Health risk from fluorid exposure of a population in selected areas of Tamil Nadu South India

Augustine Amalraj,Anitha Pius

Department of Chemistry,Gandhigram Rural Institute-Deemed University,Gandhigram,Dindigul 624302,Tamil Nadu,India

Abstract Prevalence of fluorosi is a public health problem in many states of India.It is necessary to fin out the different sources through which fluorid enters human metabolism.Only when the sources are identified suitable remedial measures can be initiated.This study was attempted to fin out the contribution of fluorid from various sources such as drinking water,staple food grains,cooked rice,green leafy vegetables and cow milk in the selected area of the study.Percentage of incidence of fluorosi was estimated using a clinical survey.Calculated community fluorosi index values in all the 22 villages were greater than the accepted index value.Fluoride intake from all manor dietary sources of different age groups such as infants,children,adolescents,adults and aged above 70 was determined.The study revealed that,of all the sources,fluorid contribution from drinking water is significan irrespective of age.Hence,it is advised that people of South India,where there similarity in diet pattern,consume drinking water with lesser fluorid to minimize the debilitating effect of fluoride The study also recommends to the government authorities concerned with supply to provide water with low fluorid level.

Keywords: Community fluorosi index;Total fluorid intake;Fluoride exposure level;Dietary sources

1.Introduction

About 80%of the diseases in the world are due to poor quality of drinking water,and the fluorid contamination in drinking water is responsible for 65%of endemic fluorosi in the world[1-3].Fluorosis is a slow,progressive,crippling malady,which affects every organ,tissue and cell in the body and results in health complaints having overlapping manifestations with several diseases.The disease“fluorosis has now become a global problem and the health impairment due to fluorosi has occurred in the citizens of about 25 nations across the globe,and more than 200 million people worldwide are at the risk of fluorosi[4].Fluoride toxicity depends for its severity on four factors(i.e.)the total dose ingested,duration of fluorid exposure,nutritional status and body’s response[5].

Different forms of fluorosi are dental,skeletal and nonskeletal.Dental fluorosi is a developmental disturbance of dental enamel caused by excessive exposure to high concentrations of fluorid during tooth development.The risk of fluorid overexposure occurs between the ages of 3 months and 8 years.In its mild forms fluorosi often appears as unnoticeable,tiny white streaks or specks in the enamel of the tooth.In its most severe form,tooth appearance is marred by discoloration or brown markings.The enamel may be pitted,rough and hard to clean.The spots and stains left by fluorosi are permanent and may darken over time.The severity of dental fluorosi depends on the amount of fluorid exposure,the age of the child,individual response,weight,degree of physical activity,nutrition,and bone growth.Skeletal fluorosi is a bone disease caused by excessive consumption of fluoride It causes pain and damage to bones and joints.Fluoride when consumed in excess,can also affect non-calcifie tissues besides bone and teeth is called as non skeletal fluorosis The soft tissue organs affected by fluorid are named in the following order:aorta,thyroid,lungs,kidneys,heart,pancreas,brain and spleen[6].

Fluoride is readily absorbed from the gastro-intestinal tract,with estimates of absorption ranging from 75%to 100%[7,8].The systemic fluorid absorption from water through the gastrointestinal tract into blood streamis nearly 100%by the process of simple diffusion without any intervention of overall water quality [9-11].Fifty percent of the ground water sources in India have been contaminated by fluorid and more than 90%of rural drinking water supply programs are based on ground water[12].Fluoride intake from water depends on the amount of water ingested through itself and the quantity of water ingested through food by means of water used for cooking and their flu oride content[13].Many previous studies,from various parts of the world reported the development of dental fluorosi even if the people consume drinking water with fluorid less than 1.0 mg/L[14,15],which implies that the optimal fluorid dose level in drinking water may vary with various features such as local climatic conditions,methods of food processing and cooking[16],amount of food and water intake and its fluorid and other nutrients level,and dietary habits of the community[17-19].

In India,there are only limited studies available in the literature on fluorid content of raw foods and it became clear that fluorosi varies within the population.Factors responsible for these variations could be fluorid intake by drinking water,dietary intake,especially intake of food grown in soil or irrigated with water rich in fluoride Therefore,the present study attempts to fin out the fluorid content of drinking water,staple food grains,green leafy vegetables,cow milk samples and also cooked rice predominantly consumed by the people of certain fluorid endemic areas of Tamil Nadu,South India and to correlate estimated daily intake with dental fluorosis The exposure doses of fluorid from the consumption of vegetable and cereal crops in children(3-14 years of age)are found to be higher than the limits laid down by Institute of Medicine,Washington,DC[20].This means the children in this age group are more likely at the risk of fluorosi from consumption of vegetables and cereal grown in the fluoridate areas.

The aim of the study is to fin out the average fluorid intake from various sources and to correlate with the percentage prevalence of fluorosis Identificatio of the source which contributes more is to be specifie so that effective interventions can be suggested to the people of the affected areas.In order to fin out the extent of fluorid contamination in drinking water and to fin out the fluorid exposure dose in Nilakottai block,an extensive study was accomplished by estimating fluorid level in drinking water.

2.Materials and methods

2.1.Selection of the study areas

The criteria for selection of the study area are the high percentage prevalence of dental fluorosi and the presence of fluorid in excess of 1 mg/L in drinking water samples.The percentage of dental fluorosi was estimated based on a clinical survey carried out among school children above 7 years(the age above which milk teeth are replaced by permanent ones)and among village adults.The villages were selected from the Nilakottai block of Dindigul district.Goggle earth satellite image of exact locations of fluorid endemic villages were prepared and is shown in Fig.1.

Fig.1.Location of fluorid endemic villages in Nilakottai block in Dindigul district.

2.2.Collection and analysis of drinking water samples

The study was carried out in 22 villages of the block 110 drinking water samples were collected from 22 villages in Nilakottai block,Dindigul district,Tamil Nadu,South India for fluorid analysis.Five drinking water samples were collected from each village of the selected areas and stored in clean,high density polyethylene bottles at 4°C before being analyzed.Fluoride levels in drinking water samples were measured by using fluorid ion selective electrode Orion ion analyzer(Thermo scientifi Orion 4 Star pH.ISE Benchtop) by diluting with total ionic strength adjustment buffer(TISAB III)in 10:1 ratio.The instrument was calibrated with standard fluorid solutions so chosen the concentration of one was ten times the concentration of the other and also that the concentration of the unknown falls between those standards.Then the concentration of the unknown was directly read from the digital display of the meter[21].

2.3.Collection and analysis of staple food grains

The chief staple food grains grown in the selected fluoroti villages were identified A total of 528 grain samples cultivated in the selected fluoroti villages were collected.Eight different grains,normally consumed by the people were collected in triplicate of different field of the selected areas using random sampling technique.To determine total fluorid in the grain samples,20 g of dried and ground grain sample was taken in nickel crucible,to which,10 mL calcium hydroxide suspension was added as a fluorid fixin agent and mixed thoroughly.The sample was evaporated to dryness in a hot air oven at 80°C and then ashed in muffl furnace at 600°C for 12 h.The ash was transferred to a distillation flask Sufficien analar silver sulphate was added to precipitate any chloride present in the sample.Pieces of glass and porcelain were also put in the flask 25 mL of 60%analar perchloric acid and 20 mL of distilled water were added and then distillation was carried out.The distillate collected between 135°C and 139°C,was neutralized using 0.2 mol/L sodium hydroxide and analyzed for fluorid using fluorid ion selective electrode[22].

2.4.Collection and analysis of solid food samples

A total of 66 cooked rice samples,three from each selected area were collected and analyzed as per the procedure adopted for food grain samples[22].

2.5.Collection and analysis of green leafy vegetables(GLV)

A total of 660 GLV samples grown and usually present in the diet of the people of selected area were collected,dried,ground and analyzed as per the standard procedure adopted [22].Ten different varieties were collected depending on the availability in an area.

2.6.Collection and analysis of cow milk samples

Among a total of 66 cow milk samples,3 from each selected area were collected in polyethylene bottles and refrigerated until the time of analysis.To determine fluorid concentration,50 mL of the cow milk sample was taken in a distillation flask sufficien silver sulphate was added to precipitate the chlorides present.50 mL of analar 60%perchloric acid was added and the distillate was collected between 135°C and 139°C and it was neutralized by using 0.2 mol/L sodium hydroxide and analyzed for fluorid using fluorid ion selective electrode[22].

2.7.Clinical survey and community fluorosis index(CFI)

Clinical survey was conducted among the people living in the selected areas.Dental examinations were made using dental mirrors and probes under indirect sunlight by two qualifie and well trained dentists.Community fluorosi index was calculated based on the symptoms of dental fluorosi using randomized sampling method,which is classifie into seven categories based on Dean’s classificatio viz.,normal,questionable,very mild,mild,moderate,moderately severe and severe and each of these seven classification were given a numerical weight such as 0.0,0.5,1.0,1.5,2.0,3.0 and 4.0,respectively[23,24].

Criteria for Dean’s fluorosi index are described as follows:Normal(0.0):the enamel represents the usual translucent semivitriform type of structure,and the surface is smooth,glossy,and usually of a pale creamy white color;Questionable (0.5):the enamel discloses slight aberrations from the translucency of normal enamel,ranging from a few white fleck to occasional white spots.This classificatio is utilized in those instances where a definit diagnosis of the mildest form of fluorosi is not warranted and a classificatio of “normal” is not justified Very mild (1.0):small opaque,paper white areas scattered irregularly over the tooth but not involving as much as 25%of the tooth surface.Frequently included in this classificatio are teeth showing no more than about 1-2 mm of white opacity at the tip of the summit of the cusps of the bicuspids or second molars;Mild(1.5):the white opaque areas in the enamel of the teeth are more extensive but do not involve as much as 50%of the tooth;Moderate (2.0):all enamel surfaces of the teeth are affected,and the surfaces subject to attrition show wear.Brown stain is frequently a disfigurin feature;Severe:includes teeth formerly classifie as “moderately severe (3.0)” and “severe(4.0).” All enamel surfaces are affected and hypoplasia is so marked that the general form of the tooth may be affected.The major diagnostic sign of this classificatio is discrete or confluen pitting.Brown stains are widespread and the teeth often present a corroded-like appearance[23-25].

People with symptoms of dental fluorosi were identifie and classifie in each category and the number of people in each category is multiplied by the corresponding numerical weight given in the brackets,the products thus obtained for the various categories are added up and the sum total divided by the total number of people surveyed,gives community fluorosi index.Only when the community fluorosi index value is greater than 0.6,fluorosi is considered to be a public health problem in that area[23,26].

The percentage incidence of fluorosi was estimated from the number of people affected by fluorosi from the particular area with total number of people surveyed.

2.8.Nutritional survey

The nutritional survey among the population of different age groups infants,children and adults was conducted in the selected endemic fluorid area.Based on household survey,quantity of drinking water,cow milk,GLVs,food grains and cooked food consumed per day by the various age groups of people and their body weight were accounted in the selected villages and the values were verifie with the data available with the local primary health centers.

2.9.Estimation of total fluoride intake(TFI)

Total fluorid intake(TFI)is the summation of the daily flu oride intake through entire diet sources.Daily fluorid intake(DFI) from the particular diet source was calculated by multiplying the fluorid concentration of the respective item with total quantity of the particular item consumed per day.

DFI=FC×QD

where FC is the fluorid concentration in the diet source and QD is the quantity of the diet intake per day.

2.10.Fluoride exposure dose through drinking water(ED)

The fluorid exposure dose through drinking water was calculated by the following equation:

where FC is the fluorid concentration(mg/L),WI is the water intake(L/day)and BW is the body weight(kg).

The assumption behind the calculation is persistent exposure and total bioavailability of fluorid concentration in water.The water intake of different age groups was estimated from the nutritional survey.Infants in their emergent life drink 250 mL of water as boiled water per day.In boiled water,fluorid level increases proportionally to the loss of volume,so the fluorid concentration in the actual drinking water was doubled[16].The estimated water intake for the infant,children and adult was 0.25,1.50 and 3.0 L per day,respectively.For the calculation,body weight of infants in the age group of 0-6 years was kept as 6 kg and children between 7 years and 19 years as 25 kg body weight and that of adults above 19 years as 60 kg body weight,the mean of water fluorid level in each village was used for fluorid exposure dose calculation.

2.11.Statistical analysis

Fluoride concentrations in water and diet sources such as grains,cooked food,green leafy vegetable and cow milk were expressed as mean and standard deviation of samples and the values are presented.The extend of linearity and correlation between water fluorid level with contribution of various diet sources was estimated using correlation coefficien (r) and coefficien of determination(R2).The quantity of numerical relationships between the total fluorid intake and the contribution of diet sources were expressed through regression analysis and were performed by using SPSS Inc,Origin Pro and Excel.

3.Results

3.1.Fluoride concentration in drinking water and CFI of the study area

All samples from 22 selected fluorid endemic villages contain more than 1.5 mg/L fluorid (Table 1).Among these villages,Akkarakaranpatti,Silukuvarpatti and Thoppinayakkanpatti contain more than 3 mg/L fluoride which is three times higher than the safe fluorid level in drinking water.14% of drinking water samples of the selected villages have more than 3 mg/L of fluoride remaining 86% drinking water samples have more than 1.5 mg/L fluoride Higher prevalence rate of fluorosi was observed from Akkarakaranpatti and Thoppinayakkanpatti that are 82.4% and 81.8%,respectively,with 1.57 and 1.34 CFI values.The numerical relationship between the water fluorid level with percentage of fluorosi prevalence and community fluorosi indices of the study areas indicated that,CFI values of all the selected endemic villages are higher than 0.6,a value above which fluorosi is considered to be a public health problem.Community fluorosi index and the percentage of fluorosi prevalence from all the selected endemic villages were shown in Table 2.The percentage of prevalence offluorosi and community fluorosi index increased significantl(r=0.977,R2=0.955,P＜0.0001) with the increase in drinking water fluorid level.The quantitative numerical relationship between the water fluorid level with percentage of fluorosi prevalence and community fluorid indices is shown in Table 3.Almost 30%of the people in the selected fluorid endemic villages are affected with more than mild form of fluorosi with higher CFI values.

Table 1 Drinking water fluorid levels in selected fluorid endemic areas of Nilakottai block.

Table 2 Community fluorosi index and percentage prevalence of fluorosi in the area of study.

Table 3 Correlation between total fluorid intake and related variables.

3.2.Correlation analysis between fluoride concentration in drinking water and maximum exposure dose

Correlation between the water fluorid levels with exposure dose of different age groups is shown in Table 4.High degree of correlation is obtained (r=0.99,R2=0.99) for all age groups.The significan levelsP＜0.0001 and the numerical relationship and the regression equations between the drinking water fluo ride level and fluorid exposure dose are listed in Table 4.The obtainedR2and analysis of variance indicate a high significan relationship between drinking water fluorid levels with the maximum exposure dose.The correlation coefficien and regression equations give the high significan linearity and numerical relationship between water fluorid levels with exposure dose.

3.3.Fluoride concentration in milk samples,green leafy vegetables,food grains and cooked foods

Table 4 Correlation between water fluorid level and exposure dose.

Table 5 Fluoride levels of cow milk samples in selected fluorid endemic areas.

The concentrations of fluorid i n samples of milk,green leafy vegetables,foodgrainsandcookedf oodscollectedfromselected endemic areas with varying fluo rid concentration in ground water are given in Tables 5-8.The results show that all the cow milk samples analyzed found to have very low levels of fluoride Fluoride levels in cow milk ranged from 0.043 mg/L to 0.147 mg/L in the selected fluorid endemic villages.All the varieties of green leafy vegetables grown in Sangarapuram,Thoppinayakanpatti,Silukuvarpatti and Akkarakaranpatti found to have high content of fluoride An interesting observation is that,in all the areas,theAcalypha indicaleaves recorded high fluorid level,but the majority of the people living in these endemic fluorid areas useMoringa oleiferaandSesbania grandiflorain their regular diet as per the nutritional survey data.BothMoringa oleiferaandSesbania grandifloraalso contain higher fluorid concentrations as shown in Table 6.In Akkarakaranpatti village the fluorid concentration inMoringa oleiferaandSesbania grandiflorawere recorded as 7.68 mg/kg and 4.47 mg/kg,respectively,and in the Thoppinayakanpatti village the fluo ride concentration inMoringa oleiferaandSesbania grandiflorawere recorded as 6.77 mg/kg and 4.88 mg/kg,respectively.Fluoride concentrations of food grain samples were analyzed and the results are given in Table 7.It was found that green gram,finge millet,pearl millet,fiel been and black-eyed bean contained the maximum amount of fluoride From the household survey,it was observed that,most of the people use blackeyed bean and fiel bean in their regular diet.Both the food grains also contain more fluorid concentration in all selected villages.Fluoride content in cooked food samples collected from selected villages was analyzed and it is given in Table 8.Samples from Akkarakaranpatti,Sankarapuram and Silukuvarpatti registered high content of fluoride Food grains particularly black-eyed bean recorded 4.21,4.12,4.12,4.11 mg/kg of flu oride content in Silukuvarpatti,Sankarapuram,Pallapatti and Akkarakaranpatti,respectively,and the fiel bean samples from Thoppinayakanpatti,Micheilpalayam and Sangalpatti recorded 3.11,2.95,2.67 mg/kg of fluoride respectively.

Basella alba 1.62 ± 0.09 1.65 ± 0.13 1.80 ± 0.10 1.64 ± 0.09 1.40 ± 0.11 0.62 ± 0.04 1.13 ± 0.12 1.22 ± 0.12 1.66 ± 0.20 2.26 ± 0.08 1.76 ± 0.09 2.30 ± 0.15 3.51 ± 0.22 1.66 ± 0.14 2.06 ± 0.12 2.23 ± 0.12 1.60 ± 0.21 4.32 ± 0.28 6.68 ± 0.19 3.39 ± 0.30 1.59 ± 0.17 1.67 ± 0.15 Acalypha indica 3.86 ± 0.10 3.48 ± 0.29 4.16 ± 0.11 4.16 ± 0.11 3.23 ± 0.19 0.58 ± 0.04 2.58 ± 0.04 2.37 ± 0.24 3.60 ± 0.21 3.48 ± 0.13 3.31 ± 0.16 3.86 ± 0.11 5.18 ± 0.08 3.40 ± 0.06 3.43 ± 0.14 1.30 ± 0.11 1.48 ± 0.03 5.93 ± 0.17 6.95 ± 0.23 4.79 ± 0.40 1.61 ± 0.19 1.48 ± 0.12 Amaranthus dubius 1.38 ± 0.15 1.17 ± 0.10 1.52 ± 0.30 1.56 ± 0.07 1.48 ± 0.20 0.88 ± 0.04 1.33 ± 0.21 1.09 ± 0.07 1.79 ± 0.09 1.41 ± 0.12 1.62 ± 0.09 1.72 ± 0.04 5.00 ± 0.17 1.47 ± 0.05 1.51 ± 0.09 1.17 ± 0.09 1.14 ± 0.07 4.75 ± 0.13 5.41 ± 0.22 5.43 ± 0.19 1.19 ± 0.10 1.49 ± 0.05 Alternanthera sessilis 1.75 ± 0.08 1.87 ± 0.25 1.84 ± 0.09 1.90 ± 0.16 0.96 ± 0.12 1.63 ± 0.10 1.92 ± 0.13 1.70 ± 0.15 2.15 ± 0.09 2.07 ± 0.10 1.51 ± 0.12 1.98 ± 0.12 4.41 ± 0.23 2.02 ± 0.10 1.91 ± 0.16 1.96 ± 0.17 1.68 ± 0.23 5.29 ± 0.17 5.79 ± 0.16 5.63 ± 0.17 1.30 ± 0.07 1.56 ± 0.14 Rumex acetosa 1.53 ± 0.29 1.55 ± 0.25 1.75 ± 0.09 1.46 ± 0.07 1.30 ± 0.07 1.02 ± 0.07 1.70 ± 0.57 1.36 ± 0.08 1.20 ± 0.16 1.46 ± 0.20 1.63 ± 0.13 2.16 ± 0.09 3.56 ± 0.32 2.04 ± 0.11 1.37 ± 0.17 1.58 ± 0.10 1.61 ± 0.16 4.43 ± 0.38 4.29 ± 0.29 4.52 ± 0.21 1.77 ± 0.13 2.06 ± 0.09 Amaranthus polygonoides 2.24 ± 0.11 1.84 ± 0.08 1.60 ± 0.20 2.37 ± 0.13 2.05 ± 0.13 1.02 ± 0.06 1.68 ± 0.06 1.58 ± 0.04 2.58 ± 0.04 2.27 ± 0.12 2.39 ± 0.10 2.17 ± 0.12 5.69 ± 0.51 1.17 ± 0.08 2.51 ± 0.14 2.35 ± 0.16 2.31 ± 0.17 5.83 ± 0.25 4.56 ± 0.29 6.39 ± 0.16 1.64 ± 0.19 1.96 ± 0.11 Amaranthus spinosus 1.97 ± 0.13 1.37 ± 0.10 1.74 ± 0.13 2.04 ± 0.11 1.67 ± 0.14 0.74 ± 0.09 1.54 ± 0.27 1.63 ± 0.21 2.35 ± 0.13 1.65 ± 0.15 1.66 ± 0.12 2.05 ± 0.09 6.13 ± 0.37 1.49 ± 0.19 2.59 ± 0.33 1.58 ± 0.25 1.68 ± 0.10 5.56 ± 0.28 4.76 ± 0.17 5.41 ± 0.20 1.76 ± 0.16 1.88 ± 0.21 Solanum nigrum 1.63 ± 0.34 1.45 ± 0.23 Fluoride levels of green leafy vegetables in selected fluorid endemic areas.1.58 ± 0.22 1.87 ± 0.08 1.10 ± 0.16 1.24 ± 0.05 1.24 ± 0.14 1.25 ± 0.03 2.04 ± 0.22 1.65 ± 0.09 1.26 ± 0.04 1.60 ± 0.29 5.09 ± 0.36 1.30 ± 0.05 0.92 ± 0.12 1.61 ± 0.20 1.62 ± 0.28 5.92 ± 0.09 5.76 ± 0.20 5.48 ± 0.16 1.97 ± 0.21 2.03 ± 0.12 Levels of fluorid (mg/kg),mean±SD Moringa oleifera 2.32 ± 0.14 1.45 ± 0.10 2.75 ± 0.12 2.32 ± 0.06 2.20 ± 0.16 3.68 ± 0.23 1.53 ± 0.25 1.74 ± 0.10 2.23 ± 0.06 1.97 ± 0.13 2.12 ± 0.09 2.58 ± 0.23 6.77 ± 0.51 2.30 ± 0.12 2.26 ± 0.10 1.58 ± 0.30 1.53 ± 0.15 5.48 ± 0.24 5.57 ± 0.28 7.68 ± 0.32 2.17 ± 0.09 3.72 ± 0.13 Sesbania grandiflora 2.79 ± 0.15 2.45 ± 0.20 3.49 ± 0.24 2.59 ± 0.29 2.60 ± 0.20 2.47 ± 0.08 2.48 ± 0.27 2.06 ± 0.12 2.32 ± 0.17 2.68 ± 0.15 2.45 ± 0.24 3.26 ± 0.17 4.88 ± 0.34 2.41 ± 0.19 2.67 ± 0.20 2.38 ± 0.33 2.54 ± 0.13 5.09 ± 0.19 4.28 ± 0.09 4.47 ± 0.26 2.75 ± 0.12 3.29 ± 0.17 Table 6 Name of the village Alagampatti Sangalpatti Micheilpalayam Ammapatti Uchanampatti Othur Chockanchettipatti Murugathuranpatti Kulalakundu Sandilarpuram Nagayakavundampatti Kolinjipatti Thoppinayakanpatti Meenachipuram Pallapatti Kattukothampatti Kanthappankottai Sangarapuram Silukuvarpatti Akkarakaranpatti Sithargalnatham Kullichettipatti

Table 7 Fluoride levels of food grains in selected fluorid endemic areas.

3.4.Impact of fluoride ion concentration in drinking water on total fluoride intake

Daily average intake level of various dietary sources by different age groups of people from the study areas as collected through nutritional survey is shown in Table 9.To calculate the range of fluorid intake of the various age groups of people in the fluoroti area,six villages from the selected area were selected.These six villages were selected based on the level of fluorid in drinking water.Two each from low,moderate and high fluoroti areas were selected and the values are given in Table 10 and shown in Fig.2.The total intake level of fluorid was compared with the average contribution from various dietary sources.The major fraction of the total fluorid intake was derived from water.For children,about 70%of the total flu oride intake was derived from drinking water.High degree of correlation was obtained (r=0.9991,R2=0.9982,P＜0.0001)between total fluorid intake and water fluorid level as shown in Table 11.

3.5.Impact of fluoride ion concentration in cow milk on total fluoride intake

All the cow milk samples analyzed found to have very low levels of fluoride Among these,milk samples from Thoppinayakanpatti and Silukuvarpatti which contain more than 0.12 mg/L fluorid was recorded as high within the samples analyzed.Fluoride levels in cow milk ranged from 0.043 mg/L to 0.147 mg/L in the selected fluorid endemic villages.Cow milk showed significan contribution on total fluorid intake(r=0.967,R2=0.935,P=0.0016)due to fluorid contamination as shown in Table 11.

Table 8 Fluoride levels of cooked rice in selected fluorid endemic areas.

Table 9 Average total intake level of various dietary sources as per nutritional survey.

3.6.Impact of fluoride ion concentration in green leafy vegetables(GLVs)on total fluoride intake

The fluorid concentrations in GLVs were determined and presented in Table 6.It was observed thatAcalypha indicaleaves grown in selected endemic villages had the maximum concentration of fluoride From the nutritional survey carried out in these areas,most of the people living in these villages reported that they predominantly consumeMoringa oleiferaandSesbania grandiflora.Both these GLVs when analyzed found to have high content of fluorid which also contributed significantl to high fluorid exposure level of people.The linear relationship between fluorid content of GLVs grown in fluoroti villages(r=0.9876,P=0.0002)and the total fluorid intake was established as shown in Table 11.GLVs are one of the major sources of daily fluorid intake.Daily fluorid consumption of teenagers from GLVs in low,medium and high fluorid endemic villages was found to be 0.38,0.58,1.66 mg/day,respectively.Green leafy vegetables contribute 13%of the fluorid intake within the sources analyzed.

3.7.Impact of fluoride ion concentration in food grain on total fluoride intake

Fluoride levels of food grain samples collected from the selected endemic fluoroti areas are shown in Table 7.From the nutritional survey,it was learnt that,most of the people living in these villages prefer to have black-eyed bean and fiel bean in their regular diet.The obtained statistical correlations in Table 11 illustrate a high linear relationship(r=0.9536,R2=0.9094,P=0.0032) between the total fluorid intake and intake through food grains.Among the dietary sources analyzed contribution of food grain is 10%.

3.8.Impact of fluoride ion concentration in cooked rice on total fluoride intake

Cooked rice is another major source of daily fluorid intake by children as well as adult people.0.34,0.38,0.50 mg/d of fluorid was consumed at an average from cooked rice by adults in low,medium and high fluoroti areas,respectively.The values ranged from 1.16 mg/kg to 4.94 mg/kg for various food grains.There is no definit pattern in the concentration of fluorid with respect to the variety or place.However,all the samples from villages like Nagayakavundampatti and Kullichettipatti registered relatively less fluorid in all the grains and samples from Alagampatti,Thoppinayakkanpatti and Silukuvarpatti recorded little higher values.Table 11 illustrates a high linear relationship(r=0.9939,R2=0.9878,P＜0.0001) between the total fluorid intake and fluorid from cooked rice.

4.Discussion

In view of the environmental and socio-economic conditions of the Indian sub continent,the desirable limit of the fluorid is set at 0.60-1.20 mg/L and maximum permissible limit in absence of any other source is set at 1.5 mg/L for drinking water[27].In this study about 98% of the daily fluorid intake by infants through their diet sources was derived from the local drinking water sources.Children residing in the fluoroti areas acquire nearly double the amount of fluorid than the optimal level.Because of powder-based infant formulae reconstituted with water from high fluorid endemic area containing more than 0.5 mg/L of fluorid may provide a daily fluorid intake of above the threshold limit of 0.10 mg/kg [28-30].Drinking water seems to be the major contributor of fluorid among all the sources analyzed.

Fluoride ingestion through food is comparatively less than through water.However,it cannot be neglected in the endemic areas because it will increase the fluorid burden in addition to water.Fluoride not only enters through water,but also through many edible items.Fluoride of food items depends upon the fluorid contents of soil and water used for irrigation.Thus,fluorid in water contributes significantl to the total exposure of an individual to this element,but not the only source of exposure.In certain areas,the major contribution comes from diet sources as in the case of Vranjska Banja where agricultural products including potatoes,beans,tomato,cucumbers,water melons,etc.which are still traditionally grown have high fluorid content[31].

Akkarakaranpatti 0.680 0.017 1.020 0.023 Silukuvarpatti 0.640 0.018 0.960 0.024 Ammapatti 0.376 0.008 0.564 0.010 Meenachipuram Average daily fluorid intake from the diet source(mg/d)0.364 0.010 0.546 0.013 Sangalpatti 0.308 0.007 0.462 0.009 Kattukuthampatti 0.304 0.007 Total fluorid intake per day from each dietary source in selected fluorid endemic areas.0.456 0.010 Average daily intake level of dietary source 200 mL 150 mL 300 mL 200 mL Dietary source Drinking water Cow milk Drinking water Cow milk Table 10 Age group and weight range Infants Birth to 6 months(2-6 kg)Above 6-12 months(6-10 kg)1.700 0.283 0.768 0.206 0.146 2.550 0.045 1.152 0.411 0.365 6.800 0.045 1.920 0.617 0.548 1.600 0.030 0.557 0.211 0.138 2.400 0.048 0.836 0.421 0.345 6.400 0.048 1.393 0.632 0.518 0.940 0.013 0.232 0.138 0.108 1.410 0.020 0.348 0.276 0.270 3.760 0.020 0.580 0.414 0.405 0.910 0.017 0.230 0.129 0.102 1.370 0.027 0.345 0.259 0.255 3.640 0.027 0.575 0.389 0.383 0.770 0.011 0.145 0.079 0.096 1.155 0.018 0.218 0.157 0.240 3.08 0.018 0.363 0.236 0.360 0.760 0.012 0.158 0.072 0.090 1.140 0.019 0.237 0.144 0.215 3.040 0.019 0.395 0.216 0.338 500 mL 250 mL 100 g 50 g 200 g 750 mL 400 mL 150 g 100 g 500 g 2000 mL 400 mL 250 g 150 g 750 g Drinking water Cow milk Greens Grains Rice Drinking water Cow milk Greens Grains Rice Drinking water Cow milk Greens Grains Rice Children From 1 to 3 years(11-18 kg)From 3 to 10 years(18-25 kg)From 10 to 18 years(25-45 kg)6.800 0.040 1.920 0.822 0.730 6.400 0.042 1.393 0.842 0.690 3.760 0.018 0.580 0.552 0.540 3.640 0.023 0.575 0.518 0.510 3.080 0.015 0.363 0.314 0.480 3.040 0.017 0.395 0.288 0.450 2000 mL 350 mL 250 g 200 g 1000 g Drinking water Cow milk Greens Grains Rice Adult From 18 to 70 years(45-70 kg)5.100 0.034 1.536 0.617 0.730 4.800 0.036 1.114 0.632 0.690 2.820 0.015 0.464 0.414 0.540 2.730 0.020 0.460 0.389 0.510 2.310 0.013 0.290 0.236 0.480 2.280 0.014 0.316 0.216 0.450 1500 mL 300 mL 200 g 150 g 1000 g Drinking water Cow milk Greens Grains Rice Old Above 70 years(50-70 kg)

Fig.2.Average body weight vs total fluorid intake of the respective age group of population.

All individual sources of exposure are important since it is the total fluorid intake from all sources that is critical in the development of fluorosi [32].Among various sources,infant foods,milk formulas,foods containing chicken,some bottled waters and beverages,were identifie as significan sources of ingested fluorid by many researchers [33-37].Fluoride is entering human food chain in increasing amount through the consumption of tea,wheat,spinach,cabbage,carrots and other Indian foods [38-40].The observations from studies done in China suggested that food can significantl contribute to the total fluorid uptake[41,42].

In Japan,it has been observed that rice and green leafy vegetables(GLVs)main staples in this country produced in polluted areas contained fluorid that is several to more than ten times higher than that found in non-contaminated products[43].The absorption rate of fluoride contained in rice and GLVs produced in fluoride-pollute areas was greater than 90%approximating that of NaF[44,45].Sorghum and ragi are consumed extensively in several areas in southern parts of India,where endemic fluoro sis is rampant[46].It is reported that in these areas,the incidence and severity of fluorosi were higher when the staple foods were sorghum or ragi,rather than rice.Rice(Oryza saliva)is a major dietary staple of nearly half the world’s population.About 95%of this cereal is produced and consumed in South East Asiancountries,including India.It is estimated that more than half of the population of India subsists on rice [47].In rural India,home-made parboiled rice is consumed even in many villages where fluorosi is endemic.The economically backward people,who are the main victims of fluorosi [46],resort to parboiling paddy at the household level.For this purpose,invariably,they use the same source of locally available water which may have unsafe levels of fluoride It seems possible that such a practice may enhance the fluorid concentration of rice.This study also shows that,the fluorid intake from cooked rice is about 6%-8%in medium and high fluoroti villages,respectively.

Table 11 Correlation between total fluorid intake per day and various dietary sources.

Calculated daily intake of fluorid from water,green leafy vegetables,food grains,cow milk and cooked rice in high flu orotic areas of the study shows 3-5 times higher fluorid than 2 mg/d that was estimated as worldwide average for fluorid intake in children and adolescent.For children and adolescents during their developmental age,fluorid daily threshold of 2 mg was recommended by the World Health Organization[3].

Cow milk is a suitable alternative for human milk;fluorid content of samples of cow milk analyzed in this study found to range from 0.04 mg/L to 0.15 mg/L.Fluoride exposure level through milk infants in low fluoroti areas and high fluoroti areas is found to be 2.1%and 2.4%,respectively.Even in high fluoroti area cow milk samples do not have excess fluoride

The WHO[3]stated that“The most serious effect is the skeletal accumulation of fluorid from long-term excessive exposure to fluorid and its effect on non-neoplastic bone disease specifically,skeletal fluorosi and bone fractures”.There is clear evidence from India and China that skeletal fluorosi and an increasing risk of bone fracture occurred at total intake of 14 mg fluoride/ and evidence suggestive of an increasing risk of bone effect at a total intake of about 6 mg fluoride/ [48].From the results obtained in this study and previous studies,it is evident that after nearly 30 years of experience,the defluoridatio system does not provide a safe and dependable supply of domestic water [48].As a defluoridatio plant using activated alumina was installed in one of the villages of the study area in the year 2001 with the financia support from Tamil Nadu water and drainage (TWAD) board,a government institution concerned with provision of safe drinking water is not properly utilized by the people of the area.Another drawback is that the plant is not properly monitored and necessary periodical regeneration of the adsorbent is not carried out.

Adults in high fluoroti areas were exposed to 2 times higher fluorid intake per day than the recommended level of 4 mg/d through all the selected diet sources analyzed.Fluoride levels of all selected diet sources analyzed in medium and high flu orotic areas are nearly 2 and 3 times higher,respectively,than the low fluoroti area diet.The drinking water fluorid level has significan influenc on fluorosi prevalence in the study area as it contributes around 66% of the total fluorid intake per day.If water used for drinking or food processing has fluorid level more than 0.65 mg/L,it enhances the daily total fluorid intake more than the standard limit of 4 mg/d for normal adults recommended by the Agency for Toxic Substance and Disease Registry[49,50].Our finding regarding fluorid levels of food grains and green leafy vegetables show that foods grown in high fluoroti areas have significantl higher fluorid than those grown in low fluoroti area.

5.Conclusion

Planning the supply of low-fluorid water for a fluorid endemic community requires a thorough knowledge of local hydro-geological conditions.An exhaustive inventory of the water sources being used by the community is essential.This study identifie water as the major causative factor for fluorid toxicity among other sources,such as cow milk,food grains,green leafy vegetables and cooked rice though,these are consumed in larger quantity by the people,their contribution is less towards the fluorid toxicity of the area studied.Supply of drinking water with less fluorid is emphasized.Rainwater harvesting techniques should be promoted since they have a dilution effect on the fluorid concentration of the ground water of the affected villages.

Acknowledgment

We would like to thank the University Grants Commission,Government of India for the financia support [F.No.39-806/2010(SR)].