Research Article

Analyses of biofilms accumulated on dental restorative materials

Suzana  Beatriz  Portugal  de  FÚcio, dds, ms,  Regina  Maria Puppin-Rontani, dds, ms, phd,

FabÍola  Galbiatti  de  Carvalho, dds, ms,  Renata  de  Oliveira  Mattos-Graner, dds, ms, phd,

LourenÇo  Correr-Sobrinho, dds, ms, phd  &  Franklin  Garcia-Godoy, dds, ms

Abstract: Purpose: To qualitatively and quantitatively assess the architectural arrangement of microorganisms in biofilm developed on the surface of different restorative materials: ceramic (C), resin composite (RC), conventional (CGIC) and resin-modified glass-ionomer cements (RMGIC). Methods: Streptococcus mutans was used to develop a biofilm that adhered to the surfaces of the selected material disks in 30 days. The specimens were stained and analyzed by confocal laser scanning microscopy and COMSTAT. Among biofilm properties, mean thickness, total bio-volume, roughness coefficient and surface-to-volume ratio were investigated, as well as characteristics of the distribution and architecture of viable/nonviable cells in the biofilm. Results: Only the mean biofilm thickness was statistically significantly different among the restorative materials tested. C and RC accumulated the thickest biofilms. Qualitative analysis showed cellular aggregates and fluid-filled channels penetrating to a considerable depth of the biofilm. In addition, images demonstrated a progression of more viable cells in superficial regions of the biofilm to proportionally more nonviable cells in the deeper regions of the biofilms near the disk. (Am J Dent 2009;22:131-136).

*: Dr. Regina M. Puppin-Rontani, Department of Pediatric Dentistry, Piracicaba Dental School – Unicamp,  Av. Limeira 901 – 13414-018, Piracicaba, SP, Brazil. E-*: rmpuppin@fop.unicamp.br

 Research Article

Cytotoxicity of resin-based light-cured liners

Josimeri  Hebling, dds, ms, phd,  Fernanda  Campos  Rosetti  Lessa, dds, ms,  

Indri  Nogueira, dds, ms, Ricardo  Marins  de  Carvalho, dds, ms, phd  &  Carlos  Alberto  de  Souza  Costa, dds, ms, phd

Abstract: Purpose: To evaluate the cytotoxic effects of resin-based light-cured liners on culture of pulp cells. Methods: Discs measuring 4 mm in diameter and 2 mm thick were fabricated from TheraCal (TCMTA), Vitrebond (VIT), and Ultrablend Plus (UBP). These specimens were immersed in serum-free culture medium (DMEM) for 24 hours or 7 days to produce the extracts. After incubating the pulp cells for 72 hours, the extracts were applied on the cells and the cytotoxic effects were determined based on the cell metabolism (MTT), total protein expression and cell morphology (SEM). In the control group, fresh DMEM was used. Data from MTT analysis and protein expression were submitted to Kruskal-Wallis and Mann-Whitney tests at the preset level of significance of 5%. Results: When in contact with the 24-hour extract, TCMTA, VIT, and UBP decreased the cell metabolism by 31.5%, 73.5% and 71.0%, respectively. The total protein expressed by the cells in contact with VIT and UBP was lower than TCMTA and DMEM (Mann-Whitney, P< 0.05). When in contact with the 7-day extract, TCMTA, VIT, and UBP decreased the metabolic activity by 45.9%, 77.1% and 64.4%, respectively. All the liners expressed statistically lower amounts of proteins when compared to the control. A reduction in the number of cells was observed for all liners. The remaining cells from TCMTA group resembled those from the control group while for VIT and UBP the cells presented significant morphological alterations. (Am J Dent 2009;22:137-142).

*: Dr. Carlos Alberto de Souza Costa, University of São Paulo State, UNESP, Rua Humaitá, 1680 - CEP: 14.801-903, Araraquara, SP, Brazil.  E-*: casouzac@foar.unesp.br

Effect of desensitizing agents on dentin permeability

Hiroshi  Ishihata, dds, phd, Masafumi Kanehira, dmd, phd, Tomoko Nagai, dds,

Werner J. Finger, phd, dr med dent, Hidetoshi Shimauchi, dds, phd  &  Masashi Komatsu, dds, phd

Abstract: Purpose: To investigate the in vitro efficacy of two dentin desensitizing products at reducing liquid permeability through human dentin discs. The tested hypothesis was that the products, in spite of different chemical mechanisms were not different at reducing or eliminating flow through dentin discs. Methods: Dentin slices (1 mm thick) were prepared from 16 extracted human third molars and their permeability was indirectly recorded in a split chamber model, using a chemiluminescence technique, after EDTA treatment (control), after soaking with albumin, and after desensitizer application. Two products were studied: MS Coat, a self-curing resin-containing oxalate product, and Gluma Desensitizer, a glutaraldehyde/HEMA-based agent without initiator. The dentin slices were mounted between an upper chamber, filled with an aqueous solution of 1% potassium ferricyanide and 0.3% hydrogen peroxide, and a lower chamber filled with 1% sodium hydroxide solution and 0.02% luminol. The upper solution was pressurized, and upon contact with the luminol solution a photochemical signal was generated and recorded as a measure of permeability throughout two consecutive pressurizing cycles at 2.5 and 13 kPa (26 and 133 cm H₂O), respectively. Results: The permeability of the control and albumin-soaked samples was similarly high. After application of the desensitizing agents, dentin permeability was reduced to virtually zero at both pressure levels (P< 0.001). (Am J Dent 2009;22:143-146).

*: Prof. Dr. Werner J. Finger, Department of Preclinical Dentistry, School of Dental Medicine, University of Cologne, Kerpener Strasse 32, D-50931 Cologne, Germany. E-*: wjfinger@aol.com

Research Article

Fracture resistance after simulated crown lengthening and forced

tooth eruption of endodontically-treated teeth restored with a fiber post-and-core system

Qing-Fei  Meng, mds,  Li-Juan Chen, mds,  Jian Meng, mds,  Ya-Ming Chen, dds, phd, 

Roger J. Smales, mds, ddsc &  Kevin H-K. Yip, bds, med, mmedsc, phd

Abstract: Purpose: To evaluate the effect of ferrule preparation length on the fracture resistance after simulated surgical crown lengthening and after forced tooth eruption of endodontically-treated teeth restored with a carbon fiber-reinforced post-and-core system. Methods: 40 extracted endodontically-treated mandibular first premolars were decoronated 1.0 mm coronal to the buccal cemento-enamel junction. The teeth were divided randomly into five equal groups. The control group had no ferrule preparation (Group A). Simulated crown lengthening provided ferrule preparations of 1.0 mm (Group B) and 2.0 mm (Group C). Simulated forced tooth eruption provided ferrule preparations of 1.0 mm (Group D) and 2.0 mm (Group E). After restoration with a carbon fiber post-and-core system, each root was embedded in an acrylic resin block from 2.0 mm apical to the margins of a cast Ni-Cr alloy crown, and loaded at 150° from the long axis in a universal testing machine at a crosshead speed of 1.0 mm /minute until fracture. Data were analyzed using ANOVA with Tukey HSD tests, and Fisher’s exact test, with α= 0.05. Results: Mean failure loads (kN) for Groups A, B, C, D and E were: 1.13 (SD= 0.15), 1.27 (0.18), 1.02 (0.11), 1.63 (0.14) and 1.92 (0.19), respectively. Significant differences were shown for the effects of treatment method and ferrule length, with significant interaction between these two sources of variation (P< 0.0001). Increased apical ferrule preparation lengths resulted in significantly increased fracture resistance for simulated forced tooth eruption (P< 0.0001), but not for simulated crown lengthening (P≥ 0.24). (Am J Dent 2009;22:147-150).

*: Prof. Ya-Ming Chen, Department of Prosthetic Dentistry, 140 Hanzhong Road, Nanjing, Jiangsu Province, 210029, PR China.  E-*: yaming_chen@yahoo.com

 Research Article

Effects of intrapulpal temperature change induced by visible light units

on the metabolism of odontoblast-like cells

Pedro Paulo Chaves de Souza, dds, ms, phd,  Josimeri Hebling, dds, ms, phd, Mariana Grecco Scalon, dds, Andreza  Maria  Fábio  Aranha, dds, ms, phd  &  Carlos  Alberto  de  Souza  Costa, dds, ms, phd

Abstract: Purpose: To investigate the effects of intrapulpal temperature changes induced by a quartz tungsten halogen (QTH) and a light emitting diode (LED) curing units on the metabolism of odontoblast-like cells. Methods: Thirty-six 0.5 mm-thick dentin discs obtained from sound human teeth were randomly assigned into three groups: QTH, LED and no light (control). After placement of the dentin discs in pulp chamber devices, a thermistor was attached to the pulpal surface of each disc and the light sources were applied on the occlusal surface. After registering the temperature change, odontoblast-like cells MDPC-23 were seeded on the pulpal side of the discs and the curing lights were again applied. Cell metabolism was evaluated by the MTT assay and cell morphology was assessed by SEM. Results: In groups QTH and LED the intrapulpal temperature increased by 6.4°C and 3.4°C, respectively. The difference between both groups was statistically significant (Mann-Whitney; P< 0.05). QTH and LED reduced the cell metabolism by 36.4% and 33.4%, respectively. Regarding the cell metabolism, no statistically significant difference was observed between both groups (Mann-Whitney; P> 0.05). However, when compared to the control, only QTH significantly reduced the cell metabolism (Mann-Whitney; P< 0.05). It was concluded that the irradiance of 0.5 mm-thick human dentin discs with a QTH in comparison to a LED curing unit promoted a higher temperature rise, which propagates through the dentin negatively affecting the metabolism of the underlying cultured pulp cells. (Am J Dent 2009;22:151-156).

*: Dr. Carlos Alberto de Souza Costa, Department of Physiology and Pathology, School of Dentistry, University of São Paulo, UNESP, Rua Humaitá, 1680 - CEP: 14.801-903, Araraquara, SP, Brazil.  E-*: casouzac@foar.unesp.br

Research Article

Radiopacity of an endodontic core material

Emre  Bodrumlu, dds, phd  &  Kahraman  Gungor,  dds, phd

Abstract: Purpose: To evaluate and compare the relative density values of a new endodontic core material (Resilon) and conventional gutta-percha. Methods: 15 Resilon and gutta-percha discs for testing radiopacity were prepared by first coiling three size 35 cones. The coiled material was evenly compressed into the well of a brass mold 10 mm in diameter and a depth of 1 mm. All radiographs were taken on No. 4 ultra-speed dental film. The radiopacity of Resilon, gutta-percha, and dentin slices were then measured with a digital transmission densitometer. Results: The mean radiopacities of Resilon and gutta-percha coiled discs were 4.06 and 7.21 mm of aluminum equivalence respectively, which exceeded the minimal requirement of 3 mm. Both tested materials had lower radiopacity values than dentin. There were highly significant differences in radiopacity values among the tested specimens (P< 0.01). Additionally, the mean radiopacity values in root canals were 5.50 and 8.52 mm Al for Resilon and gutta-percha, respectively (P< 0.01). (Am J Dent 2009;22:157-159).

*: Dr. Emre Bodrumlu, Department of Operative Dentistry and Endodontics, Faculty of Dentistry, Ondokuz Mayis University, 55139 Kurupelit-Samsun, Turkey. E-*:  bodrumlu@omu.edu.tr

Research Article

Comparison of the color stability of flowable and universal resin composites

Bin  Yu, mche  &  Yong-Keun  Lee, dds, phd

Abstract: Purpose: To evaluate the color stability of flowable resin composites after ultraviolet (UV) irradiation and thermocycling, and to compare the color stability with the corresponding shade universal resin composites of the same brand. Methods: Five specimens each ( 10 mm in diameter and 2 mm in thickness) of four brands of flowable and universal resin composites (shade A2) were light-cured. Baseline color was measured on a reflection spectrophotometer over a white background. Artificial aging was performed with a filtered xenon lamp (irradiation intensity: 550 W/m²) for 24 hours and then color was measured. Afterwards, thermocycling was performed for 3,000 times between 5 and 55°C and color was measured again. Changes in color (∆E^*_ab) and color parameters (∆L^*, ∆a^*, ∆b^* and ∆C^*_ab) during the UV irradiation, the thermocycling and both aging protocols were calculated. Differences in the color change of the flowable and the universal resin composites were analyzed with two-way repeated measures ANOVA by the combination of brand and type (eight groups) of materials and aging protocol (P< 0.05). Results: Color changes during the UV irradiation, the thermocycling and both aging protocols were in the range of 2.3-6.8, 0.6-0.8 and 2.6-6.2 ∆E^*_ab units, respectively. Flowable resin composites showed higher color change during the UV irradiation and was significant in three of the four brands (P< 0.05). The same results were obtained after both aging protocols. Changes in color parameters were varied by the brand and type of resin composites (P< 0.05). (Am J Dent 2009;22:160-164).

*: Dr. Yong-Keun Lee, Department of Dental Biomaterials Science, School of Dentistry, Seoul National University, 28 Yeongeon-dong, Jongro-gu, Seoul, Korea. E-*: ykleedm@snu.ac.kr

Research Article

Effect of irradiance and light source on contraction stress, degree of

conversion and push-out bond strength of composite restoratives

Leonardo GonÇalves Cunha, dds, ms, phd, Roberta Caroline Bruschi Alonso, dds, ms, phd,

Carmem Silvia Costa Pfeifer, dds, phd, Mario Fernando De Góes, dds, ms, phd, Jack Liborio Ferracane, ms, phd &  Mário Alexandre Coelho Sinhoreti, dds, ms, phd

Abstract: Purpose: To evaluate the influence of five curing methods on contraction stress, stress rate, and degree of conversion (DC) of a composite and on bond strength of composite restoratives. Methods: For the stress test, composite was applied between two 5-mm diameter glass rods, mounted in a servohydraulic machine. Stress rates were calculated as the change in stress vs. time.  DC was measured by FTIR. Bond strength testing was performed using a push-out test in bovine incisors. The C-factor was 3.0 for all tests. Five methods were evaluated: High Intensity LED (LED HI), Continuous Halogen Light (QTH CL), Medium Intensity LED (LED MI), Low Intensity LED (LED LI), and Pulse Delay Halogen Light (QTH PD). Results were analyzed by ANOVA and Tukey’s test (α = 0.05). Results: Stress values ranged from 9.25 MPa (QTH PD) to 10.46 MPa (LED MI). No statistical difference was observed among the methods. Bond strength values ranged from 24.6 MPa (LED HI) to 35.4 MPa (QTH PD), with the QTH PD presenting a statistically higher value compared to the other methods. Stress rate and bond strength presented an inverse linear correlation (r² = 0.79). LED HI presented the highest maximum stress rate, followed by LED MI, QTH CL, LED LI, and QTH PD. The reduction in stress rate observed for the low intensity groups was associated with a general increase in bond strength, with no adverse effect on the degree of conversion of the restorative composite. (Am J Dent 2009;22:165-170).

*: Dr. Leonardo Gonçalves Cunha, UNITAU - Department of Dentistry - Prosthodontics Area, Rua Expedicionário Ernesto Pereira, 110, Centro CEP 12020-330, Taubaté, SP, Brazil.  E-*: leonardogcunha@yahoo.com

Research Article

Effect of a bleaching agent on abrasion of resin-based restoratives

Juliana Jendiroba Faraoni-Romano, dds, ms, phd,  Cecilia Pedroso Turssi, dds, ms, phd 

&  MÔnica Campos Serra, dds, ms, phd

Abstract: Purpose: To evaluate the effect of a 10% carbamide peroxide-containing bleaching agent on brushing abrasion of esthetic restorative materials. Methods: Using a randomized complete block design, 150 specimens (n = 15) measuring 3 x 3 x 2 mm were fabricated into acrylic resin cylinders, using one of the restorative materials: a microfilled resin composite (A1), a hybrid resin composite (Ch), a flowable resin composite (Wa), a resin-modified glass-ionomer cement (Fj) and a polyacid-modified resin composite (Dy). The bleaching agent or artificial saliva (control) was applied for 2 hours/day. After that, 120 brushing strokes were simulated automatically and the samples were kept in artificial saliva. Such bleaching/brushing cycle was performed daily for 21 days. Wear depth was assessed using profilometry. Results: Bleaching did not show significant effect on wear depth. There was a significant difference among the restorative materials. Tukey’s test showed that (A1=Ch) < (Wa) < (Fj) and that Dy was only different from Fj. (Am J Dent 2009;22:171-174).

*: Dr. Mônica Campos Serra, Faculdade de Odontologia de Ribeirão Preto – USP, Departamento de Odontologia Restauradora, Av. do Café, s/nº - Monte Alegre - CEP 14040-904, Ribeirão Preto - SP – Brazil. E-*: mcserra@forp.usp.br

Research Article

The effect of coffee solution on tooth color during home bleaching

applications

Mariana  Lerner  Attia, dds,  FlÁvio  Henrique  Baggio Aguiar, dds, ms, phd, Paula  Mathias, dds, ms, phd, Glaucia  Maria  Bovi  Ambrosano, ms, phd,  CÉres  MendonÇa  Fontes, dds, ms

&  Priscila  Christiane  Suzy  Liporoni, dds , ms , phd

Abstract: Purpose: To quantify the change in color of human and bovine teeth exposed to a coffee solution during a 16% carbamide peroxide (16%CP) home application bleaching treatment using photoreflectance analysis. Methods: 40 enamel slabs (4x4x2 mm) were obtained from seven unerupted third human molars and seven bovine incisors, which were allocated into four groups: G1: human control group whitened and not exposed to a coffee solution; G2: bovine control group whitened and not exposed to a coffee solution; G3: human teeth whitened and exposed to a coffee solution; G4: bovine teeth whitened and exposed to a coffee solution. The home bleaching procedure was performed using 16%CP gel applied in a 1 mm-thick layer on the tooth surface over a period of 6 hours per day, for 28 days. Tooth color was evaluated using photoreflectance analysis throughout the bleaching procedure at intervals of 7, 14, 21 and 28 days, and post-treatment at 7, 15 and 30 days. Results were statistically analyzed using ANOVA and Tukey-Kramer tests. Results: After 28 days of bleaching treatment, no significant difference was detected between photoreflectance analysis of specimens exposed to coffee solution and specimens not exposed to coffee solution (P< 0.05). However, when the teeth were exposed to a coffee solution during home bleaching treatment, the whitening effect was observed to be less stable (P< 0.05). Bovine and human enamel substrates behaved similarly in terms of staining and bleaching effects, although they presented inherent differences in color (P< 0.05). (Am J Dent 2009;22:175-179).

Research Article

In vitro assessments of experimental NaF dentifrices containing

a prospective calcium phosphate technology

Robert L. Karlinsey, phd, Allen C. Mackey, bs, George K. Stookey, phd, ms  &  Aaron M. Pfarrer, bs

Abstract: Purpose: To determine the fluoride dose response of experimental NaF dentifrices containing a prospective calcium phosphate technology, along with the corresponding relative enamel and dentin abrasion values. Methods: 3 mm diameter bovine enamel specimens were mounted, ground and polished, and softened in a carbopol-lactic acid solution (pH = 5.0) for 36 hours at 37°C. Specimens were then measured for baseline Vickers microhardness and stratified (N=18, mean VHN =33) into eight groups. These groups consisted of a placebo paste, four test dentifrices (A, B, C, D) with three of the four (A, B, C) containing a promising calcium phosphate ingredient, Crest Cavity Protection, MI Paste Plus, and PreviDent Booster 5000. The groups were cycled in a lesion reversal pH cycling model consisting of four 2-minute treatment periods (diluted 1:3 with DI water) and one 4-hour acid challenge (carbopol-lactic acid, pH = 5.0) per day. Between these events, specimens were immersed in artificial saliva (pH = 7.0). After 20 days of cycling, the specimens were microdrilled and analyzed for fluoride content, and also measured for Vickers surface microhardness after 10 and 20 days of cycling and after a 2-hour and 16-hour post-cycle acid challenge (carbopol-lactic acid, pH = 5.0). Separately, relative dentin and enamel abrasion (RDA and REA) were performed using the ADA recommended radiotracer method. Results: A fluoride dose response was observed for the test dentifrices after 10 and 20 days of pH cycling, with test dentifrice C promoting the highest remineralization among the groups while both the MI Paste Plus and PreviDent systems provide the least remineralization (one-way ANOVA, SNK, P< 0.05). With respect to enamel fluoride uptake, the group facilitating the highest incorporation of fluoride into the enamel lesion was test dentifrice C, while the least effective NaF system was the MI Paste Plus (one-way ANOVA, SNK, P< 0.05). In terms of formulation abrasion, the REA scores were similar among the test dentifrices, MI Paste Plus, and PreviDent and compared favorably to the ADA reference material score (one-way ANOVA, SNK, P< 0.05); relative to the ADA reference material RDA score, the data indicate that MI Paste Plus was essentially non-abrasive, while PreviDent was significantly more abrasive to dentin (one-way ANOVA, SNK, P< 0.05). Altogether, these data show the developmental test dentifrices demonstrate a fluoride dose response and show great promise in remineralizing white-spot enamel lesions relative to MI Paste Plus and PreviDent. (Am J Dent 2009;22:180-184).

*: Dr. Robert L. Karlinsey, Indiana Nanotech, LLC,  351 West 10th Street, Suite 309, Indianapolis, IN 46202, USA.  E-*: rkarlins@gmail.com

Research Article

In vitro remineralization efficacy of NaF systems

containing unique forms of calcium

Robert L. Karlinsey, phd, Allen C. Mackey, bs  &  George K. Stookey, phd, ms

Abstract: Purpose: To determine the in vitro remineralization potential of a new calcium phosphate technology in a 1000 ppm F system. Methods: 3 mm diameter bovine enamel specimens were mounted, ground and polished, and softened in a carbopol-lactic acid solution (pH = 5.0) for 36 hours at 37°C. Specimens were then measured for baseline Vickers microhardness and stratified (N=10, mean VHN =35) into the following groups: Group A: distilled water (negative control); Group B: MI Paste Plus (900 ppm F); Group C: Theramed SOS (1450 ppm F); Group D: "control" dentifrice (1000 ppm F); and, Group E: "test" dentifrice (1000 ppm F) admixed with a new functionalized calcium phosphate system. The groups were then cycled for 10 days in a pH cycling model consisting of four 2-minute treatment periods (diluted 1:3 with distilled water) and one 4-hour acid challenge (carbopol-lactic acid, pH = 5.0) per day. Between these events, specimens were immersed in artificial saliva (pH = 7.0). After 10 days of cycling, the specimens were measured for Vickers surface microhardness and were subsequently microdrilled, with the powder measured for fluoride content using a calibrated fluoride-sensitive electrode. Results: Significant differences resulted between the distilled water and fluoride-containing groups. Among the fluoride-containing groups, Group B demonstrated statistically low levels of enamel fluoride deposition and ΔVHN, while Group E statistically outperformed Group D. Among the groups with different calcium systems (Groups B, C, and E), Groups C and E were found to be statistically equivalent and superior to Group B with respect to both bioavailable fluoride and ΔVHN. Based on our results, these data demonstrated the combination of a new calcium phosphate technology plus 1000 ppm F, produced significantly greater remineralization relative to both the 1000 ppm F test dentifrice and MI Paste Plus, and was statistically equivalent to Theramed SOS. (Am J Dent 2009;22:185-188).

*: Dr. Robert L. Karlinsey, Indiana Nanotech, LLC,  351 West 10th Street, Suite 309, Indianapolis, IN 46202 , USA . E-*: rkarlins@gmail.com

 Research Article

Replicate single-use comparative study of plaque removal

with two contemporary manual toothbrushes

GÉza T. TerÉzhalmy, dds , ma ,  Pat Walters, rdh, msdh, msob, Robert D. Bartizek, ms, Julie M. Grender, phd &  Aaron R. Biesbrock, dmd, phd, ms

Abstract: Purpose: To compare the plaque removing efficacy of Oral-B Advantage 123 and Colgate 360º manual toothbrushes on a single brushing after a 24-hour void in oral hygiene. Methods: This study had a two-treatment (Advantage 123 versus Colgate 360°), examiner-blind, randomized, four-period (visit) crossover design. At the first (baseline) visit, subjects received a plaque examination using the Rustogi Modified Navy Plaque Index (RMNPI) after which they were instructed to brush for 1 minute in their normal manner with their assigned toothbrush and a marketed dentifrice and unaided by access to a mirror. Post-brushing plaque was then assessed. At three further visits, each separated by a period of 3-8 days, brushes were assigned to subjects according to their treatment sequence and the same brushing and plaque grading procedure as at baseline was followed. Results: 50 subjects were enrolled in the replicate single-use study; all were included in the analysis. Oral-B Advantage 123 was significantly better than Colgate 360º at reducing whole mouth (P= 0.006), gingival margin (P= 0.010) and approximal (P= 0.040) plaque scores. (Am J Dent 2009;22:189-192).

*: Dr. Aaron Biesbrock, P&G Health Care Research Center , 8700 Mason-Montgomery Road, Mason , OH 45040 , USA .  E-*: biesbrock.ar@pg.com

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