Current guidance for fluoride intake – is it appropriate?

Marília Afonso Rabelo Buzalaf

Professor of Biochemistry and CariologyDepartment of Biological Sciences, Bauru School of Dentistry, University of São Paulo, Brazil

1. Introduction

Fluorides have been extensively employed to control dental caries since the first half of the 1900s.

Since the classical epidemiological studies by H. Trendley Dean, it was known that there should be an

optimum level of exposure to fluoride that would be able to provide the maximum protection against caries,

with minimum dental fluorosis 1. Up to the 1980s, it was believed that fluoride exerted its protective effect

against caries through a ‘systemic’ action, after being absorbed and taken up in the apatite crystals of the

forming teeth. According to this concept, it was unavoidable to ingest fluoride in order to have protection

against caries. In this sense, the occurrence of dental fluorosis was considered a necessary risk in order that

the cariostatic benefits of fluoride could be achieved 2. In the 1980s and 1990s, a paradigm shift was

proposed regarding the mechanisms of action of fluorides against caries3. It was observed that the amount

of fluoride that could be taken up in the apatite was not enough to provide significant protection against

acid dissolution 4. On the other hand, the presence of low levels of fluoride in the oral fluids surrounding

the enamel was effective to inhibit demineralization and enhance remineralization. The concept that

fluoride interferes in the dynamics of caries formation mainly when it is constantly present at low

concentrations in the fluid phases of the oral environment became widely accepted (‘topical’ action) 5-9 and

made it possible to obtain substantial caries protection without significant ingestion of fluorides 10. Having

this in mind and being aware of the increase in the prevalence of dental fluorosis in both fluoridated and in

non-fluoridated areas 11-13, researchers all over the world turned their attention toward controlling the

amount of fluoride intake 10. It is important to point out that even the methods of fluoride delivery

classically classified as ‘systemic’, such as water and salt fluoridation, can have a ‘topical’ effect against

caries when fluoride is in contact with the teeth. In addition, after the ingestion of fluoride, this ion can

return to the oral cavity through saliva and crevicular fluid and then exert its anticariogenic action, by

interfering in de- and remineralisation processes 2. In fact, most of the anticaries effect of ‘systemic’

sources of fluoride, such as fluoridated water or salt, is nowadays attributed to the ‘topical’ contact with the

teeth while these vehicles are in the oral cavity or when fluoride is redistributed to the oral environment

though saliva 2. This does not mean that fluoride does not possess a pre-eruptive effect on caries control.

The pre-eruptive effect has been described for decades based on data from epidemiological studies, such as

the classical Tiel-Culemborg fluoridation study in The Netherlands 14 (for review see Murray et al. 15).

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More recent cohort studies have reported that the pre-eruptive effect of fluoridated drinking water is

important for caries prevention especially in pit and surface surfaces of permanent molars, since these areas

are of difficult access to ‘topical’ fluoride 16-18.

In the present review, we will discuss the appropriateness of the current guidance for fluoride

intake, considering the windows of susceptibility to caries and fluorosis, the modern trends of fluoride

intake from multiple sources, individual variations in fluoride metabolism, as well as recent

epidemiological data. Based on the available evidence, further research that is required to provide

additional support for future decisions on guidance in this area will be suggested.

2. Etiology and window of susceptibility to the development of caries and fluorosis

When we try to use fluorides to obtain the best balance in the protection from caries while limiting

the risk of dental fluorosis, it is necessary to have in mind the etiology of these lesions and the windows of

susceptibility to both of them.

Caries is a multifactorial disease caused by the simultaneous interplay of different factors – dietary

sugars, dental biofilm and the host – within the context of the oral environment 2,19. Whenever there is an

unfavorable balance leading to increased periods of demineralization and reduced periods of

remineralization, an initial caries lesion might form. This can take place from the crib to the grave,

provided that the risk factors exceed the protective factors 9. There are some periods in life when the

unfavorable balance is more likely to occur, such as in the primary dentition in the preschool years, in the

mixed dentition in early school years, in the permanent dentition of adolescents at high school, young

adults at college or along adulthood 20.

Dental fluorosis is caused by excessive fluoride intake during tooth formation. Considering that

fluorotic changes in teeth cannot be reversed but may be easily prevented by controlling fluoride intake

during the critical period of tooth formation, the identification of periods during which fluoride intake most

strongly results in enamel fluorosis is of great importance and subject of extensive investigation10.

For the whole permanent dentition (excluding the third molars), the window of susceptibility for

fluorosis development has been regarded to be the first 6-8 years of life 21-23. Most of the studies concerning

the window of maximum susceptibility to dental fluorosis development, however, have focused on the

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permanent maxillary central incisors, which are of greatest cosmetic importance. While there is general

agreement that the early maturation stage of enamel development is more critical for fluorosis than the

secretory stage 24-29, the evidence considering the age at which maxillary central incisors are most

susceptible to dental fluorosis is not completely conclusive. Table 1 summarizes the results of studies

focused on this topic 10. They can be divided into two categories: studies involving subjects whose exposure

to fluoride started at different ages during tooth formation 30-37 and those involving subjects that had been

exposed from birth and then had an abrupt reduction in daily fluoride intake 28,38-41. Most of them were

cross-sectional, retrospective, and focused on just one or two sources of fluoride intake. Only one study

(Iowa Fluoride Study), used longitudinal data on individual fluoride intake 36,37. While one study reported

that the first year of life was the most critical period for developing fluorosis in the permanent central

maxillary incisors 33, three studies found the first 3 years critical 37-39 and another one recognized a later

period (between 35 and 42 months) 41, most of the them agreed that the first two years of life are most

important 30-32,34,35. This was also reported in a meta-analysis 42, where it was reported that the duration of

exposure to fluoride during amelogenesis, rather than specific risk periods, would seem to explain the

development of dental fluorosis in the maxillary permanent central incisors. In other words, long periods of

fluoride exposure (>2 out of the first 4 years) led to an odds ratio (OR) of 5.8 (95% CI: 2.8 – 11.9) versus

shorter periods of exposure (<2 out of the first 4 years of life). This is in agreement with a more recent

longitudinal study which concluded that (1) although the first 2 years of life were generally found to be

more important compared with later years, fluoride intake during each individual year (until the fourth year

of life) was associated with fluorosis and (2) subjects with higher levels of fluoride intake (estimated mean

daily ingestion of 0.059 mg per kilogram body weight) during the whole first three years of life had the

highest risk of fluorosis 36. Thus, the development of fluorosis appears to be related not only to the timing

of fluoride intake relative to the periods of tooth formation, but also to the cumulative duration of fluoride

exposure 36,42. From the available evidence, it seems rational to monitor fluoride intake of children during

the first three years of life in order to minimize the risk of developing dental fluorosis of the permanent

maxillary central incisors 10,36,37,42.

In summary, the fluoride intake of importance to dental fluorosis occurs in early childhood while

that of importance to dental caries occurs along the whole life course. This implies that policies aiming at

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reducing the fluoride intake to diminish the risk of dental fluorosis should be targeted to early childhood,

while fluoride exposure should be maintained across life for the control of dental caries 43.

3. ‘Optimal’ fluoride intake: can it be precisely determined?

The ‘optimal’ intake of fluoride (between 0.05 and 0.07 mg fluoride per kilogram body weight)

44 that is still accepted worldwide was in fact empirically established. Its origin comes from the 1940s,

when McClure 45 estimated that the “average daily diet” contained 1.0-1.5 mg of fluoride and this would

provide about 0.05 mg fluoride per kilogram body weight for 1-12-year-old children. Farkas and Farkas

46, in the 1970s, cited various sources that suggested 0.06 mg fluoride per kilogram body weight as

“generally regarded as optimum”. In the 1980s, this range of estimates started being used as a

recommendation for ‘optimal’ fluoride intake 47. However, it is not clear if this level of intake is ‘optimal’

for caries prevention, for fluorosis prevention, or both 10. In addition, some authors consider 0.1 mg

fluoride per kilogram body weight per day to be the exposure level above which fluorosis occurs 48, while

others report the occurrence of dental fluorosis with a daily fluoride intake of less than 0.03 mg fluoride

per kilogram body weight per day 24.

Some prospective studies have attempted to add evidence to the empirically established range of

‘optimal’ fluoride intake. In a small-scale study, Martins et al. 49 evaluated the relationship between

fluoride intake and dental fluorosis in permanent central incisors and first molars of 49 children. When

the children were aged 19-39 months, fluoride intake from diet, dentifrice and the combination of both

was evaluated using the ‘duplicate-plate’ method and ‘simulated toothbrushing’ 50. Six years later, when

the permanent central incisors and first molars of these children had erupted, they were evaluated for

dental fluorosis. Dentifrice was the most contributor for the total fluoride intake, but no association was

found between dental fluorosis in permanent teeth and fluoride intake from diet, dentifrice, or combined.

The study had limitations such as the fact that fluoride intake was measured only once and the absence of

children with severe dental fluorosis 49. The most comprehensive study on the association between

fluoride intake, dental caries and dental fluorosis is the Iowa Fluoride Study (IFS), which is still ongoing.

It is a longitudinal cohort study of children recruited soon after birth from 8 Iowa hospitals during March

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1992 to February 1995 51-57. Initially, 1882 newborns were recruited and their mothers completed baseline

questionnaires between 1992 and 1995. After this, mothers were sent questionnaires on a regular basis (3-

and 4-month intervals from birth to 48 months of age and every 6 months thereafter), which included

detailed information regarding children´s fluoride ingestion from different sources, such as water,

beverages, food products, dietary fluoride supplements and fluoride dentifrice 51,52,54,55. Some years ago,

the authors presented results relating longitudinal fluoride intake of the participants to dental caries and

dental fluorosis. The main aim was to relate longitudinal fluoride intake to optimal oral health (absence of

dental caries and dental fluorosis in the permanent teeth), in order to add scientific evidence to the

“optimal” fluoride intake. Six-hundred and one children were included. Of these, 153 had neither

fluorosis at age 9 or caries experience at age 5 or age 9; 202 had caries but no fluorosis at age 9; 96 had

fluorosis but no caries; and 150 had both. Children with no caries history and no fluorosis at age 9 years

had estimated mean daily fluoride at or below 0.05 mg/kg during different periods of the first 48 months

of life, and this level declined thereafter. Children with caries or fluorosis had slightly lower or higher

fluoride intakes, respectively. These results suggest that the accepted range of 0.05 – 0.07 mg fluoride per

kilogram body weight may not be optimal in preventing fluorosis. However, given that most fluorosis

was mild or very mild, and not of esthetic concern, even at high intake levels, recommendations to limit

daily fluoride intake to less than 0.05 mg fluoride per kilogram body may not be justified. On the other

hand, considering that most caries prevention results from topical fluoride exposure, it does not make

much sense trying to establish what is the ‘optimal’ fluoride ingestion level for caries prevention. It was

disappointing that after conducting this extensive and well designed cohort study the authors had to

conclude that “Given the overlap among caries/fluorosis groups in mean fluoride intake and extreme

variability in individual fluoride intakes, firmly recommending an “optimal” fluoride intake is

problematic” 56. Moreover, the authors agree with Burt and Eklund 58 that “perhaps it is time that the term

optimal fluoride intake be dropped from common usage”. This study also had limitations, since it relied

on parental reports of fluoride use and ingestion; it was conducted in one area of the United States with a

sample that was not representative of any defined population; and there were missing data. In addition,

most of fluorosis was mild or very mild, and not of esthetic concern. On the same way, most of children

with caries had relatively few decayed or filled surfaces. Despite these limitations, it is the best outcome-

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based assessment of the ‘optimal’ fluoride intake available so far. Even so, the overlap among

caries/fluorosis groups in mean fluoride intake and the high variability in individual fluoride intakes for

those caries and fluorosis-free discourage a strict recommendation of an ‘optimal’ fluoride intake,

especially at the individual level. This recommendation, however, seems to be desirable at the population

level to guide programs of community fluoridation. For this purpose, and having in mind the windows of

susceptibility to the development of dental fluorosis and dental caries, maybe it would be helpful to have

different ranges of ‘optimal’ fluoride intake for small children and adults, but additional studies are

required before this can be implemented.

4. Factors that modify the metabolism or effects of fluoride

It is not surprising that the ‘optimal’ range of fluoride intake has not been precisely determined

so far. In fact, this is not an easy task, as many factors modify the metabolism and effects of fluoride in

the organism and alter the relationship between fluoride intake and the risk of developing dental fluorosis,

especially when we consider the ‘optimal’ range of fluoride intake at the individual level 59,60.

4.1. Acid-base disturbances

Many aspects of fluoride metabolism, such as absorption, distribution and renal excretion are pH-

dependent, since the coefficient of permeability of lipid bilayer membranes to hydrogen fluoride (HF) is

one million times higher than that of ionic fluoride 61. This implies that fluoride crosses cell membranes as

HF, going from the more acidic to the more alkaline compartment 59. The kidneys are the major route of

fluoride removal from the body. When the pH of the tubular fluid is lower, higher amounts of HF cross the

tubular epithelium, returning to the systemic circulation 59. Thus, any condition that leads to acidic urine

will increase the retention of fluoride in the organism. This includes diet composition (protein- 62 and

sorghum-rich diets 63,64, certain drugs (ascorbic acid, ammonium chloride, chlorothiazide diuretics,

methenamine mandelate), metabolic and respiratory disorders leading to acidosis 60,65, as well as the altitude

of residence 65. Significantly higher prevalence of dental fluorosis has been observed in communities at

high altitude in comparison to those living at low altitude 66-71. It is believed that hypoxia in high altitude

areas ultimately leads to a decrease in urinary pH, increasing fluoride retention in the body 65.

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4.2. Renal impairment

Considering that the kidneys are the major route of fluoride removal from the body, it could be

expected that renal impairment would increase fluoride retention in the organism, thus augmenting the risk

of dental fluorosis. Intake of fluoride by nephrectomized rats increases plasma fluoride levels 72,73. In

addition, children with renal disease present more severe dental fluorosis than healthy children 74.

4.3. Physical activity

Depending on the balance of several factors, exercise can be associated with either increased or

decreased plasma fluoride levels. Upon prolonged physical activity, production of lactic acid might

promote the diffusion of HF from the extracellular to the intracellular fluids leading to an increase in the

rate of fluoride uptake by bone and other tissues, which would reduce plasma fluoride concentration. On

the other hand, plasma fluoride concentration may increase during exercise because of reduced renal

fluoride excretion. The factors associated with reduction in renal fluoride excretion are vasoconstriction

within the kidneys due to increased sympathetic nervous system activity during exercise, which reduces the

renal blood flow and glomerular filtration rate, and acidification of tubular fluid due to the production of

lactic acid, increasing fluoride reabsorption in the tubules 75. Studies conducted with animals have reported

reduced plasma fluoride levels 65,76 and increased bone fluoride levels in exercised (light exercise) rats

compared with non-exercised ones 76. In one of these studies, the rats were submitted to acute exercise and

exposure to fluoride 65 while in the other one fluoride and exercise were administered on a chronic basis

(during 30 days) 76. Information available for humans is limited to a small-scale pilot study that investigated

evaluated urinary fluoride excretion and plasma fluoride concentration in nine young adults undergoing

acute exercise with different intensities following ingestion of 1 mg fluoride. Contrarily to what had been

reported to occur in animal studies, it was observed a trend of a rise in plasma fluoride concentration and

decline in rate of fluoride renal clearance with increasing exercise intensity 77. It seems that the intensity of

the exercise and its nature (acute or chronic), as well as the doses of fluoride administered are important

factors that influence the effect of exercise on fluoride balance in the organism. Additional human studies

involving larger sample size and taking these variables into account are necessary to provide more evidence

on this important matter.

4.4. Nutritional status

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The evidence for the association between malnutrition and dental fluorosis is controversial and

must be interpreted with caution. A fasting child may absorb fluoride more quickly than a well-fed child

since in an empty stomach complexes of fluoride do not exist. On the other hand a malnourished child

presents slower bone growth, which is expected to reduce the fluoride deposition along time 59.

A study conducted in Saudi Arabia showed a significant relationship between water fluoride

concentration, socioeconomic status and prevalence of diffuse enamel lesions (using the DDE index) 78.

However, the DDE index includes dental fluorosis amongst the enamel defects recorded but it is not

specific for evaluating dental fluorosis 79. A study conducted with Tanzanian children also suggested a

relationship between malnutrition and dental fluorosis. However the high prevalence of dental fluorosis

was correlated with previous information on nutrition, but direct comparison between children with or

without malnutrition regarding the prevalence of dental fluorosis was not made 69. A recent study found a

significant association between dental fluorosis (TF ≥4) and low height-for-age in Mexican children. The

children lived in communities with 0.56, 0.70 or 1.60 mg/L fluoride in the drinking water and were also

exposed to fluoridated salt. The association between malnutrition and dental fluorosis was only found for

those living in the area with the highest fluoride concentration in the drinking water 80. In a study conducted

with Brazilian children, a relationship between malnutrition (assessed by height-for-age and weight-for-

age) and dental fluorosis (TF index). The children lived in rural areas containing three different ranges of

fluoride concentrations in the drinking water (below 0.7 mg/L, between 0.7 and 1.0 mg/L and above 1.0

mg/L) 81. In the Mexican study, however, the association between malnutrition and dental fluorosis was

only found for the children living in the area containing 1.6 mg/L fluoride in the drinking water and also

exposed to fluoridated salt 80. These children had a higher exposure to fluoride than the Brazilian children

81, since they were exposed to fluoride both from water and salt. Thus, it is possible that a high intake of

fluoride is necessary in order that an association between dental fluorosis and malnutrition can be

identified.

In view of the contradictory study of the studies that evaluate the relationship between

malnutrition and dental fluorosis, future studies on this topic are necessary. These studies should involve a

longitudinal design, where nutritional status, dietary habits and fluoride intake are assessed during the

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period of tooth formation. It is an important topic especially for developing countries, where malnutrition

and dental fluorosis are prevalent and fluoride-containing products are used to control dental caries 59.

4.5. Diet composition

As mentioned above, any dietary component that promotes acidification of urine will intensify

the fluoride retention in the organism and could be associated with increased risk of dental fluorosis. This is

the case of protein- 62 and sorghum-rich diets 63,64. On the other hand, dietary components that lead to

alkalinisation of urine will increase fluoride excretion and are expected to reduce the likelihood of dental

fluorosis. Among these are tamarind 82,83 and vegetarian diet 62.

Calcium-rich diets have also been associated with lower prevalence of dental fluorosis. This

might be due to the fact that calcium can reduce the extent of fluoride absorption by forming insoluble

complexes with fluoride 84. In addition, during amelogenesis, high calcium concentrations can enhance

amelogenin secretion into the enamel space, thereby increasing the local buffering capacity at the

mineralization front thus fighting the deleterious effect of fluoride on the generation of excess protons 85. In

an endemic area of fluorosis in China, the prevalence of dental fluorosis of the milk-drinking and non-milk-

drinking groups was 7.2 and 37.5%, respectively 86. Low calcium concentrations in the drinking water were

inversely related to the prevalence of dental fluorosis in endemic areas of India 87,88. Some authors have

reported an improvement in dental fluorosis in children living in endemic areas of fluorosis in India after

daily supplementation with ascorbic acid, calcium and vitamin D3 89,90. These findings have led to the

suggestion that calcium supplementation should be implemented in areas with endemic fluorosis 88.

However, more evidence is required before this measure can be adopted. Further well-designed studies

should be conducted to determine the effect of calcium alone in communities with similar background

exposure to fluoride.

4.6. Genetic factors

The possibility of genetic predisposition to dental fluorosis was raised in epidemiological studies

that found marked variations in dental fluorosis prevalence in subjects from areas with comparable levels of

fluoride intake 69,91. This possibility gained strength when a study done with 12 inbred mice strains revealed

different susceptibilities to dental fluorosis among the strains. It was observed, both by clinical examination

and by quantitative light induced fluorescence, that the A/J strain was highly affected by dental fluorosis,

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while the 129P3/J strain was remarkably unaffected, even after exposure to water containing 50 mg/L

fluoride 92. It was hypothesized that the different susceptibility to dental fluorosis between these two mice

strains could be due to differences in fluoride metabolism. The resistant strain (129P3/J) was expected to

excrete more fluoride, which in turn would decrease the susceptibility to dental fluorosis. Thus, a metabolic

study was designed to test this hypothesis. In the metabolic study, it was observed that the susceptible

animals (A/J) ingested a significantly higher amount of water when compared with their 129P3/J

counterparts 93. This was later on attributed to an increased expression of Alpha-aminoadipic semialdehyde

dehidrogenase (α-AASA dehydrogenase) in the latter 94. This enzyme metabolyzes irreversibly betaine

aldehyde to betaine, which is the most effective osmoprotectant accumulated by eukariotic organisms to

cope with osmotic stress. This can explain the lower volume of water consistently ingested by the 129P3/J

mice throughout the metabolic study 93. In the metabolic study, in order to equalize the fluoride intake

between the two strains, the fluoride concentrations in the drinking water given to the A/J mice

(susceptible) were reduced, which led to similar amounts of fluoride intake between the two strains. After

controlling for fluoride intake, surprisingly, the resistant strain (129P3/J) excreted a significantly lower

amount of fluoride in urine when compared with the susceptible strain (A/J). Consequently, the resistant

129P3/J mice had significantly higher plasma and bone fluoride concentrations and even so, they were not

affected by dental fluorosis93.

Histological examination of maturing enamel of A/J and 129P3/J mice revealed that exposure to

fluoride increased the accumulation of amelogenins in the maturing enamel of A/J mice, but not of the

129P3/J mice 95. In attempt to identify the possible genes involved in the resistance/susceptibility to dental

fluorosis, quantitative trait locus (QTL) detection mapping was conducted. Dental fluorosis-associated

QTLs were identified on mouse chromosomes 2 and 11 95,96.

Recently, the profile of protein expression in secretory and maturation-stage enamel of

susceptible (A/J) and resistant (129P3/j) mice exposed to fluoride (50 mg/L in the drinking water) or not

was examined, in attempt to identify proteins related to susceptibility/resistance to dental fluorosis 97.

Amelogenin was exclusively identified in the maturation-stage of the susceptible mice (A/J) treated with

fluoride, in agreement with a previous study 95. Another structural protein that was identified exclusively in

the maturation-stage of the susceptible mice (A/J) treated with fluoride was type I collagen (COL1A1) 97.

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The possible relationship of COL1A1 with increased susceptibility to dental fluorosis cannot be ruled out

for two reasons: 1) Epidemiologic data support an association between polymorphisms in the gene of

COL1A2 and dental fluorosis in populations exposed to high fluoride levels in the drinking water,

suggesting the possibility of a gene-environment interaction 98 and 2) In mice, Col1a1 gene is located in

chromosome 11 within the QTL interval associated with dental fluorosis 96. Pigment epithelium-derived

factor, also known as Serpinf1, was also identified exclusively in the maturation-stage of susceptible mice

(A/J) treated with fluoride. Serpinf1 is an inhibitor of serine protease and its gene is also located in

chromosome 11, near the QTL interval associated with dental fluorosis 96. It is important to mention that

Kallikrein 4 (KLK4) is a serine protease actively involved in the degradation of the enamel matrix proteins

during the maturation-stage 99. Vimentin, a protein whose gene resides in chromosome 2 and is involved in

the stabilization of type 1 collagen was also identified exclusively in the maturation-stage of susceptible

mice (A/J) treated with fluoride 97. Considering that the genes of these proteins differentially expressed

between the two strains are all located in chromosomes 2 and 11, where QTLs associated with dental

fluorosis have been identified 95,96, the existence of polymorphisms associated with dental fluorosis

development should be investigated initially in these mice and then in humans.

In the last years, several studies conducted in areas of endemic fluorosis in China investigated the

relationship between dental fluorosis and gene polymorphisms in humans (Table 2). Significant

associations between Col1A2 98, estrogen receptor 100, myeloperoxidase 101 and calcitonin receptor 102 gene

polymorphisms and dental fluorosis were reported. These studies, however, had modest sample sizes and

should be replicated in different populations with larger sample sizes.

5. Sources of fluoride intake

The most important risk factor for fluorosis is the total amount of fluoride consumed from all

sources during the critical period of tooth development 10. Reports of increased prevalence of mostly mild

but also some moderate to severe dental fluorosis have prompted, all over the world, investigations on the

fluoride concentrations of potential sources as well as on the fluoride intake from all sources, mainly in

children at the age of risk for fluorosis development 50,53,55,103-161. Case-control studies, cohort studies and

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randomized clinical trials whose results were compiled in systematic reviews identified four major risk

factors for dental fluorosis: fluoridated drinking water 162-165, fluoride supplements 166, fluoride toothpaste

167-169 and infant formulas 163,170. These major sources of fluoride intake, as well as recommendations on

how to reduce fluoride intake from them, will be discussed below.

5.1. Fluoridated drinking water

Community drinking water fluoridation is included among the top ten greatest public health

achievements in the world in the last century 171. Despite several fluoride-containing products are nowadays

being used, water fluoridation is still the most equitable and cost-effective method of delivering fluoride to

all members of most communities, regardless of income level age, educational attainment or. A recent

systematic review on economic evaluation of community water fluoridation attested that the economic

benefit of this measure exceeds the intervention cost. It was estimated that per capita annual intervention

cost ranges from $0.11 to $4.92 for communities with at least 1,000 population, while the per capita annual

benefit ranges from $5.49 to $93.19. The benefit-cost ratios ranged from 1.12:1 to 135:1 and were

positively associated with community population size 172. Additionally, there is evidence that water

fluoridation reduces the oral health gap between social classes 164, despite this has been questioned in a

recent systematic review 165.

A systematic review on the safety and efficacy of water fluoridation published in 2000 included

214 studies classified as low to moderate quality and reported that water fluoridation was associated with

an increased proportion of children without caries and a reduction in the number of teeth affected by caries.

The range of mean differences in the proportion of children without caries was -5% to 64% (median

14.6%), with a mean in dmft/DMFT of 0.5 to 4.4 (median 2.25) 162,165. A recent systematic review released

by the Cochrane group used more restrict inclusion criteria. Only prospective studies with a concurrent

control that compared at least two populations – receiving or not fluoridated water – with outcome(s)

evaluated at at least two points in time were included, which resulted in the inclusion of 155 studies with

only 107 providing data for quantitative synthesis. A reduction in dmft of 1.81 (95% CI 1.31 to 2.31) and in

DMFT of 1.16 (95% CI 0.72 to 1.61) was reported, which translates to 35% and 26% reductions in dmft

and DMFT, respectively, compared to the median control group mean values. The increases in the

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percentage of caries free children were 15% (95% CI 11 to 19%) in deciduous dentition and 14% (95% CI

5% to 23%) in permanent dentition. The majority of studies (71%) were conducted prior to 1975 (before

the widespread use of fluoridated dentifrice) and with children, since no studies conducted with adults met

the inclusion criteria 165. However, this systematic review has been criticized for many reasons 173. First, the

nature of water fluoridation programs is quite different from RCTs, mainly because of practical, ethical and

financial factors. Evaluation of water fluoridation efficacy should take these differences into account and

use appropriate inclusion criteria, such as inclusion of single cross-sectional surveys of fluoridated and non-

fluoridated groups with control for confounding factors. Most of the recent studies were of this design and

were excluded from the review. Second, reports by local authorities or Government agencies that present

robust data, but are not published in academic journals, could have been included. Third, the effect of water

fluoridation on the prevalence of fluorosis should have been isolated from the confounding effect of other

fluorides, especially dentifrices. Fourth, it is very important when interpreting evidence from trials that the

lack of evidence (or the existence of poor quality evidence) is not confused with an absence of effect 173.

Since the classic epidemiological studies by Dean, in the 1940s, it is known that about 10% of

children in optimally fluoridated areas (around 1.0 ppm) are affected by mild or very mild fluorosis of the

permanent teeth, and less than 1% in low-fluoride areas 1. Systematic reviews, however, have estimated

that, at a fluoride level of 0.7-1.0 ppm in the drinking water, the prevalence of any fluorosis is around 40-

48% and of aesthetically concerning dental fluorosis is around 12.0-12.5% 162,165. The increased prevalence

of dental fluorosis found more recently all over the world indicates that some young children are ingesting

fluoride from sources other than drinking water. In fact, a review that compiled results of fluoride intake

from different sources indicated that most of the fluoride intake of young children comes from the use (and

ingestion) of fluoridated dentifrice 174.

The U.S. Department of Health and Human Services set up a Panel on Community Water

Fluoridation that recently proposed a new recommendation on water fluoride levels, that is 0.7 ppm

fluoride for the entire nation, and replaces the 1962 U.S. Public Health Service Drinking Water Standards

which were based on ambient air temperature of geographic areas and ranged from 0.7-1.2 ppm fluoride.

The aim of this measure is to maintain caries prevention benefits while reducing the risk of dental fluorosis.

This guidance is based on several considerations that include: a) scientific evidence related to effectiveness

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of water fluoridation on caries prevention and control across all age groups; b) fluoride in drinking water as

one of several available fluoride sources; c) trends in the prevalence and severity of dental fluorosis; d)

current evidence that fluid intake in children does not increase with increases in ambient air temperature

due to augmented use of air conditioning and more sedentary lifestyles 175. This measure has been object of

criticism, since more evidence should have taken into account before its implementation. In addition, it is

possible that water fluoridation in other countries is affected by this decision 43. One aspect that should have

been taken into account before this change is the different window of susceptibility to caries (across the

whole life course) and dental fluorosis (early childhood). Thus, actions to reduce fluorosis should be

targeted in controlling fluoride intake in early childhood, and fluoride exposure across life should be

maintained for the prevention of dental caries. Another aspect is that most dental fluorosis is mild, a

condition not considered as a public health problem. Contrarily, people react to low severity fluorosis in a

positive manner, due to the desire for whiter teeth that is widely disseminated among the young people

nowadays 176. On the other hand, caries is a disease with well-established sequela, great impact among oral

diseases and great cost for its treatment 177. One of the reasons claimed by the Panel to support the change

was the current evidence of fluid intake of children across outdoor air temperature. However, the more

recent studies employed methodologies (24-h diet recall focused on individual analysis using linear

regression) that differ from the classic study by Galagan & Vermilion (5-day observational period repeated

across the seasons) 178. Thus, it is necessary that additional studies compare these two study designs.

Moreover, there has been increased consumption of bottled water 179 and soda and manufacturers of soda

have moved to use distilled water, which reduces potential fluoride exposure from fluoridated tap water 43.

It should be noted that the increase in the prevalence of any fluorosis all over the world across the

1980s and 1990s coincides with the introduction of many different fluoridation vehicles. Any measure

targeted to reduce the prevalence and severity of dental fluorosis should precisely know the individual

contribution of all fluoridated vehicles for the total daily fluoride intake of children in the risk for dental

fluorosis. A study estimated the total daily fluoride intake from different constituents of the diet and from

dentifrice by 1- 3-year-old children living in an optimally fluoridated area. Standard fluoride concentration

dentifrice alone was responsible for, on average, 81.5% of the daily fluoride intake, while among the

constituents of the diet, water and reconstituted milk were the most important contributors and were

15

responsible for about 60% of the total contribution of the diet 115. For 4-6-year-old children living in the

same community, however, the impact of fluoride ingested from dentifrice was less and water alone

provided a mean of 34% of the estimated daily fluoride from the diet, which corresponds to about 0.014 mg

fluoride per kilogram body weight 116,117. Research conducted in the US reports similar findings. In a

nonfluoridated area, 65% and 34% of fluorosis cases were related to the use of fluoride supplements and

tooth brushing, respectively. As for children grown up in fluoridated areas, 68% of fluorosis cases were

attributed to the use of dentifrice during the first year of life. 180 These results indicate that measures to

reduce the risk of fluorosis should be targeted not on reducing fluoride concentration of the drinking water,

but at reducing fluoride concentration and rationalizing use other vehicles that have a more prominent

association with dental fluorosis, such as dentifrices.

Since fluoride present in water contributes only a small portion of intake from the dietary

constituents, fluoridated water probably has its greatest impact on dental fluorosis prevalence indirectly,

through being used in the reconstitution of infant formulas and in the processing of other children’s foods

and beverages 44. Taking into account the low risks and great benefits of public water fluoridation, as well

as the levels of prevalence and especially severity of dental fluorosis found today, this measure must be

maintained in the areas where it already exists and extended to the areas where it is feasible to implement

water fluoridation.

In order to minimize the possible impact of water fluoridation on dental fluorosis, some measures

should be taken. One of them is external monitoring of water fluoridation by an independent assessor. This

measure has been shown to be successful in improving the consistency of fluoridation 181 and ideally should

be implemented wherever there is adjusted fluoridation but, at least, in the communities where fluctuations

in water fluoride levels commonly occur 182.

Another important advice is to reconstitute infant formula given for infants and small children

with water containing less than 0.5 ppm fluoride 10. A recent meta-analysis found that a 1.0 ppm increase

in the fluoride level in the water supply is associated with a 67% increased odds ratio for dental fluorosis

associated with infant formula 163. Thus, bottled water with relatively low fluoride content can be used

instead of fluoridated water from the public supply 120,124,183. Several brands of bottled water commercially

available have low fluoride content and should be adequate for this purpose 103,179,184-189. However, one

16

difficulty is that in many cases fluoride concentrations are not stated or are stated inaccurately on the labels,

and unexpectedly high fluoride concentrations can be found 179,187. This reinforces the need for global

labeling of fluoride levels in bottled water and rigorous surveillance by the competent public health

authorities 10.

5.2. Fluoride dentifrice

During many decades, fluoridated water was recognized as the main risk factor for dental fluorosis

as a consequence of the studies by Dean and little use of fluoride from other sources 1. However, increases

in the prevalence of dental fluorosis both in non-fluoridated areas than in fluoridated areas 190 turned the

attention of the researchers to mapping the impact of other potential sources of fluoride ingestion. Among

these, fluoride dentifrices were identified as a potential risk factor for dental fluorosis, since an inverse

relationship can be observed between the age of the child and the mean percentage ingestion of dentifrice

191,192. In fact, there is an extensive relationship between the use of fluoridated dentifrice and the risk of

developing dental fluorosis. A review compiled data for estimated total fluoride intake of children living in

different locations 174. It was noted that dentifrice was usually the main contributor for young children.

Thus, dentifrice is an important source of fluoride during the critical period of tooth development. A

plethora of studies of distinct designs, conducted in different countries, both in fluoridated and non-

fluoridated communities, investigated the association between the use of fluoride dentifrice and the

prevalence or severity of dental fluorosis. A positive association was found in most of these studies, mainly

related to the early use of fluoride toothpaste (before age 24 months), regardless of the community

fluoridation status (for review see Table 2 in Buzalaf & Levy 10). A Cochrane systematic review and meta-

analysis 167 compiled the results of 25 studies published between 1988 and 2006 and investigated the

relationship between the use of fluoride dentifrices and dental fluorosis. Among the 25 studies included,

only one RCT was considered at low risk of bias 193. The main findings were: (a) a significant reduction in

the risk of dental fluorosis was found if toothbrushing with fluoride dentifrice did not start until the age of

12 months, but the evidence for starting toothbrushing with fluoride dentifrice before the age of 24 months

was inconsistent (data from case-control and cross-sectional studies); (b) no significant association was

found between frequency of toothbrushing or amount of dentifrice used and fluorosis (data from cross-

17

sectional surveys); and (c) using dentifrice with a higher concentration of fluoride increased the risk of

dental fluorosis (data from two RCTs; evidence from cross-sectional studies was inconsistent). Based on

the available evidence, the authors concluded that decisions involving the use of topical fluorides

(including dentifrices) should balance their benefits in caries prevention and the risk of causing dental

fluorosis. They noted: “if the risk of fluorosis is of concern, the fluoride level of toothpaste for young

children is recommended to be lower than 1,000 ppm” 167. In fact, risk-benefit considerations are critical for

counselling parents on the ideal dentifrice to be indicated to their children. A recent systematic review and

meta-analysis of 83 independent trials concluded that only dentifrices containing 1,000 ppm fluoride or

more have been proven to be beneficial for preventing caries in children and adolescents 194. However, for

the deciduous dentition (age related with the development of dental fluorosis), uncertainty regarding the

effectiveness of low-fluoride dentifrices for preventing caries was reported due to the lack of trials and the

evidence is inconclusive 194. Another systematic review and meta-analysis released in 2013 evaluated the

effects of low and standard fluoride toothpastes on caries in the primary dentition and aesthetically

objectionable fluorosis in the permanent dentition. The authors concluded that low-fluoride toothpastes

significantly increased the risk of caries in primary teeth (RR = 1.13 (1.07-1.20)) and did not significantly

decrease the risk of aesthetically objectionable (moderate to severe) fluorosis in the upper anterior

permanent teeth (RR = 0.32 (0.03-2.97)) 195. However, these results have been criticized 196. Only 5 studies

were included in the meta-analysis. Among them, only one (which was also included in the Cochrane

systematic review 194) showed that children using low-fluoride dentifrice had a significant increase in the

mean caries incidence at tooth level compared to those using standard fluoride toothpaste. In addition, in

this study, children lived in non-fluoridated areas and were at high risk for dental caries. The other 4 studies

included did not find any significant difference between low- and conventional fluoride dentifrices in

preventing caries in the primary teeth. These studies had high variation among the dmfs values. Regarding

dental fluorosis, only two studies, performed in non-fluoridated and non-optimally fluoridated areas were

analysed, with mean RR of 0.32 and individual RR very discrepant (0.09 and 0.8). These studies presented

different age of the patients at baseline and time of follow-up. Thus, no final conclusion can be drawn,

especially considering that there was not evaluation of the use of low-fluoride toothpaste in a fluoridated

area. Furthermore, the use of fluoride dentifrice is more related to mild fluorosis and in a lesser extent to

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moderate to severe fluorosis, which was object of analysis in the Santos et al. 195 study. Many other

limitations of the Santos et al. 195 study can also be pointed out, such as: the selected studies presented bias,

such as high rates of losses at follow-up, no report of baseline caries balance and the tested formulations

had different formulations regarding fluoridated salt, concentration and pH 196. In fact, limited evidence

demonstrates that for children younger than 6 years, fluoride toothpaste use is effective in caries control.

However, ingesting pea-sized amounts or more can lead to mild dental fluorosis 169. In summary, there is no

evidence supporting the use of low-fluoride toothpastes for caries prevention in the primary dentition

because of the low number of RCTs on children <7 years old. In addition, there is also no evidence that

low-fluoride toothpaste leads to the same risk of dental fluorosis as standard toothpaste, with lack of

information in fluoridated areas 196. These are urgent research needs. While additional research is not

available, it should always be kept in mind that the lack of evidence must not be interpreted as lack of

effect.

It has been recommended that young children use a small amount of dentifrice, in order to reduce

the intake of fluoride. This measure has achieved success in reducing mild to moderate fluorosis 197,198, but

in fact a systematic review did not find an association between the amount of dentifrice used and fluorosis

167. A recent study showed that the low-fluoride dentifrice applied using transversal technique led to a

significantly higher AUC of salivary fluoride than the conventional dentifrice in a pea-sized amount 199.

These results suggested that the use of a small amount of conventional dentifrice may not be as effective as

the use of a low-fluoride formulation applied using the transversal technique. The authors quoted:

“Therefore, the current recommendation of very small quantities of a conventional toothpaste should be re-

evaluated, as it is not based on sound scientific evidence that considers intraoral fluoride retention or any

other relevant parameter impacting clinical efficacy. As the controversies surrounding low-fluoride

toothpastes have not yet been fully addressed, this issue should be constantly discussed and evaluated by

authorities to determine the best possible therapy to patients. In this sense, while more consistent evidence

is not available on the recommendations of toothpastes to children, professionals, parents and caregivers

should reserve the right to choose the treatment that better suits the children’s need, based on risks,

benefits, costs, and personal preferences. ” 199 In order to provide more evidence on this important topic, in

situ and clinical studies should evaluate the anticaries efficacy of brushing with a low-fluoride dentifrice

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(550 ppm) applied using the transversal technique compared with a conventional toothpaste (1,100 ppm) in

a pea-sized amount.

When we think about the ideal dentifrice that should be indicated to obtain the best risk/benefit

relationship, there should not be a universal recommendation, i.e., the characteristics of the

individual/population should be respected. Both in non-fluoridated 200 and in fluoridated areas 201 the low-

fluoride dentifrice (500-550 ppm) has similar efficacy as the conventional one (1000-1100 ppm fluoride)

for caries prevention in caries-inactive 2-4-year children. However, in caries-active children the

conventional dentifrice significantly reduces the progression and net increment of initial caries in

comparison with the low-fluoride dentifrice 200,201. Thus, the caries risk of the child is in important factor to

be taken into account to indicate the type of dentifrice to be used.

Due to the uncertainties in the literature regarding the anticaries potential of low-fluoride

dentifrices, researchers have proposed alterations in their formulations in order that their anticaries

potential can be similar to the one of the conventional dentifrices, such as pH reduction 201 or phosphates

supplementation 202. Randomized clinical trials evaluating these low-fluoride (500-550 ppm) dentifrices

revealed anticaries efficacy similar to the one of the conventional dentifrices (1000-1100 ppm fluoride)

201,202. This indicates that they are the ideal alternative for children, taking into account risk-benefit

considerations. In the Brazilian market, a low-fluoride liquid acidic dentifrice is available on the market.

Clinical trials revealed good results for the prevention of caries and also fluorosis 201, since long-term use of

this dentifrice by young children results in nails fluoride concentrations lower than 2 mg/Kg, which has

been associated with reduced risk of developing dental fluorosis in the permanent maxillary central incisors

203. The tested formulation could be an alternative to standard fluoride concentration dentifrice in order to

avoid dental fluorosis in young children, but additional clinical trials are necessary to provide unequivocal

evidence on this matter. Also further work should be done in an attempt to enhance the anti-caries efficacy

of low-fluoride dentifrices in order to further maximize benefits and minimize risk of accidental ingestion.

In conclusion, based on the available evidence regarding the risks of caries and dental fluorosis, it

seems reasonable to recommend low-fluoride (500-550 ppm) dentifrices for young children who are at risk

of developing dental fluorosis in the permanent maxillary central incisors (less than 3 years of age) but

have low-caries risk, especially if they live in a fluoridated area. In all other cases, dentifrices containing at

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least 1,000 ppm fluoride should be used. Although to date there is not unequivocal evidence supporting the

association between the amount of dentifrice used and dental fluorosis 167, it seems rational to recommend

the use of a small amount of dentifrice by young children, which can be easily achieved using the

‘transverse’ 204 or ‘drop’ 205 techniques. It is equally important that young children brush under adult

supervision and be instructed to expectorate the foam after toothbrushing as much as possible 10.

5.3. Dietary Fluoride Supplements

A plethora of studies that evaluated the association between different sources of fluoride intake

and dental fluorosis, include dietary fluoride supplements, besides fluoride dentifrices, among the main risk

factors for dental fluorosis (for review, see Table 2 in Buzalaf & Levy 10).

Dietary fluoride supplements were developed to prevent dental caries in children living in

fluoride-deficient areas. The recommended daily dose was based on the age of the child and fluoride

concentration in the drinking water. In 1999, a systematic review of studies evaluating the association

between the use of fluoride supplements by children living in non-fluoridated areas and dental fluorosis

was done 206. By analyzing studies conducted between 1966 and 1997, the authors performed a qualitative

review of 10 cross-sectional/case-control studies and found a strong association between the use of fluoride

supplements and dental fluorosis. The estimated odds ratio of dental fluorosis in children living in non-

fluoridated areas who had regularly used supplements during the first six years of life when compared with

non-users was about 2.5 206. This systematic review was updated with the inclusion of another four studies

in the meta-analysis, which confirmed the positive association between the use of supplements and the

occurrence of dental fluorosis. The odds ratio for dental fluorosis increased by 84% for each year of use of

fluoride supplements between the ages of younger than 6 months and 7 years, but the first 3 years of life

were considered more important 166. Most cases of dental fluorosis associated to the use of fluoride

supplements were graded as mild, with little probability of causing social impact 166,207. The effectiveness

of fluoride supplements for preventing caries was also evaluated. The authors found a weak, inconsistent

evidence that fluoride supplements are effective at preventing caries in the primary dentition. However,

they are able to help prevent caries in the permanent teeth of school-aged children (older than 6 years)

21

when used on a regular basis, primarily due to topical effect 166. This was confirmed in a Cochrane

systematic review that included randomized or quasi-randomized controlled trials with minimum follow-up

of 2 years, comparing fluoride supplements with no fluoride supplement or with other preventive measures

such as topical fluorides in children that were less than 16 years of age at the start. The main outcome was

change in dmfs/DMFS. Eleven studies were included (7,196 children). The use of fluoride supplements

when compared with no fluoride supplement was associated with a 24% (95% CI 16-33%) reduction in

DMFS, but the effect was unclear on deciduous or primary teeth. When compared with the administration

of topical fluorides, no differential effect was observed. The authors commented that 10 trials were rated as

being at unclear risk of bias and one at high risk of bias. Therefore, the provided evidence about the

efficacy of fluoride supplements is weak 208.

The available evidence regarding the associations of supplements with dental caries and dental

fluorosis indicates that consideration of the risk-benefit ratio is necessary when prescribing supplements,

similarly to what has been discussed above for fluoride dentifrices. There is general consensus that fluoride

supplements should not be prescribed in optimally fluoridated areas, for infants less than 6 months of age

nor for children who are at low risk of developing dental caries. Different policies, however, have been

adopted by distinct countries and dental associations regarding the recommendations for the appropriate use

of supplements to prevent caries (for review, see Buzalaf & Levy 10). Considering the available evidence

that fluoride supplements only help prevent caries when regularly used by children older than 6 years of

age, and that their use before this age (but especially during the first 3 years) is associated with dental

fluorosis 166, the view of the a group of European experts, in 1991, that recommend a dose of 0.5 mg/day

fluoride for at-risk individuals from the age of 3 years 209 seems to be the most rational one. However, for

remote/special populations not receiving other fluoride and caries prevention measures, fluoride

supplementation may also be considered 10.

5.4. Infant formulas

Breastfeeding is undoubtfully the best nutrition for infants and young children. Sometimes,

however, it is not feasible. Furthermore, as infants are weaned from breast milk, they are given infant

formula, mainly in the first 4 to 6 months of life before they start receiving solid foods. Commercially

22

prepared infant formulas are available as powder and liquid concentrates that have to be diluted with water

before use, or as ready-to-feed formulations. Several studies have analyzed the fluoride content of infant

formulas worldwide. This varies from negligible to high and is dependent on the fluoride concentration in

the water used to reconstitute the powder or liquid concentrate formulas 103,108,124,153,155,183,210,211. Considering

that infants ingest high volumes of milk per day, they are likely to exceed the upper tolerable limit of

fluoride intake if they are exclusively fed powdered infant formula reconstituted with 0.7-1.0 ppm

fluoridated water. Thus, it has been suggested that the intake of fluoride by infants from formulas is

influenced more by the water used to reconstitute the formula than by the formulas themselves

103,120,183,211,212. Soy-based infant formulas, however, usually have higher fluoride concentrations than milk-

based ones 124,183,210,211,213. It has been reported that substantial consumption of fluoride-rich soy-based infant

formulas, even if they are reconstituted with deionized water, would provide a fluoride intake above the

upper tolerable limit for 1-month-old children 124,183,213.

Due to their intrinsic fluoride content and mainly to the use of fluoridated water to reconstitute

infant formulas, their consumption by infants and young children has been considered among the risk

factors for dental fluorosis. A systematic review evaluated the relationship between use of infant formula

from birth to age 24 months and dental fluorosis 163. The authors compiled the results of 19 studies

including 17,429 subjects with ages ranging from 2 to 17 years. The summary odds ratio from 17 studies

relating infant formula use to dental fluorosis in the permanent dentition was 1.8 (95% CI 1.4-2.3). There

was, however, significant heterogeneity in the magnitude of the odds ratios among the studies, which

indicates that the summary OR must be interpreted with caution. A meta-regression provided weak

evidence that the fluoride in the infant formula resulted in an increased risk of developing dental fluorosis.

In fact, the dental fluorosis risk associated with the use of infant formula depended on the concentration of

fluoride in the water supply. An increase in the dental fluorosis OR of 5% was seen as the fluoride level of

the water supply increased by 0.1 ppm (OR 1.05, 95% CI 1.02-1.09). This means that a 1.0 ppm increase in

the fluoride concentration in the water supply is associated with a 67% increased OR for dental fluorosis

associated with infant formula (OR 1.67, 95% CI 1.18-2.36). The Iowa Fluoride Study revealed that greater

fluoride intakes from reconstituted infant formulas at ages 3-9 months increased the risk of mild dental

fluorosis of the permanent maxillary incisors 214.

23

A panel of experts convened by ADA provided evidence-based clinical recommendations

regarding fluoride intake from reconstituted infant formula (from birth to 12 months) and dental fluorosis.

Based on the systematic review mentioned above 163 and two other clinical studies the panel suggested that

“when dentists advise parents and caregivers of infants who consume powdered or liquid concentrate

formulas as the main source of nutrition, they can suggest the continued use of powdered or liquid

concentrate infant formulas reconstituted with optimally fluoridated drinking water while being cognizant

of the potential risks of enamel fluorosis development. These recommendations are presented as a resource

to be considered in the clinical decision-making process. As part of the evidence-based approach to care,

these clinical recommendations should be integrated with the practitioner´s professional judgement and

the patient´s needs and preferences.” 170 If the parents mention their concern with the occurrence of dental

fluorosis, water containing less than 0.5 ppm fluoride should be recommended for the reconstitution of

infant formulas. Bottled water with low fluoride concentrations could be used for this purpose 124,179,183.

However, fluoride concentrations both in infant formula and bottled water must be correctly displayed on

the products’ labels. Periodical analyses of fluoride concentrations present in infant formula and bottled

water by government or private laboratories could contribute to assure that the fluoride levels are

adequately displayed on the labels.

6. Biomarkers of exposure to fluoride

As mentioned above, estimating fluoride intake, especially for children at the age of risk to

dental fluorosis, is extremely important. However, it is a quite difficult task, due to the multiplicity of

sources of fluoride ingestion. In addition, taking into account that only absorbed fluoride is involved in

the development of dental fluorosis, the monitoring of fluoride absorption rather than fluoride intake, is

much more informative 215. In this context, biomarkers of exposure to fluoride assume great importance.

According to the WHO, “a fluoride biomarker is of value primarily for identifying and monitoring

deficient or excessive intakes of biologically available fluoride” 216. The biomarkers of exposure to

fluoride have been classified as contemporary (that assess present of very recent exposure) (for review,

see Rugg-Gunn et al. 217) or recent and historical (that assess chronic or subchronic exposure to fluoride

(for review see Pessan & Buzalaf 218).

24

Among the contemporary biomarkers of exposure to fluoride, urine has been regarded as the

most useful one 219. Since a known proportion of ingested fluoride is excreted in urine 220, daily urinary

fluoride excretion is a useful biomarker of contemporary fluoride exposure for groups of people, but not

for individuals 217,219. Normal values have been published for different ages 219. Fluoride concentrations in

parotid and submandibular saliva, but not in whole saliva, have also been proposed to be related to

plasma fluoride concentrations (for review, see Rugg-Gunn et al. 217) but to date there are not enough data

to establish a normal range of fluoride concentrations in ductal saliva that would enable the use of this

contemporary biomarker of fluoride exposure 217. In fact, the usefulness of contemporary biomarkers of

exposure to fluoride to predict the risk of dental fluorosis is limited, since these biomarkers reflect very

recent exposure to fluoride (typically in the last 24 hours) and dental fluorosis is related to increased

consumption of fluoride on a chronic basis. In this sense, recent biomarkers of exposure to fluoride are

more useful.

Regarding recent biomarkers, nails seem to be promising since they have many advantages: can

be easily collected in a non-invasive manner, can be stored for long time at room temperature without

degradation and their concentration of fluoride reflects the average level of intake over a protracted time,

which is desirable when the risk of developing dental fluorosis is considered 218. Recently, the use of

fingernail fluoride concentrations at ages 2-7 years was validated as predictor of the risk of developing

dental fluorosis in the permanent dentition. It was observed that children with dental fluorosis had

significantly higher fingernail fluoride concentrations than those without the condition and the

concentrations tended to increase with the severity of fluorosis. With a water fluoride concentration of 2

ppm at ages 2-7 years as a threshold, this biomarker had high sensitivity (0.84) and moderate specificity

(0.53) as a predictor for dental fluorosis. It was suggested that this biomarker might be useful in public

health research, since it has the potential to identify nearly 80% of children at risk of developing dental

fluorosis 203. If there is concern in avoiding dental fluorosis in the permanent maxillary central incisors,

fingernail fluoride concentrations could be assessed periodically during the first 3 or 4 years of life to

guide counselling on sources of fluoride intake.

7. Concluding remarks and research needs

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It is quite difficult to think about a strict recommendation for an ‘optimal’ range of fluoride

intake at the individual level in light of the actual knowledge on: 1) the mechanisms of action of fluoride

to control caries, 2) the mechanisms involved in dental fluorosis development, 3) the distinct factors that

interfere in the metabolism of fluoride, 4) the windows of susceptibility to both dental caries and fluorosis

development. An ‘optimal’ range of fluoride intake is, however, desirable at the population level to guide

programs of community fluoridation. Since caries can be controlled along the whole life course mainly by

the local contact of the teeth with fluoride present in the fluid phases of the oral environment and

fluorosis develops following excessive fluoride intake during the critical period of tooth formation (first

6-8 years of life), we need to define what the term ‘optimal intake’ stands for. Is it to control caries, to

avoid dental fluorosis or both? Considering that the effect of fluoride to control caries occurs mainly due

to its presence in the oral fluids and the role of ingested fluoride is regarded as less important, the term

‘optimal’ intake looks more suitable when we think on the prevention of dental fluorosis. Consequently, it

seems rational to monitor fluoride intake during the first 6-8 years of life. In this sense, maybe it would be

helpful to have different ranges of ‘optimal’ fluoride intake for small children and adults; nevertheless

additional studies are required before this can be implemented. But what should be our target for

‘optimal’ range of fluoride intake to avoid dental fluorosis? Incredibly, even after extensive efforts in

attempt to add scientific evidence to the empirically established range of 0.05 – 0.07 mg/kg fluoride per

day, not much progress has been achieved. Even the most well designed study on this topic (Iowa

Fluoride Study), after years of extensive investigation, came to the conclusion “Given the overlap among

caries/fluorosis groups in mean fluoride intake and extreme variability in individual fluoride intakes,

firmly recommending an ‘optimal’ fluoride intake is problematic 56”. This is not surprising in view of the

many factors that affect fluoride metabolism and alter the relationship between the amount of fluoride

ingested and the risk of developing dental fluorosis. What should we do then? Despite the ‘optimal’ range

of fluoride intake is not precisely known and might vary from individual to individual, it seems rational to

monitor the amount of ingested fluoride during the first 6-8 years of life or at least during the first 4 years

of life, to avoid fluorosis in the permanent central maxillary incisors. This can be done by controlling the

ingestion of fluoride from its main sources (water, dentifrice, infant formula and supplements). In

26

addition, periodical analysis of fingernails that reflect recent exposure to fluoride might help. Additional

gaps of knowledge in this field that would benefit from investigation are:

- Pre-eruptive effect of fluoride on caries progression into dentin;

- Effect of different types of exercise on the metabolism of fluoride;

- Relationship between malnutrition and dental fluorosis;

- Supplementation with calcium to reduce dental fluorosis;

- Relationship between gene polymorphisms and dental fluorosis;

- Comparison of 24-h diet recall with 5-day observational period to evaluate the patterns of

fluid intake of children across outdoor air temperature;

- RCTs on the efficacy of low-fluoride toothpastes to prevent caries in the primary dentition;

- Effect of brushing with a low-fluoride dentifrice (550 ppm) applied using the transversal

technique compared with a conventional dentifrice (1,100 ppm) in a pea-sized amount on

caries progression in situ and in vivo;

- Additional clinical trials of low-fluoride toothpastes whose formulations have been

modified to increase the anticaries efficacy;

- Periodical analyses of fluoride concentrations in infant formula, bottled water and infant

foods;

- Validation of biomarkers of exposure to fluoride.

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Table 1. Window of maximum susceptibility (WMS) to the development of dental fluorosis in the permanent maxillary central incisors 10

Type of studya

nb WMS Fluoride source References

1 86 6-23 months Toothpaste, supplements

Holm & Andersson, 1982 30

2 16 35-42 months Water Ishii & Suckling, 1986 41

1 139 1st two years Toothpaste Osuji et al., 1988 31

2 22-26 months Water Evans & Stamm, 1991 28

2 15-24 months for males21-30 months for females

Water Evans & Darvel, 1995 40

1 113 1st two years Toothpaste Lalumandier & Rozier, 1995 32

1 48 1st year Water Ismail & Messer, 1996 33

1 383 0-20 months Toothpaste, supplements

Wang et al., 1997 34

1 66 1st two years Water, toothpaste, supplements

Bårdsen & Bjorvatn, 1998 35

1 and 2c NAd 1st two years but duration of exposure more important

Variable Bårdsen, 1999 42

2 1,896 1st three years Water Burt et al., 2000 38

Burt et al., 2003 39

1e 579 1st two years Total intake Hong et al., 2006 36

1e 1st three years Total intake Hong et al., 2006 37

a 1- Study in individuals who were introduced to fluoride at different ages; 2- Study in populations which have experienced an abrupt reduction in daily fluoride intake.b Volunteers who completed the study.c Meta-analysis.d Not applicable.e Longitudinal design.

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Table 2. Studies assessing the association between dental fluorosis and gene polymorphisms in humans.

Main finding Reference

Col1A2 gene PvuII polymorphism is associated with dental fluorosis in populations highly exposed to fluoride

Huang et al., 2008 98

Osteocalcin gene HindII polymorphism is not associated with dental fluorosis Ba et al., 2009 221

Estrogen receptor gene RsaI polymorphism is associated with dental fluorosis in populations highly exposed to fluoride

Ba et al., 2011 100

Parathyroid hormone gene Bst Bi polymorphism is not associated with dental fluorosisWen et al., 2012 222

Myeloperoxidase gene polymorphism is associated with dental fluorosis Zhang et al., 2013 101

Calcitonin receptor gene AluI polymorphism is associated with dental fluorosis in populations highly exposed to fluoride

Jiang et al., 2015 102

42