes (33). The in vitro result in Figure 4 is almost in accordance with those previous results, but as shown in the IR spectra in Figure 2, the in situ results fairly depend on each individual. It seems that the carefully regulated in vitro conditions in the whole mixed saliva are different from the in situ ones, where pure salivas are secreted separately from several different glands. Involvement of the calcium-bridging mechanism in a protein-binding process has been suggested previously by some researchers (31,36). In the present work, it has been shown that Mg2+ ions as well as Ca2 + ions considerably affect the adsorption of the salivary proteins on the HAP surface, but in totally different manners, respectively. In the simple solutions of albumin, which is one of the acidic proteins, these two divalent cations cause an increase in the amount of adsorbed proteins. But this is not the case, interestingly,