Due to practical difficulties in quantifying fluoride exposure, the ability of various biomarkers to predict it has been investigated previously. However, the results are inadequate for validation of their application and usefulness. This study aimed to investigate the association between contemporary/recent biomarkers of fluoride exposure and total daily fluoride intake (TDFI) of children with large differences in fluoride exposure through drinking water. TDFI was assessed in 61 healthy 4 to 5 years old children who provided at least one biomarker sample; 32 lived in a low-fluoride area (0.04 mgF/L) and 29 lived in a high-fluoride area (3.05 mgF/L). Validated questionnaires were administered to evaluate fluoride intake from diets (including water) and toothpaste ingestion. Daily urinary fluoride excretion (UFE) as well as fluoride concentrations in plasma, fasting whole saliva, hair and nails (toenails/fingernails) were evaluated and related to total fluoride exposure. TDFI, UFE and fluoride concentration of biomarkers were statistically significantly higher in the high-fluoride area than in the low-fluoride area. There was a strong statistically significant positive correlation between TDFI and: UFE (ρ=0.756, p<0.001); plasma fluoride concentration (ρ=0.770, p<0.001); toenail fluoride concentration (ρ=0.604, p<0.001). The statistically significant positive correlation between TDFI and fingernail fluoride concentration (ρ=470, p<0.001) as well as between TDFI and fasting whole saliva fluoride concentration (ρ=0.453, p=0.001) was moderate, whereas it was weak between TDFI and hair fluoride concentration (ρ=0.306, p=0.027). In conclusion, the current study confirmed the suitability of 24h urine samples for estimating F exposure in children. The strong correlations between TDFI and F in plasma and toenails also suggest these biomarkers may be considered for health risk assessments of fluoride in children who are susceptible to development of dental fluorosis. Sample preparation and analysis Urine samples collected over the 24h period were pooled for each participant, and the total volume was measured. Saliva samples were centrifuged at 1000 rpm (MSE Harrier 18/80, UK) for 2 min to separate any food debris [Martínez-Mier et al., 2011]. Each blood sample was centrifuged for 10 min at 1500 rpm and the plasma collected. Nail samples from each participant (finger-and toenails , separately) were weighed, and then surface contamination from the nail clippings was removed by sonication in distilled water for 15 min [Whitford, 2005]. The cleaned samples were dried at 95ºC and weighed again. Food samples were weighed and then homogenised using an industrial blender (Cookwork BL9292, UK). Water and urine samples were analysed directly using a F ion selective electrode (ISE, 720A series; Orion Research Inc., Boston, MA, USA) after addition of TISAB III [Martínez-Mier et al., 2011]. Food, toothpaste, plasma, whole saliva, hair and nail samples were analysed after overnight hexamethyldisiloxane-acid diffusion [Martínez-Mier et a...