Effect of two NaF dentifrices (1450 and 5000 ppm F) applied 2 or 3 times/day on F retention and pH drop in plaque
A Nordström, D Birkhed
Department of Cariology, Institute of Odontology, Sahlgrenska Academy, University of Gothenburg, Sweden
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Introduction
The prevalence of dental caries has declined in many industrialised countries since the late 1960s. This is mainly a result of the daily use of fluoride (F) dentifrices [Glass, 1982; Bratthall et al., 1996; Marinho et al., 2003]. The Swedish Council on Technology Assessment in Health Care [SBU, 2002] reported strong scientific evidence that daily use of F toothpaste is an effective method for preventing caries in permanent teeth.
Various factors influence the efficacy of F toothpaste, including the concentration of F in the paste, the amount of toothpaste applied on the brush, the frequency of tooth brushing and the post-brushing water rinsing behaviour [Davies et al., 2003]. In a recent study, we concluded that 5000 ppm F toothpaste without post-brushing water rinsing resulted in higher F concentration in both plaque and saliva compared to 1450 ppm F toothpaste [Nordström et al., 2009]. Several trials have reported an association between brushing frequency and caries incidence. In 3-year clinical trials, the caries increment in subjects who brushed only once a day was 20-30% higher than those who brushed twice a day [Chester et al., 1992; Chestnutt et al., 1998]. Also, the penetration of F into plaque biofilms seems to increase with the duration of NaF exposure [Watson et al. 2005].
According to instructions by the manufacturer, (delivered in the package) the 5000 ppm toothpaste (Duraphat®) is recommended for tooth brushing “3+3+3” i.e. three minutes, three times a day, during three months. However, most persons who need extra F therapy may not be motivated to brush their teeth up to 3 times a day. Seppä et al. [1997] showed that just rinsing with a toothpaste–water mixture (without using a toothbrush) appears as a good alternative for adults who need extra F therapy. We believe that using toothpaste on the fingertip and as a “lotion” massaging the buccal surfaces could be an alternative for a third application of F during daytime. There are no studies evaluating the effect of a third daily use of a 5000 ppm F dentifrice compared to two applications, on the retention of F in plaque and saliva. The aim of this study is therefore to investigate the retention of F in plaque and saliva and the pH drop in plaque using a 5000 ppm dentifrice or 1450 ppm F (as NaF) two or three times a day, in order to evaluate the potential benefit of using toothpaste three times a day. In addition, the “toothpaste massage method” will be evaluated in this respect.
Material and methods
Study Design
The investigation is a 6-cell controlled randomized, non blinded study. The outcome measure will be retention of F in plaque and saliva. Plaque-pH change in situ after a sucrose rinse is used as an indicator of the microbiological activity.
Subjects
A total of 16 volunteers will carefully be selected in order to be capable of following the instructions given. They should be in general good health and have a minimum of 24 teeth. The subjects are instructed not to use any other F products apart from the test toothpaste, not to eat or drink within two hours after last tooth brushing, not to use any proximal cleaning and not to use chewing gum or snuff, during the 2-week test periods. The study will be submitted to the Ethics Committee of Sahlgrenska Academy at Göteborg University.
Treatments
Six different tooth-brushing methods are used (see table below). One week prior to each brushing period, the subjects use F-free toothpaste twice daily and a waxed, F-free dental floss (Johnson & Johnson Products, New Brunswick, N.J., USA, or Sunstar Butler Laholm, Sweden) once daily. The teeth are cleaned professionally with a hand-piece and a rubber cup, using RDA 170 polish paste (CCS AB, Borlänge, Sweden) and flossed. The subjects are then carefully instructed to use one gram of toothpaste on the toothbrushes and to brush for two minutes, followed by expectoration and a single rinse with 10 ml of tap water for 10 seconds. Each 2-weeks test period, is followed by a wash-out period of one week, using the F-free toothpaste and floss. The dentifrices are Duraphat 5000 ppm F as NaF (Colgate-Palmolive AB, Danderyd, Sweden) and Pepsodent Superfluor 1450 ppm F as NaF (Lever Faberge´, Stockholm, Sweden). The subjects are asked to brush twice a day (in the morning after breakfast and in the evening before bedtime) or three times a day (in the morning after breakfast, after lunch and in the evening before bedtime). A new method, using F toothpaste as a “lotion” to massage the buccal surfaces is also evaluated. After lunch, the subjects are instructed to apply at least one centimetre of toothpaste on the fingertip and massaging the buccal surfaces for one min, followed by exploration of the toothpaste slurry and no water rinsing afterwards.
Toothpaste (ppm F) / Brushings/day / Massaging/day5000 / 2 / 1
5000 / 2
5000 / 3
1450 / 2 / 1
1450 / 2
1450 / 3
Measurements
Three measurements will be carried out, the first after one week and the second and third after 2 weeks in the following order: 1) F concentration in whole saliva after brushing/massaging with the test toothpastes (5000 or 1450 ppm F), 2) F accumulation in proximal plaque and 3) plaque-pH after a sucrose rinse.
1. Fluoride Concentration in Whole Saliva
After one week, the subjects are asked to brush or to massage their teeth with the same toothpaste and method as during the 2-week test period. Whole saliva samples are collected before the brush and repeated after 1, 3, 5, 10, 30 and 60 min. The samples are frozen at -20˚C until analysed for F. The saliva samples will be analysed blinded regarding subjects and methods. A volume of 200 μl of liquid, consisting of distilled water and TISAB III (dilution10:1) (Thermo Electron Corp., Waltham, USA) is added to the plaque samples. The suspension is homogenised by sonification for 20 s (Branson W185D, Dansbury, Connecticut, USA) in order to disperse the plaque. The samples are kept in a refrigerator at +4˚C over night. The tubes are then vibrated in a Minishaker MS1 (IKA, Wilmington, USA) for 20 s. 100 μl of the solution is placed as a drop on a Petri dish. The F concentration is measured by an ion-specific electrode (model 96-09, Orion Research, Boston, Mass., USA.) by carefully lowering the electrode into the fluid. The surface tension of the drop will ensure that the liquid enclose the entire membrane surface of the electrode. In order to calibrate the electrode in saliva, three standard solutions are used (0.1, 1.0, and 10 ppm F).
2. Fluoride Accumulation in Proximal Plaque
Proximal plaque is collected 2 hours after the last brushing occasion (in order to measure the F accumulated in plaque during the 2 weeks), using a F-free, extra-wide, waxed dental floss (Johnson & Johnson or Sunstar Butler), according to Sjögren and Birkhed [1993]. Plaque is collected from all proximal sites in two of the four quadrants. The collected plaque is transferred from the floss to a 2-ml plastic tube (Eppendorf PRC, Sarstedt, Nümbrecht, Germany). This is carried out by drawing the floss through a slit, which is cut in the lid of the tube. Plaque adhering to the floss is thereby left on the inside of the tube. The samples are frozen at -20˚C until analysed. The Eppendorf tubes are weighed before and after the plaque sampling and the plaque wet weight determined to the nearest 0.1 mg and the F concentration expressed as both ppm F and ng F per mg plaque. The plaque samples will be analysed in a similar way as the saliva samples (see above).
3. Plaque-pH
Plaque-pH is measured in two proximal sites in the remaining two quadrants (where plaque has not been collected) with the microtouch method, at baseline and at various time-points during 45 min. Plaque-pH is registered by inserting an iridium microelectrode (Beetrode® MEPH-1; W.P. Instrument, New Haven, Conn., USA) into 2 proximal sites in the premolar/molar region [Lingström et al., 1993]. For each individual, the same sites are used throughout the whole study, i.e. for all six test periods and all time points. The electrode is connected to an Orion SA 720 pH/ISE Meter (Orion Research, Boston, Mass., USA), equipped with a porous glass reference electrode (MERE 1; W.P. Instruments). A salt bridge is created in a 3 M KCL solution between the reference electrode and one of the subject’s fingers. After measuring resting pH (0 min), a mouth rinse with 10 ml 10% sucrose for 1 min is carried out. Plaque pH is then measured at 6 different time points (2, 5, 10, 15, 30 and 45 min) after the rinse.
Statistical Methods
Two-way ANOVA followed by Scheffe´s test are used to compare the six brushing methods, regarding accumulation of F in the proximal plaque, F concentration in saliva and plaque-pH response (as AUC). P-values < 0.05 are considered statistically significant.
Clinical Importance
In Sweden, the F concentration in dentifrices sold in ordinary stores varies between 400 and 1500 ppm. Dentifrice with 5000 ppm F can only be obtained at a pharmacy as an over-the-counter (OCT) product. Adults and teenagers with high caries risk are a suitable target group for this 5000 ppm F product, for example during orthodontic treatment and for elderly with hyposalivation due to medication or radiation. From a cariological point of view, it is interesting to evaluate the effect of a third application of a 5000 ppm F dentifrice on the retention of F in plaque and saliva. A new method using the toothpaste as a “lotion” and massaging the buccal surfaces is included in this study. The “massage method” should not replace tooth brushing, but might be used as an additional F application during daytime.
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