Science of the Total Environment 407 (2009) 4168–4173

Contents lists available at ScienceDirect

Science of the Total Environment

j ourna l homepage: www.e lsev ie r.com/ locate /sc i totenv

Trace element levels in whole blood of riparian villagers of the Brazilian Amazon

Jairo Lisboa Rodrigues a, Bruno Lemos Batista a, Myriam Fillion b, Carlos J.S. Passos c,Donna Mergler b, Fernando Barbosa Jr. a,⁎a Laboratório de Toxicologia e Essencialidade de Metais, Depto. de Análises Clínicas, Toxicológicas e Bromatológicas, Faculdade de Ciências Farmacêuticas de Ribeirão Preto-USP,Avenida do Café s/n, Monte Alegre, 14040-903, Ribeirão Preto-SP, Brazilb Centre interdisciplinaire de recherche sur la biologie, la santé, la société et l'environnement (CINBIOSE), Université du Québec à Montréal, Canadac Faculdade UnB Planaltina (FUP), Universidade de Brasília, Planaltina (DF), Brazil

⁎ Corresponding author. Tel.: +55 16 36024701; fax:E-mail address: fbarbosa@fcfrp.usp.br (F. Barbosa).

0048-9697/$ – see front matter © 2009 Elsevier B.V. Adoi:10.1016/j.scitotenv.2009.02.041

a b s t r a c t

a r t i c l e i n f o

Article history:Received 19 November 2008Received in revised form 14 February 2009Accepted 26 February 2009Available online 29 March 2009

Monitoring the nutritional status of essential elements is of critical importance in human health. However,trace element concentrations in biological fluids are affected by environmental and physiological parameters,and therefore considerable variations can occur between specific population subgroups. Brazil is a largecountry with much food diversity. Moreover, dietary habits differ from north to south. As an example, thetraditional populations of the Brazilian Amazon basin are heavily dependent on fish, fruits, vegetables andmanioc for their daily sustenance. However, very few studies have examined to what extent these dietsreflect adequate nutritional status for essential elements. Then, in the present study we have evaluated thelevels of some trace elements (Cu, Co, Zn Sr, and Rb) in the whole blood of a riparian Brazilian Amazonianpopulation and estimated the influence of age and gender on levels and inter-element interactions in thesame population. For this, 253 subjects, aged 15 to 87, from 13 communities situated on the banks of theTapajós, one of the major tributaries of the Amazon, were randomly selected. The values found for cobalt,copper and strontium in whole blood are in the same range as in other populations. On the other hand, thelevels of rubidium and zinc may be considered higher. Moreover, gender was shown to influence Zn and Culevels while age influenced the concentrations of Sr and Rb in men and Cu in women. Given the scarcity ofstudies examining nutritional status in traditional communities of the Amazon, our study is the first toprovide relevant insight into trace element values in this region and inter-element interactions. This paper isalso of particular importance for future studies looking at the possible protective effects of traditionalAmazon riparian diets against mercury intake from fish consumption.

© 2009 Elsevier B.V. All rights reserved.

1. Introduction

The significance of trace elements in human health and disease hasbeen reviewed in numerous scientific publications (Hambidge 2000,2003). Trace elements may be either essential or non-essential.Essential elements are those that are required by an organism tomaintain its normal physiological function. This definition of essentialelements is based mainly on the observation that inadequate intakecauses impairment of human health which can be reversed byadministering the element. Many essential trace elements are parts ofprotein complexes, (e.g., metalloproteins), which are required forenzymatic activities and can play structural roles in connective tissueor cell membranes. Non-essential trace elements are considered toxicand are not required for physiological processes, but essentialelements can also cause toxicity when concentrated at excessivelevels (Bryan et al., 2007). Without essential elements the organismcannot complete its normal life cycle or achieve normal healthy

+55 16 3602 4881.

ll rights reserved.

growth. Thus, monitoring the nutritional status of essential elementsis of critical importance in human health. Today, biomonitoring oftrace elements in human blood has become an important tool fordiagnosing essential element deficiency (Angerer et al., 2007; Bryanet al., 2007; Parsons and Barbosa, 2007; Zanao et al., 2002). However,essential trace element concentrations in biological fluids are affectedby environmental and physiological parameters, and thereforeconsiderable variations can occur between specific population sub-groups (Kristiansen et al., 1997, Christensen, 1995). For this reason,several large-scale studies have been undertaken in an effort toestablish the baseline ranges of trace elements in specific populationsaround the world (Cornelis et al., 1994; Gil et al., 2006; Goullé et al.,2005; Kucera et al., 1995; Iyengar, 1987; Kouremenou-Dona et al.,2006; Kristiansen et al., 1997; Minoia et al., 1990; Moreno et al., 1999;Schuhmacher et al., 1994; Templeton et al., 1994). In the United States,the analysis of some essential elements in the U.S population, such asselenium and iron, is part of the U.S. National Health and NutritionExamination Survey (NHANES) (CDC, 2008). In European countries,many studies describe baseline levels of essential elements in adults(Minoia et al., 1990; Kristiansen et al., 1997;White and Sabbioni, 1998)

Fig. 1. Map of the study area. Participating communities are identified by a large red dot.

Table 1Socio demographic characteristics of the study population (n =253).

Characteristics Total Women Men

Age (years) % % %15–30 28 31 2431–45 29 30 2846–60 26 27 2461–75 14 10 1976–90 4 2 6

Years of education0–3 25 21 314–6 39 41 367–9 19 14 2510–12 14 19 613–15 3 4 1

4169J.L. Rodrigues et al. / Science of the Total Environment 407 (2009) 4168–4173

and children (Bárány et al., 2002a; Malvy et al., 1993). However, nolarge surveys including the determination of trace elements in bloodsamples have been carried out among Brazilians.

Brazil is a large country with much food diversity. Moreover,dietary habits differ from north to south. As an example, thetraditional (e.g., riparian and indigenous) populations of the BrazilianAmazon basin are heavily dependent on fish, fruits, vegetables andmanioc for their daily sustenance. Earlier studies have shown that therelative food diversity and strong seasonality of Amazon riparian diets(Murrieta, 1998; Murrieta et al., 1999; Passos et al., 2001, 2007) caninfluence the dietary intake and bioavailability of trace elements.However, very few studies have examined to what extent these dietsreflect adequate nutritional status for essential elements. Moreover,nutritional status may modulate the toxic effects of mercury exposurefrom contaminated fish (Grotto et al., 2009).

The main aims of the present study are to 1) evaluate the levels ofessential trace elements (Cu, Co, Zn Sr, Rb) in the whole blood of ariparian Amazonian population in Brazil; and 2) estimate the influenceof age and gender on levels and inter-element interactions in the samepopulation.

2. Materials and methods

2.1. Population and study design

This study is part of a larger interdisciplinary investigation studyingHg sources and environmental dynamics, human exposure and healtheffects in the Tapajós River valley (Brazilian Amazon, state of Pará). In

Mayand July 2006 a cross-sectional studywas carried out with a total of450 people from the Tapajós region. Participants provided bloodsamples and responded to interview-administered questionnairescovering socio-demographics, lifestyle, dietary habits, and residentialand work history. Of these, 253 (47% women and 53% men), aged 15 to87, from13 communities situated on the banks of the Tapajós, one of themajor tributaries of the Amazon (Fig.1), were randomly selected for thepresent study. Table 1 summarizes the socio-demographic character-istics of this study population. Ethical approval was obtained from theEthics Review Board at the University of São Paulo in Ribeirão Preto(Brazil).

Table 2Analytical performance for the determination of trace elements in blood referencesamples (PT/EQAS) from the Institut National de Sante' Publique du Quebec (INSP)with the proposed method.

Analyte ICP 07B03 (n=5) ICP 07B06 (n=5)

Valueobtained(µg/L)

Target value(acceptable range)µg/L

Valueobtained(µg/L)

Target value(acceptable range)µg/L

Co 8.7±0.2 8.7±1.2 4.2±0.1 4.1±0.3Cu 801±18 888±78 2385±23 2350±114Rb – – – –

Sr – – – –

Zn 6377±22 6770±380 6140±633 6416±101

(Values obtained are denoted as mean±SD, n=5).

4170 J.L. Rodrigues et al. / Science of the Total Environment 407 (2009) 4168–4173

2.2. Sample collection

A trained Brazilian nurse collected 3-mL blood sample from eachparticipant. Blood samples were collected in “trace metal free”evacuated tubes (BDVacutainer®) containing heparin as anticoagulant.Two (2) ml of blood was then pipetted into an eppendorf tube (2 mLvolume) previously cleaned in a 100 clean roomand immediately frozenat−20 °C before analysis.

2.3. Sample analysis

2.3.1. ReagentsAll reagents used were of analytical-reagent grade except for HNO3,

which was previously purified in a quartz sub-boiling still (Kürner)before use. A clean laboratory and laminar-flow hood capable ofproducing class 100 were used for preparing solutions. High purity de-ionized water (resistivity 18.2 MΩ cm) obtained using a Milli-Q waterpurification system (Millipore, Bedford,MA, USA)was used throughout.All solutions were stored in high-density polyethylene bottles. Plasticbottles and glassware materials were cleaned by soaking in 10% (v/v)HNO3 for 24 h, rinsed five times with Milli-Q water and dried in a class100 laminar flow hood before use. All operations were performed in a10000 clean room.

2.3.2. InstrumentationWe used a PE ELAN DRC II ICP-MS instrument (Perkin Elmer Life

and Analytical Sciences) for the determination of elements in blood.Typical daily instrumental parameters are provided by Palmer et al.(2006). The ICP-MSwas operated with Pt sampler and skimmer cones,both purchased from Perkin Elmer. Argon 99.999% (White Martins,São Paulo, Brazil) was used. Multielement stock solutions containing1000 mg L−1 of each element were obtained from Perkin-Elmer(PerkinElmer, Norwalk, CT). Rhodiumwas used as internal standard atthe concentration of 10 µg/L Rh. The internal standard was dilutedfrom a 1000 mg/L stock standard solution (PerkinElmer, Norwalk, CT,

Table 3Descriptive statistics for trace elements in whole blood measured in the present study and

Mean(µg/L)

Standard Deviation(µg/L)

Median(µg/L)

Range(µg/L)

10th percenti

Cu 920 176 911 495–1578 735

Co 0.40 0.30 0.32 0.005–2.090 0.048

Zn 8945 1019 9341 3518–12294 7744

Sr 30.9 12.0 27.10 11.56–78.60 18.95Rb 6638 1293 6557 3328–9429 5110

USA). A rinse solution consisting of 0.005% (v/v) Triton X-100® in 2%(v/v) nitric acid was prepared before each run.

2.3.3. Determination of trace elements in whole bloodTrace metal levels in whole blood were determined by Inductively

Coupled Plasma-Mass Spectrometry (ICP-MS) as proposed by Palmeret al. (2006).

The method detection limit was 0.09, 0.005, 0.01, 3.8 and 0.1 µg/Lfor Cu, Sr, Co, Zn and Rb, respectively.

2.3.4. Analytical quality controlQuality control (QC) of trace metal determination in blood

specimens was guaranteed by analyzing two reference materialsprovided by the Institut national de santé publique du Québec, Canada(INSP-external quality assessment scheme (EQAS) for trace elementsin blood). Reference samples were analyzed before and after tenordinary samples. Values obtained together with reference values areshown in Table 2. Since no blood reference material containingreference values for Rb and Sr is available, we validated our data for Rband Sr by comparing the obtained values in 5 ordinary blood samplesby our method and by graphite furnace atomic absorption spectro-metry (GF AAS). The results for both elements were in closeagreement between the two techniques.

2.3.5. Statistical analysisGeneral descriptive statistics was employed to present the main

findings of this study. A non-parametric t test was used to verifywhether or not there were significant differences between meanconcentrations of the elements according to gender dichotomization.When evaluating possible mean differences in element concentrationsamong the various age categories, analysis of variance (ANOVA) wasemployed. Spearman correlation was used to verify the inter-elementcorrelations in whole blood. Statistical procedures were performedusing Statview for Windows Version 5.0.1 and JMP 5.0.1a (SAS InstituteInc.) and GraphPad Prism Version 4.0.

3. Results and discussion

The present study is the first to provide data for levels of traceelements (cobalt, copper, rubidium, strontium and zinc) in the wholeblood of a riparian population living in the Amazon region.

The essentiality of cobalt, copper, strontium, and zinc is extremelywell-known. However, even if the essentiality of rubidium has notbeen proven, toxic effects have not been reported so far. This may bedue to the lack of reference levels for this element in the generalpopulation. Accordingly, we also provide rubidium levels in the wholeblood of our population as complementary data.

Sociodemographic characteristics of the study population are showninTable 1. Age varied from15 to 87years (mean42.0 years±16.7). A lowlevel of formal education (mean 4.9 years±3.5), characteristic of rural

elsewhere (n=253).

le 90th percentile Concentrations of trace elements (µg/L) for otherpopulations

1122 Mean of 950±100 µg/L Heitland and Koster (2006)Mean of 920 µg/L Bárány et al. (2002a,b)

0.683 Mean of 0.5 µg/L Kristiansen et al. (1997)Mean of 0.3 µg/L Barany et al. (2002a)

8996 Mean of 6100 µg/L Bárány et al. (2002a)median 5450 µg/L Rosborg et al. (2007)

44.57 Range of 20–70 µg/L Heitland and Koster (2006)7821 Mean of 2408 µg/L Heitland and Koster (2006)

Median of 2030 µg/L Rosborg et al. (2007)Mean of 2800 µg/L Barany et al. (2002a)

Table 4Trace elements in whole blood of men and women, and results of inter-sex comparisons.

Elements Mean(µg/L)

Standard deviation(µg/L)

Median(µg/L)

Range(µg/L)

p valuet-Test

Cu men 874 139 865 495–1235 b0.0001Cu women 965 178 930 591–1578Co men 0.367 0.270 0.324 0.005–1.258 0.8662Co women 0.361 0.291 0.323 0.010–2.090Zn men 9102 1089 9503 5605–11841 0.0252Zn women 8432 1199 8138 3518–12291Sr men 30.14 11.33 27.00 15.42–78.60 0.3438Sr women 28.91 9.24 27.10 11.56–54.31Rb men 6562 1115 6709 3328–8792 0.4339Rb women 6454 1061 66428 4198–9429

4171J.L. Rodrigues et al. / Science of the Total Environment 407 (2009) 4168–4173

riparian communities in remote forest areas, was observed for thepopulation of this study.

For the population in general, the mean concentrations of Cu, Co,Zn, Sr and Rb were 920±176, 0.4±0.3, 8945±1019, 30.9±12.0 and6638±1293 µg/L, respectively. The values found for cobalt, copper andstrontium inwhole blood are in the same range as in other populations(Bárányet al., 2002a,b,c;Caroli et al.,1994;Clark et al., 2007;Goullé et al.,2005;Minoia et al.,1990).On the otherhand, levels of rubidiumand zincmay be considered higher than those typically reported for otherpopulations (Barany et al., 2002a; Caroli et al., 1994; Goullé et al., 2005;Heitland and Koster, 2006; Minoia et al., 1990). Table 3 provides thedescriptive statistics for trace elements inwhole blood measured in thepresent study.

3.1. Cobalt

Cobalt exists in the earth's crust as a relatively rare metal and, as acomponent of the vitamin B12 complex, it is an essential trace element.Thedaily intake of cobalt from foodhas beenestimated to bebetween5–45 µg/day, mainly fromvegetables,meat and pluck (Christensen,1995).

In this populationwe foundmean cobalt levels of 0.4±0.3 µg/L. Thevalues found are in close agreementwith those reported byMinoia et al.(1990), Goullé et al. (2005) and Barany et al. (2002a,b) in Italian, Frenchand Swedish populations, respectively. Minoia et al. (1990) evaluated350healthy Italian subjects andobserved cobalt levels varying from0.01to 0.91 µg/L in blood. Goullé et al. (2005) have assessed cobalt in the

Table 5Trace elements according to age categories.

Age (years) n Mean Cu ± SD (µg/L) Mean Co±S.D (µg/L)

Men15–30 26 853±159 0.457±327

(p=0.7648) (p=0.2025)31–45 30 864±113 0.364±0.198

(p=0.9735) (p=0.2580)46–60 26 865±150 0.283±0.254

(p=0.1547) (p=0.2517)61–75 20 925±136 0.375±0.283

(p=0.4997) (p=0.7127)76–90 6 881±136 0.328±0.308

(p=0.6590) (p=0.2942)

Women15–30 45 1015±209 0.409±0.367

(p=0.0483) (p=0.5590)31–45 43 950±161 0.372±0.292

(p=0.6015) (p=0.3396)46–60 39 930±172 0.310±0.217

(p=0.4134) (p=0.9263)61–75 15 974±112 0.303±0.191

(p=0.3065) (p=0.8879)76–90 3 819±191 0.345±0.206

(p=0.095) (p=0.7136)

blood of 100 healthy French volunteers. They reported levels from 0.04to 0.64 µg/L. Barany et al. (2002a) found that cobalt values in the bloodof Swedish adolescents varied from b0.2 to 0.9 µg/L. On the other hand,we found much higher variations in cobalt levels between thevolunteers when compared to other studies, from 0.005 to 2.090 µg/L.In a more recent study, we evaluated the composition of essentialelements, including cobalt, in the typical diet of this riparianpopulation.Certain components of their diet such as Brazil nuts and fish are richsources of cobalt. Nardi et al. (2009) also showed a mean cobalt level of9.1 µg/g in some fish species in this region.

3.2. Copper

Copper (Cu) is in the structure of various enzymes, especiallysuperoxide-dismutase, which also participates in cleaning up freeoxygen radicals from the intracellular environment (Terrés-Martoset al., 1997). Copper is widely distributed in foods. The accumulationof copper in plants is not affected by the copper content of the soil inwhich they grow. Organ meats, seafood, nuts, and seeds are majorcontributors of dietary copper (Pennington et al., 1999).

The concentration of copper in blood (mean 920±176 µg/L) thatwe found is verymuch in line with the values reported by Bárány et al.(2000a,b) for a Swedish adolescent population (950±100 µg/L), andby Heitland and Koster (2006) for an adult French population(1042 µg/L).

3.3. Rubidium

Rubidium is another element added to the list of possible essentialelements. Rubidium resembles potassium, especially in its pattern ofabsorption, and may play a role similar to that of potassium (Nielsen,1998; Kosla et al., 2002).

The concentration of rubidium found in the blood of ourpopulation (mean 6638±1293 µg/L) is considerably higher thanthe values observed by Goullé et al. (2005) (1289–2358 µg/L) for aFrench population and byMinoia et al. (1990) for an Italian population(900–4145 µg/L). Unfortunately, we do have an explanation for thisdifference. Probably, rubidium levels may be higher in some typicalfoods consumed by this population. However, future studies arenecessary to better explain our findings.

Mean Zn±S.D (µg/L) Mean Sr±S.D (µg/L) Mean Rb±S.D (µg/L)

9469±1140 26.87±12.30 6757±792(p=0.8601) (p=0.3534) (p=0.8827)9417±1046 29.66±8.82 6714±1288(p=0.9013) (p=0.7555) (p=0.8966)9453±1031 30.59±10.89 6752±990(p=0.6054) (p=0.8007) (p=0.1871)9283±1303 36.43±10.74 5975±1164(p=0.3714) (p=0.0502) (p=0.0208)9745±746 40.41±17.79 5997±1276(p=0.993) (p=0.0085) (p=0.0500)

9020±1113 30.12±10.15 6290±865(p=0.7731) (p=0.3053) (p=0.7556)8946±1012 28.08±8.64 6360±1166(p=0.3119) (p=0.6377) (p=0.3223)9216±1056 29.05±8.47 6593±909(p=0.7505) (p=0.3956) (p=0.4634)9132±2111 26.64±7.56 6830±1532(p=0.4394) (p=0.3542) (p=0.7090)10922±216 32.11±0.19.11 6579±1280(p=0.0205) (p=0.7203) (p=0.6477)

Table 6Statistically significant Spearman's rank correlation coefficients (R) for the correlationbetween trace elements in blood.

Elements Cu Co Zn Sr Rb

Cu – 0.1139 – – −0.1186(0.048) (0.0395)

Co – – – 0.177 (0.002) –

Zn – – – 0.1623 0.4034(0.0047) (b0.0001)

Sr – – – – –

Rb – – – – –

(p values in parenthesis).

4172 J.L. Rodrigues et al. / Science of the Total Environment 407 (2009) 4168–4173

3.4. Strontium

There is evidence to suggest that strontium (Sr) is an essential tracemineral (Shahnazari et al., 2006). It is similar to calcium in chemicalcomposition and is necessary for proper bone growth and prevention ofdental caries (Curzon and Losee, 1977). Relatively large concentrationsof strontium are deposited in bones and teeth, replacing part of thecalcium in hydroxapatite crystals (Bechkoff et al., 2008).

This population's blood concentrationof strontium(mean30.9 µg/L)is in linewith that reported by Goullé et al. (2005) (range of 9–41 µg/L).

3.5. Zinc

Zinc is an important essential element and plays a major role inprotein synthesis and gene expression. Moreover, zinc stabilizes thestructures of proteins and nucleic acids and is also involved in immunephenomena (Camara and Amaro, 2003; Prasad, 1998; Salgueiro et al.,2000).

The concentration of zinc found in the blood of this population(mean 8945 µg/L) is much higher than the values observed for otherpopulations (Barany et al., 2002a; Goullé et al., 2005;Minoia et al.,1990;Rosborg et al., 2007). The difference could be explained by the higherlevels of zinc in the diet of Amazonianpopulations compared to the dietsof other populations as demonstrated by Nardi et al. (2009). Brazil nuts(34±8 µg/g) and, to a lesser extent, beans (29±3 µg/g) are the majorsources of this essential element in the food typically consumed by theAmazonian riparian population (Nardi et al., 2009).

3.6. The influence of age and gender in the concentration of elements inblood

Results show that there were significant differences in bloodconcentration for Cu and Zn (pb0.0001, p=0.025, respectively), butnot for cobalt, strontium and rubidium, between men and women.Results are summarized in Table 4. It can be seen that zinc levels arelower inwomen thanmen.Conversely, Cu levelswere lower inmen thanwomen. These findings could be explained by differences betweenmen and women in both dietary habits and element absorption. Thebioavailabilityof copper ismarkedly influencedby the amountof copperin thediet. Bioavailability ranges froma75-percent adsorptionofdietarycopper when the diet contains only 400 µg/day to a 12-percentadsorption for a diet containing 7.5mg/day (Turnlund,1998). Themainsources of copper in the typical diet of this population are beans andBrazil nuts (Nardi et al., 2009)while zinc comesmainly from rice, beans,corn and Brazil nuts (Nardi et al., 2009). It is also well known that thecomposition of blood samples from males and females is different. Dueto lower concentration of red cells in women, the concentrations ofelements bound to red blood cells will be lower in females (Iyengar,1987).

Concentrations of trace elements by age and gender are summarizedin Table 5. We observed relatively constant concentrations of zinc andcopper in the blood of males irrespective of age. However, the bloodcopper levels of women decreased and their zinc levels increased, with

age. In linewith ourfindings, Kouremenou-Dona et al. (2006) found thehighest mean zinc concentration in serum in Greek women between 50and60years old.Morenoet al. (1999) alsoobserved a strong influence ofage on zinc level in a Spanish population.

Our findings also show that blood strontium concentrations werehigher in men over 60. However, older men have lower blood rubidiumlevels than younger men.

3.7. The inter-element correlation

With Spearman's rank correlation test we found a number ofsignificant correlations between the trace elements in the blood of thestudy population.

The statistically significant inter-element correlations in blood areshown in Table 6. Themajor correlationwas observed between Zn andRb (r=0.4034, pb0.05). Similarly to our finding, Barany et al. (2002b)found a moderate correlation between Sr and Zn levels in bloodcollected from a Swedish population.

We also observed positive correlations between Cu and Co, Zn and Srand Co and Sr. On the other hand, there was a negative correlationbetween Rb and Cu (r=−0.119, pb0.05). This negative correlationcould bedue to increaseduptakeof oneelement as a consequenceof lowlevels of another.

3.8. Seasonality of Amazonian diet

The seasonality of food consumption is one of its most outstandingfeatures in the study area (Passos et al., 2001). These authors showedthat food composition varies throughout the very different seasons,according to river's water levels. Such seasonality can be more clearlyseen for fish consumption and ecology (Lebel et al., 1997; Dolbec et al.,2001; Da Silva et al., 2006), but it also holds for consumption of othersources of animal proteins, fruits, and spices (Passos et al., 2001, 2003,2004, 2007). The cross-sectional design used for the presentinvestigation did not allow us to examine seasonal influences on theconcentrations of the various trace elements assessed, but furthersurveys should explore such issue as well as the implications forhuman nutritional status throughout the year.

4. Conclusion

The present work provides values for Cu, Co, Zn, Sr and Rb in theblood of riparian villagers living in the Brazilian Amazon. Gender wasshown to influence Zn and Cu levels. Age influenced the levels of Srand Rb in men and Cu in women. Given the scarcity of studiesexamining nutritional status in traditional communities of theAmazon, our study is the first to provide relevant insight into traceelement values in the Amazon region and inter-element interactions.This region has a diversity of foods. Some of them are consideredimportant sources of antioxidants but little is known about theirchemical composition. Moreover, dietary habits of traditional popu-lations of the Amazon basin (e.g., riparian and indigenous) arestrongly dependent on fish. Fish is the most important source ofprotein but is also an important source of mercury exposure. Analternative public health approach to reducing the possible effects ofmercury would be to identify essential elements in the bodies oftraditional populations that might interact and influence Hg absorp-tion and/or toxicity. This paper is therefore of particular importancefor future studies looking at the possible protective effects of tradi-tional riparian diets against mercury exposure and toxicity in theriparian Amazon population.

Acknowledgements

The authors are grateful to Fundação deAmparo àPesquisa doEstadode São Paulo (FAPESP), Canadian Institutes of Health Research (CIHR)

4173J.L. Rodrigues et al. / Science of the Total Environment 407 (2009) 4168–4173

and Conselho Nacional de Desenvolvimento Científico e Tecnológico(CNPq) for financial support and fellowships.

References

Angerer J, Eweres U, Wihelm M. Human biomonitoring: state of art. Int J Hyg EnvironHealth 2007;210(3–4):201–28.

Bárány E, Bergdahl IA, Bratteby LE, Lundh T, SamuelsonAG, Schutz BA, et al. Trace elementsin blood and serum of Swedish adolescents: relation to gender, age, residential area,and socioeconomic status. Environ Res 2002a;89:72–84.

Bárány E, Bergdahl IA, Bratteby LE, Lundh T, Samuelson AG, Schutz BA, et al. Trace elementlevels in whole blood and serum from Swedish adolescents. Sci Total Environ2002b;286:129–41.

Bárány E, Bergdahl IA, Bratteby LE, Lundh T, Samuelson G, Schutz A, et al. Relationshipsbetween trace element concentrations in human blood and serum. Toxicol Lett2002c;134:177–84.

Bechkoff G, Radisson J, Bessueille L, Bouchekioua-Bouzaghou K, Buchet R. Distinct actionsof strontium onmineral formation in matrix vesicles. Biochem Biophys Res Commun2008;373:378–81.

Bryan CE, Cristopher SJ, Balmer BC. Establishing baseline levels of trace elements inblood and skin of bottlenose dolphins in Sarasota Bay, Florida: implications for non-invasive monitoring. Sci Total Environ 2007;388:325–42.

Camara E, Amaro MA. Nutritional Aspect of zinc availability. Int J Food Sci Nutr2003;54:143–51.

Caroli S, Alimonti A, Coni E, Petrucci F, Senofonte O, Violante N. The assessment ofreference values for elements in human biological tissues and fluids: a systematicreview. Crit Rev Anal Chem 1994:363–98.

CDC. Centers for the Disease Control and Prevention. 2008, www.cdc.gov/nchs/nhanes.htm, accessed in January 2009.

Christensen JM. Human exposure to toxic metals— factors influencing interpretation ofbiomonitoring results. Sci Total Environ 1995;166:89-135.

Clark NA, Teschke K, Rideout K, Copes R. Trace element levels in adults from the westcoast of Canada and associations with age, gender, diet, activities, and levels ofother trace elements. Chemosphere 2007;70:155–64.

Cornelis R, Sabbioni E, Van der Venne MT. Trace element reference values in tissues frominhabitants of the European Community. VII. Reviewof trace elements in blood, serumand urine of the Belgian population and critical evaluation of their possible use asreference values. Sci Total Environ 1994;158:191–226.

Curzon MEJ, Losee FL. Strontium content of enamel and dental caries. Caries Res1977;11:321–6.

Da Silva DS, Lucotte M, Roulet M, Poirier H, Mergler D, Crossa M. Mercury in fish of theTapajós River in the Brazilian Amazon. INTERFACEHS 2006;1(1) art 6.

Dolbec J, Mergler D, Larribe F, Roulet M, Lebel J, Lucotte M. Sequential analysis of hairmercury levels in relation to fish diet of an Amazonian population, Brazil. Sci TotalEnviron 2001;271:87–97.

Gil F, Capitan-VAllvey LF, de Santiago E, Ballesta J, Pla A, Hernadez AF, et al. Heavy metalconcentrations in the general population of Andalusia, South of Spain. Acomparison with the population within the area of influence of Aznalcollar minespill (SW Spain). Sci Total Environ 2006;372:49–57.

Goullé JPM, Mahieu L, Castermant J, Neveu N, Bonneau L, Lainé Bouige D, et al. Metaland metalloid multi-elementary ICP-MS validation in whole blood, plasma, urineand hair. Reference values. Forens Sci Intern 2005;153:39–44.

Grotto D, Barcelos GRM, Valentini J, Antunes LMG, Angeli JPF, Garcia SC, et al. Low levelsof methylmercury induce DNA damage in rats: protective effects of selenium. ArchToxicol 2009;82. doi:10.1007/s00204-008-0353-3.

Hambidge M. Human zinc deficiency. J Nutr 2000;130:1344S–9S.HambidgeM. Biomarkers of trace mineral intake and status. J Nutr 2003;133:948S–55S.Heitland P, Koster HD. Biomonitoring of 37 trace elements in blood samples from

inhabitants of northern Germany by ICP-MS. J Trace ElemMed Biol 2006;20:253–62.Iyengar GV. Reference values for the concentrations of As, Cd, Co, Cr, Cu, Fe, I, Hg, Mn,

Mo, Ni, Pb, Se and Zn in selected human tissue and body fluids. Biol Trace Elem Res1987;12:263–95.

Kristiansen J, Christensen JM, Iversen BS, Sabbioni E. Toxic trace element referencelevels in blood and urine: influence of gender and lifestyle factors. Sci Total Environ1997;204:147–60.

Kosla T, Skibniewska E, Debski B, Urbanska-Slomka G. Rubidium in the trophic chainsoil–plants–animals. Trace Elem Electrolytes 2002;19:171–6.

Kouremenou-Dona E, Dona A, Papoutsis J, Spiliopoulou C. Copper and zinc concentra-tions of healthy Greek adults. Sci Total Environ 2006;359:76–81.

Kucera J, Bencko V, Sabbioni E, Van der Venne MT. Review of trace elements in blood,serum and urine for the Czeck and Slovak populations and critical evaluation oftheir possible use as reference values. Sci Total Environ 1995;166:211–34.

Lebel J, Roulet M, Mergler D, Lucotte M, Larribe F. Fish diet and mercury exposure in ariparian Amazonian population. Water Air Soil Pollut 1997;97:31–44.

Malvy DJM, Arnaud J, Burtschy B, Richard MJ, Favier A, Houot O, et al. Reference valuesfor serum zinc and selenium of French healthy-children. Eur J Epidemiol1993;9:155–61.

Minoia C, Sabbioni E, Apostoli P, Pietra R, Pozzoli L, Gallorini M, et al. Trace elementreference values in tissues from inhabitants of the European community I. A studyof 46 elements in urine, blood and serum of Italian subjects. Sci Total Environ1990;95:89-105.

Moreno MA, Marin C, Vinagre F, Ostapczuk P. Trace elements in whole blood samplesfrom residents of the city Badajoz, Spain. Sci Total Environ 1999;229:209–15.

Murrieta RSS. Food consumption among Caboclo communities on Ituqui island,Amazonia, Brazil. Am J Human Biol 1998;10:132–3.

Murrieta RSS, Dufour DL, Siqueira AD. Food consumption and subsistence in three Caboclopopulations on Marajo Island, Amazonia, Brazil. Human Ecol 1999;27:455–75.

Nardi EP, Evangelista FSB, Lemire M, Fillion M, Mergler M, Passos CJS, et al. Thecomposition of essential elements in the typical diet of the Amazonian riparianpopulation. Food Chem 2009;112:727–32.

Nielsen FH. Ultrace elements in nitrition:current knowledge and speculation. J TraceElem Exp Med 1998;11:251–4.

Palmer CD, Lewis Jr ME, Geraghty CM, Barbosa F, Parsons PJ. Determination of lead,cadmium and mercury in blood for assessment of environmental exposure: acomparison between inductively coupled plasma–mass spectrometry and atomicabsorption spectrometry. Spectrochim Acta Part B 2006;61:980–90.

Parsons PJ, Barbosa Jr F. Atomic spectrometry and trends in clinical laboratory medicine.Spectrochim Acta B 2007;62:992-1003.

Passos CJ, Mergler D, Gaspar E, Morais S, Lucotte M, Larribe F, et al. Characterization ofthe diet of a riverside population in the Brazilian Amazon [In Portuguese]. RevSaúde Ambiente 2001;4(1/2):72–84.

Passos CJ, Mergler D, Gaspar E, Morais S, Lucotte M, Larribe F, et al. Eating tropical fruitreduces mercury exposure from fish consumption in the Brazilian Amazon. EnvironRes 2003;93:123–30.

Passos C,Mergler D, Larribe F. Response to “fruits,fish, andmercury: further considerations”.Environ Res 2004;96:102–8.

Passos CJ, Mergler D, FillionM, LemireM,Mertens F, Guimarães JRD, et al. Epidemiologicconfirmation that fruit consumption influences mercury exposure in ripariancommunities in the Brazilian Amazon. Environ Res 2007;105:183–93.

Pennington JA, Schoen SA, Salmon GD, Young B, Johnson RD, Marts RW. Composition ofcore foods of the U.S. food supply,1982–1991. III. Copper, manganese, selenium, andiodine. J Food Comp Anal 1999;58:171–217.

Prasad S.Zincdeficiency inhumans: aneglectedproblem. J AmCollNutr 1998;17(6):542–3.Rosborg I, Hyllén E, Lidbeck J, Nihlgard B, Gerhardsson L. Sci Total Environ 2007:385–90.Salgueiro MJ, Zubillaga M, Lysionek A, Sarabia MI, Caro R, De Paoli T, et al. Zinc as an

essential micronutrient: a review. Nutr. Res. 2000;20:737–55.Schuhmacher M, Domingo JL, Corbella J. Zinc and copper levels in serum and urine:

relationship to biological, habitual and environmental factors. Sci Total Environ1994;148:67–72.

Shahnazari M, Sharkey NA, Fosmire GJ, Leach RM. J Bone Miner Res 2006;21:1696–703.Templeton DM, Sunderman FW, Herber RFM. Tentative reference values for nickel

concentrations in human serum, plasma, blood and urine: evaluation according tothe TRACY protocol. Sci Total Environ 1994;148:243–51.

Terrés-Martos C, Navarro-Alarcón M, Martín-Lagos F, López G, López-Martinez MC.Determination of copper levels in serum of healthy subjects by atomic absorptionspectrometry. Sci Total Environ 1997;198:97-103.

Turnlund JR. Human whole-body copper metabolism. Am J Clin Nutr 1998;67:960S–4S.White MA, Sabbioni E. Trace element reference values in tissues from inhabitants of the

European union. X. A study of 13 elements in blood and urine of a United Kingdompopulation. Sci Total Environ 1998;216:253–70.

Zanao RA, Barbosa F, Souza SS, Krug FJ, Abdalla AL. Direct determination of selenium inwhole blood by electrothermal atomic absorption spectrometry using W–Rh-coatedplatform and co-injection of Rh as thermal stabilizer. Spectrochim Acta2002;57:291–301.