INVESTIGATIONS OF THE POTENTIAL EFFECTS OF DIFFERENT PREVENTIVE PRODUCTS ON TREATMENT AND PREVENTION OF ENAMEL SURFACE LOSS By Fowziya M. Ali Submitted in accordance with the requirements for the Degree of Doctor of Philosophy Faculty of Medicine and Health Department of Paediatric Dentistry, Leeds Dental Institute, Leeds, UK University of Leeds April 2018 "I hereby confirm that this submitted work is my own and that appropriate credit has been given where reference has been made to the work of others." This copy has been supplied on the understanding that it is copyright material and that no quotation from the thesis may be published without proper acknowledgement” © 2018 The University of Leeds and Fowziya M. Ali Dedicated to my family (My husband, my children, my parents, my sisters and brothers.) ACKNOWLEDGEMENTS Many people deserve thanks for help while this extremely challenging task was being done. Special gratitude goes to the head of the Paediatric Dentistry Department, Professor Monty Duggal, for his continued encouragement, guidance and his generosity in offering his long experience and knowledge during my PhD study. Many thanks go to Prof. Jack Toumba for his help and support, Dr Simon Strafford for offering his previous laboratory technical experience and guidance during the initial stages of the work which contributed considerably to this thesis. I gratefully value the comments and the scientific guidance involvement of The Head of Clinical Management trial / (Dentistry)/Clinical Trial Research / University of Leeds, Dr Catherine Fernandez, for her valued comments and scientific guided involvements during the clinical trial work. I sincerely appreciate the help and the assistance of the clinical trial study coordinator, Gillian Dukanovic, for her helpful well-organised involvement in clinical trial and Ashna Chavda for her assistance and helpful pleasant character. I gratefully acknowledge all the volunteers who participated in this clinical trial for their lovely and cooperative characters. I acknowledge the statisticians Dr Theresa Munyombwe and the Biostatistician Jing Kang for their precious advice and guidance on the statistical analysis. I would like to thank the Dental Laboratory Technicians for making the research appliances for the study and Ms Clare Verity for her coping with my requirements and materials for the study and my colleague Marina Malinowski who offered her help in designing the blindness of the study test products. Special thanks go to the Libyan authorities for their scholarship provision throughout the whole period of this demanding study. I express my very deep appreciation and gratitude to my husband, my children, my parents, my sisters and brothers for their unlimited love and everlasting support during my study life. ii ABSTRACT Maintaining strong healthy teeth for life is important; therefore the present study was concerned with the recognition of the significance of exploring some of the preventive and therapeutic products on the surface loss of enamel slabs subjected to acidic erosion and tooth brushing abrasion challenges using in vitro and in situ experiments. Study 1 In vitro assessment of the effect of fluoridated toothpastes on bovine and human enamel subjected to acidic erosion and tooth brushing abrasion. Aims: To assess the anti-erosive potential of toothpastes containing different delivery fluoride systems using bovine dental enamel (Part Ι) and human enamel (Part ІΙ) subjected to both acidic erosion and tooth brushing abrasion. Part Ι: Methods: Fifty bovine enamel slabs were mounted in acrylic blocks, ground flat, tested for flatness by scanning profilometry (Proscan 2000, Scantron, UK), standardised for hardness using a Knoop hardness tester (under 100 g load for 15 s) and divided into five experimental groups (E1-E5) including 10 specimens per group. The tested toothpastes were : E1 Meridol® group, (AmF/SnF2; 0.14%), E2 Elmex® anti- caries group (AmF; 0.14%F), E3 Pronamel®, (NaF, 1450 ppm F), E4 Elmex® sensitive plus (AmF; 0.14%F), and E5 Aronal® 0 ppm F. De/remineralisation cycling procedures were achieved with immersion of the slabs under static conditions in aliquots of citric acid (0.3%, pH 3.6) for 2 mins followed by soaking for 1hr in artificial saliva between the erosive challenges. The erosive challenges procedures were repeated 5 times daily for 28 days. All groups were subjected twice a day to brushing abrasion during application of a slurry of toothpaste/saliva (1: 3) with 15 tooth brushing strokes using a 300 g load and incubated overnight at 37 ˚C in night time saliva at the end of the last brushing of the enamel slabs. Erosion and abrasion depths (µm) were quantified using profilometry iii scanning (Proscan 2000, Scantron, UK) at 7, 14, 21 and 28 days. Part ΙΙ: Similar erosive / abrasive experimental procedures were used on human enamel specimens. Results: After 28 days of erosion cycling with tooth brushing abrasion, all groups showed different enamel surface loss (µm). One-way ANOVA, demonstrated that E1 showed similar trends with E5, whereas (E2, E3, E4 showed significantly less surface loss (p<0.05) compared to E1 and E2. Conclusions: The effect of dental erosion and tooth brushing abrasion combined can be decreased by toothpastes containing amine fluoride and sodium fluoride, whereas no protection was observed with the stannous and amine fluoride paste. Similar result trends were observed for both bovine and human enamel specimens under this erosive/abrasive model. Study 2 The effect of therapeutic products in combination on prevention of tooth surface loss. Aims: To study combined topical fluoridated measures on erosive and abrasive enamel wear in vitro. Methods: Sixty bovine enamel specimens were divided into four groups containing (15 samples per group) were subjected to cyclic de /remineralisation procedures. Demineralisation was performed with 0.3% citric acid (pH 3.6) 2 min / five times per day. The enamel slabs were immersed in day artificial saliva between each erosive challenge for 1 hour and incubated overnight in night saliva at 37 oC. Two times daily fluoride application with toothpaste and artificial saliva slurry (1:3 ratio) plus a tested mouthrinse with automated tooth brushing (15 strokes, 300 g load) for (2x2 min/ day) during the experimental process. The test product treatments were: group 1 Elmex® sensitive plus toothpaste (AmF, 1400 ppm F) Plus Elmex® erosion mouthrinse containing (500 ppm F as AmF/NaF and 800 ppm Sn as SnCl2) stannous chloride rinse (Elmex TP plus MR) 2 times / day; group 2 Pronamel® toothpaste (NaF, 1450 ppm F+ ® ® 5%KNO3) + Pronamel (NaF) mouthrinse (Pronamel TP plus MR) 2 mins x2 times / iv day; group 3 Pronamel® toothpaste (NaF, 1450 ppm F) 2 / day plus GC tooth mousse™ once/ a day for 5 mins (Pronamel® TP plus TM) after the last brushing; group 4 (0 ppm F toothpaste) as a control. Enamel loss was determined by scanning profilometry after the last experimental days at 7, 14, 21, and 28 days. Results: after 28 days significantly less enamel surface loss (µm) was observed with group1: (Elmex TP plus MR) (0.40±0.23) and group 2: (Pronamel TP plus MR) (0.60±0.28)(Pronamel TP plus TM) had (2.65±1.79) (p<0.01) compared to 0 ppm F control (1.84±1.85). Conclusions: The topical applications of preventive therapeutic measures in the form of AmF, 1400 ppm F toothpaste plus Elmex® erosion dental rinse (500 ppm F + 800 ppm Sn) or applications of NaF/5% KNO3 toothpaste plus NaF mouthrinse significantly decreased the effect of erosion and tooth brushing abrasion compared to combinations containing remineralising agent or fluoride-free toothpaste. Study 3: Investigations of therapeutic products on prevention of enamel surface loss under erosive and abrasive challenges in situ Aims: To investigate the effect of fluoridated toothpaste alone or in combination with mouth rinse on the prevention of tooth surface loss under acidic erosion and tooth brushing abrasion in situ. Methods: Seventeen healthy participants wore a small upper removable mouth appliance holding two sterilised bovine enamel slabs that were randomly assigned to investigate one of the five test products for 14 days entered a prospective controlled, double-blind, crossover with five phases trial. All participants that signed informed consents and fulfilled the inclusion criteria were enrolled in the study. The five treatment groups were: group A (Elmex® erosion protection toothpaste ® (1400 ppm F as AmF/NaF and 3500 Sn2); group B (non-fluoride toothpaste as a ® control); group C (Sensodyne Pronamel 1450 ppm F as NaF/5%KNO3 toothpaste); group D (.Sensodyne Pronamel® 1450 ppm F as NaF toothpaste plus Pronamel® v mouth wash as 450 ppm F NaF) and group E (Elmex® erosion protection toothpaste ® (1400 ppm F as AmF/NaF and 3500 Sn2) with Elmex erosion protection dental rinse as AmF and NaF 500 ppm F plus stannous chloride 800 stannous (Sn2). The enamel slabs were dipped extra-orally in 0.3% citric acid solution at predetermined times for 2 minutes/ five times daily. In addition the enamel slabs were brushed for 1min extra- orally then 1 min intra-orally using the standard toothpaste/natural saliva slurry and 10 ml mouthrinse was used for 60s twice per day (10 ml x2). Enamel loss was determined by surface profilometry (Proscan 2000, Scantron, UK). Results: Treatment with therapeutic products Elmex® erosion protection toothpaste, Sensodyne Pronamel® toothpaste, Sensodyne Pronamel® toothpaste plus Sensodyne Pronamel® mouthrinse and Elmex® protection erosion toothpaste