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Ca Pre-rinse Greatly Increases Plaque and Plaque Fluid FAmerican Dental Association Foundation, Paffenbarger Research Center, 100 Bureau Drive Stop 8546, National Institute of Standards and Technology, Gaithersburg, MD 20899-8546, USA Correspondence: * corresponding author, jvogel{at}nist.gov
Previous studies demonstrated that a Ca pre-treatment greatly increases salivary F from a subsequent NaF rinse. This study examines if these increases are found in plaque and plaque fluid F. Thirteen individuals accumulated plaque before rinsing with: (1) 12 mmol/L NaF (228 µg/g F), (2) 150 mmol/L Ca rinse, or (3) the Ca rinse followed by the F rinse. One hr later, plaque samples were collected, the plaque fluid was recovered, and the plaque residues were extracted 5 times with pH 6.8 or pH 4.8 buffers, and then by acid. The F in each extract after the Ca rinse/F rinse greatly exceeded the corresponding F from the NaF rinse. Consequently, the Ca rinse/F rinse increased the total plaque F and the plaque fluid F by 12x and 5x, compared with the NaF rinse alone. These and the previous salivary results suggest that a Ca pre-treatment may increase the cariostatic effects of topical F agents.
Key Words: fluoride • rinse • plaque • calcium • pre-rinse
The concentration of fluoride (F) in oral fluids has a profound effect on the de-/remineralization process (Margolis and Moreno, 1990; ten Cate, 1990; Featherstone, 2000). Given the role of plaque in the caries process, this observation has focused attention on the "bioavailable" plaque F reservoirs that can persistently increase F concentrations in plaque fluid. Calcium-to-fluoride (Ca-F) binding appears to play a central role in the formation of these plaque F reservoirs, either in the form of phosphate-contaminated "calcium fluoride-like" plaque deposits (Arends and Christoffersen, 1990; Rølla and Saxegard, 1990; Vogel et al., 2006a,b), or as bacterial calcium-fluoride complexes (Rølla and Bowen, 1977; Rose et al., 1996). Unfortunately, the low concentration of plaque fluid or salivary Ca (  1 mmol/L to 3 mmol/L) relative to the amount of applied F after use of F dentifrice or rinse ( 12 mmol/L) implies that Ca-F reservoir formation is limited by the rate at which additional Ca can be scavenged from Ca reservoirs in enamel, plaque, or saliva during the short period of F application. The relatively small amounts of these oral Ca-F reservoirs appear to produce only a transient increase in plaque fluid and salivary F (Vogel et al., 1992; Whitford et al., 2005). Previous studies have shown that if this limitation on the amount of easily available Ca is ameliorated by first pre-rinsing with a concentrated calcium lactate solution, a large increase in salivary fluoride can be obtained from a commercial dentifrice or, optimally, a F rinse (Vogel et al., 2006a,b). The purpose of this study was to examine if these increases are also found in plaque fluid F. Since, except shortly after F administration, the source of the plaque fluid F is the bioavailable Ca-F reservoirs that can be easily released from the plaque mass (Vogel et al., 1992), a second goal of this study was to examine the amount of buffer-extractable F or Ca (pH 6.8 and pH 4.8) in plaque samples after a F application with or without the Ca pre-rinse procedure.
Study Participant Protocol All procedures were performed with the informed consent of the study participants following protocols approved by the institutional review board (IRB) of the American Dental Association and reviewed by the IRB of the National Institute of Standards and Technology. These participants were screened before inclusion, to ensure their good oral health and normal salivary gland function (assessed by the measurement of unstimulated salivary flow in each potential participant). The same 13 individuals participated in all experiments; however, in some experiments, adequate plaque samples could not be recovered from all the participants. All study participants lived in an area with fluoridated water (2003 average F = 1.01 µg/g; range, 0.69 µg/g to 1.13 µg/g) and used their normal oral hygiene procedures between periods of plaque accumulation (i.e., no special tooth-cleaning procedures were performed). The rinses (described below) were given in a random order, and the participants were not aware of the identity of the test material. The analysts were not aware of the identity of the recovered samples. Before each experiment, the participants accumulated plaque for 48 hrs and fasted overnight. In the morning, these individuals received the experimental treatment. Plaque samples were collected 1 hr later.
Rinse Administration
Plaque Fluid and Plaque Recovery
Extraction of F and Ca from Plaque
Analytical Procedure
Statistical Methods
The total amounts of F in the 2 (pH 6.8 and pH 4.8) recovered samples (Table 1  ) were not statistically different (average difference 12%). Except for the 5th pH 4.8 extraction and subsequent acid extraction, where the NaF rinse samples were not different from the "no rinse" samples, both experimental rinses increased the F in all extractions. The total one-hour post-rinse plaque F recovered after the NaF rinse was significantly increased ( 5x) over the "no rinse" samples (Table 1). With regard to the Ca rinse/F rinse, every buffer extraction, as well as the subsequent acid extraction, was significantly (p < 0.001) increased over the NaF rinse samples. Reflecting these very large increases, the average total F of the Ca rinse/F rinse samples was significantly (p < 0.001) increased 64x (Table 1) over the "no rinse" samples. The Ca rinse alone did not significantly increase the F in any of the extracts.
The plaque fluid F from the pH 6.8 and pH 4.8 sample values also did not differ significantly for any test regimen (average difference
Since, as with the plaque total F and plaque fluid F, the total Ca extracted from the 2 (pH 6.8 and pH 4.8) samples was quite similar (average difference
The "no rinse" and one-hour NaF total F and total Ca concentrations observed in this study are similar to those reported in previous studies (Tatevossian, 1990; Vogel et al., 2000a,b, 2001; Whitford et al., 2005). The agreement in these quantities for the 2 collected samples demonstrates that they are nominally identical. The repeated extraction of the plaque samples with large volumes of pH 6.8 buffer failed to remove all the F. Indeed, the concentrations in these extracts were relatively similar (except extract 1, where a smaller extraction volume was used). However, each Ca rinse/F rinse plaque extract had much more F than the corresponding extract for the NaF rinse samples. In fact, the F remaining in the plaque after the 5th pH 6.8 extraction (i.e., the acid F value) for the Ca rinse/F rinse plaque samples exceeded the total F in the NaF rinse samples by 5x. With the pH 4.8 extractions, 4 extractions of plaque appeared to have exhausted the F reservoir in the NaF rinse samples, while the F remaining in the plaque after the 5th extraction (the acid F) of the Ca rinse/F rinse contained more F than the sum of the F in the first 2 pH 4.8 extracts of the NaF plaque samples. As a consequence of these increases, the average total F of the Ca rinse/F rinse group increased nearly 12x relative to the NaF rinse samples. There was a trend toward higher total plaque Ca levels in the NaF and Ca rinse/F rinse samples, but only the Ca rinse group showed a significant increase above the "no rinse" levels, and no correlation was seen between the plaque Ca and F except in the NaF rinse samples. Other studies (Whitford et al., 2005; Pessan et al., 2006) also found a significant correlation between plaque Ca and F, as well as a significant increase in total plaque Ca, in study participants who used a NaF dentifrice. Such results are compatible (Whitford et al., 2005) with the fixing of salivary Ca into plaque Ca-F reservoirs by the applied F. Unfortunately, as noted in the INTRODUCTION, the low concentration of salivary Ca relative to the amount of applied F severely limits the accumulation in plaque of such Ca-F moieties from saliva, while the low amount of plaque fluid Ca limits the in situ formation of plaque Ca-F from this source. The large increase in plaque F from the 2 rinse procedures, as well as the large Ca plaque reservoir formed by the Ca-only rinse, suggests that the Ca pre-rinse procedure can overcome these limitations. The relatively modest increase in plaque Ca when the Ca rinse was used immediately before the F rinse, relative to the NaF rinse alone, and the decrease in Ca relative to the Ca rinse alone, suggest that the water component of the subsequent F rinse was extracting Ca not yet firmly fixed in plaque (Vogel et al., 2006b). These results also suggest that there is little increased potential for calculus formation, relative to the NaF rinse group, as a result of the Ca pre-rinse procedure. In fact, several studies (Schaeken and van der Hoeven, 1990, 1993) suggest that the Ca lactate used in this study decreases calculus formation. The 12x increase in total plaque F from the Ca rinse/F rinse was reflected in a 5x increase in the average plaque fluid F relative to the NaF rinse. This increase in plaque fluid F was similar to the increases we have previously observed in saliva (Vogel et al., 2006a,b). Recently, however, a 39x increase in whole saliva F, but only about a 4x increase in total plaque F, was reported at the 2007 IADR General Session (New Orleans, LA, USA) in studies done with this same Ca pre-rinse/NaF rinse system (Whitford, personal communication). It should also be noted that studies with a Ca pre-rinse and a F dentifrice found a much smaller increase in salivary F (Pessan et al., 2006; Vogel et al. 2006), and no increase in plaque F (Pessan et al., 2006). Although we have attributed these results to Ca binding by dentifrice surfactants (Vogel et al., 2006b), two recent Ca rinse/F dentifrice studies involving an in situ model found large increases in plaque F (M. Buzalaf, private communication). In conclusion, although these very large increases in salivary F and, more importantly, in plaque fluid F suggest that a Ca pre-rinse, or other Ca pre-treatment, may increase the effectiveness of F-containing therapeutic agents, the persistence of these increases, which appears to be the key to obtaining an increased cariostatic effect, remains to be examined. It is noteworthy in this regard that the increase in plaque F induced by this treatment appears to persist, despite repeated neutral or low pH extractions.
This study was supported by a grant from the American Dental Association Foundation, and by USPHS Research Grants DE05354 and DE14707 to the American Dental Association Foundation from the National Institutes of Health–National Institute of Dental and Craniofacial Research and is part of the dental research program conducted by the National Institute of Standards and Technology in cooperation with the American Dental Association Foundation. Certain commercial materials and equipment are identified in this paper to specify the experimental procedure. In no instance does such identification imply recommendation or endorsement by the National Institute of Standards and Technology or the American Dental Association Foundation, or that the material or the equipment identified is necessarily the best available for the purpose. Received for publication April 17, 2007. Revision received January 2, 2008. Accepted for publication January 23, 2008.
Journal of Dental Research, Vol. 87, No. 5,
466-469 (2008)
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ABSTRACT
INTRODUCTION
-tricalcium phosphate-containing chewing gum and a subsequent sucrose challenge. J Dent Res 79:58–62.