www.elsevier.com/locate/foodcont

Food Control 17 (2006) 719–726

Quality assurance of volumetric glassware for the determinationof vitamins in food

I. Castanheira a,*, E. Batista b, A. Valente a, G. Dias a, M. Mora b, L. Pinto b, H.S. Costa a

a Department of Food Safety and Nutrition, National Institute of Health Dr. Ricardo Jorge, Av. Padre Cruz, 1649-016 Lisboa, Portugalb Central Metrology Laboratory, Portuguese Institute for Quality Rua Antonio Giao, 2 Caparica, Portugal

Received 2 December 2004; received in revised form 7 April 2005; accepted 11 April 2005

Abstract

Volumetric measurements are currently used in the analytical process to determine the vitamins content in foods. The variationsin stability and integrity of volumetric apparatus, although relatively small when compared with other sources of error, should bemonitored. The main purpose of this study was to investigate and identify a number of variables which, in daily use, can affect thelife cycle of volumetric glassware. The role of in-house metrological hierarchy, the operator�s performance and the effect of temper-ature was evaluated for single volume flasks, single- and multi-mark pipettes. Biased nominal capacity was applied in the calculationof water-soluble vitamins (B1, B2, B6, niacin) and fat-soluble vitamins (A, D, E) content. Persistent bias due to volumetric glass-ware, can reach 11%, in the results obtained for different vitamins content. The largest bias was observed for vitamin B1, perhapsdue to the high amount of volumetric glassware used in the analytical procedure. This is an indication that the cost–benefit of acalibration plan should be evaluated in a rational way. The regular checking of volumetric glassware for stability and integrity isdiscussed on the light of the latest revision of ISO 17025.� 2005 Elsevier Ltd. All rights reserved.

Keywords: Volumetric glassware; Bias; Vitamins; Food; Metrology in chemistry

1. Introduction

Quality assurance of volumetric glassware is a rele-vant step of the food analytical process (Anglov, Peter-sen, & Kristiansen, 1999; Wood, Nilsson, & Wallin,1998), and there are cases where volumetric methodsare adopted as CEN methods: Foodstuffs––Determina-tion of vitamins by high performance liquid chromatog-raphy (HPLC), (EN 12821; EN 12822; EN 14164; EN14152; EN 12823-1; EN 14122; CEN/TC 275/WG 9).Calibrated glassware and traceability to SI units is

0956-7135/$ - see front matter � 2005 Elsevier Ltd. All rights reserved.doi:10.1016/j.foodcont.2005.04.010

* Corresponding author. Tel.: +351 21 751 93 17; fax: +351 21 7526400.

E-mail address: isabel.castanheira@insa.min-saude.pt (I. Castan-heira).

now explicitly requested in guidelines dealing with foodmeasurements (Codex, 2002; EURACHEM, 2000).

The quality criteria of volumetric glassware has beenstudied in recent decades and granted through the use ofmetrological procedures, supported on traceability ofnominal value and its confidence interval. ISO (ISO835/1; ISO 4787) in the early 1980s published a seriesof standards where the concept of tolerance limits basedon repeatability of the calibration procedure was intro-duced. Studies on stability of volumetric glassware weredescribed later in the 1980s by Burfield and Hefter(1987). These concepts have recently been reviewed inthe literature (EURACHEM, 2000; Kadis, 2004): Thetolerance limits are calculated in compliance with theGuide ‘‘Quantifying Uncertainty in Analytical Measure-ment’’, issued by EURACHEM/CITAC (2000); volu-metric glassware is considered appropriate if the

720 I. Castanheira et al. / Food Control 17 (2006) 719–726

nominal capacity value and its associated uncertainty re-main within acceptable limits. In the case of volumetricglassware used in daily measurement practice, the expirydate of the calibration result, calibration plans andcalibration intervals are defined by the laboratory andusually operation conditions are not mentioned.

However, this procedure tends to change in the futuredue to improved actions introduced by accreditationbodies, as consensus exists, in the revision of ISO/IEC17025 to include regular checking of equipment forstability and integrity (FDIS ISO 17025, 2004).

Confidence in the whole life cycle of the measurementequipment, from its design to use, framed by metrolog-ical parameters is a thematic area that has receivedmuch attention in the field of legal metrology. Substan-tial information on this area exists in OIML D 20(1988), OIML D 27 (2001), Vaz, Castanheira, Fino,and Van der Veen (2000), Sommer, Chappell, andKochsiek (2001). The equipment is considered appropri-ate during the time in which deviation from the truevalue remains under acceptable limits. The checkingintervals for the equipment in place are designated asa periodic verification, and the decisions on equipmentperformance are based on maximum permissible errors,as described in OIML recommendation.

This work is a three-step procedure dealing withmonitoring of volumetric glassware used in the measure-ment of vitamins content in food in compliance with thelatest revision of ISO 17025 (FDIS ISO 17025, 2004). Inthe first step, an approach to establish in-house metro-logical hierarchy is discussed. The identification of crit-ical parameters in the calibration procedure is presentedin the second step. Finally, in the third step, the impactof systematic error due to volumetric glassware in themeasurement of vitamins B1, B2, B6, niacin, A, D andE content was estimated.

2. Materials and methods

2.1. Metrological qualification of volumetric glassware

2.1.1. Metrological hierarchy for volumetric glassware

Laboratory glassware was classified according to itsmetrological grade. This was based on an establishedhierarchy of traceability chain to SI units through na-tional standards of measurements and framed by stan-dard definitions adopted by the InternationalVocabulary of Metrology (VIM) (BIPM, 1993). The fol-lowing Reference Standards Volumetric Glassware(RSVG) were calibrated at the National MetrologyInstitute (NMI): burettes, pipettes, flasks and cylinderswith capacities in the range of 5–500 ml, and storedunder appropriate conditions.

Working Standards Volumetric Glassware (WSVG)(pipettes of 2 ml and 5 ml; flasks of 20 ml and 50 ml) were

calibrated against reference standards at a Food Analy-sis Laboratory (FAL) using an appropriate protocol.

The nominal value of volumetric glassware, routinelyused in food analysis, was checked against the assignedvalue of the working standard. A set of three or five20 ml and 50 ml flasks and 1 ml, 2 ml and 5 ml pipettescurrently used in vitamin content determinations weresubmitted to several tests described below.

2.1.2. Metrological criteria

Class A volumetric glassware was calibrated at theNMI and calibration results issued by the NMI wereanalysed in terms of the assigned value and associateduncertainty. The acceptance criterion was definedaccording to ISO 4787 (1984). Compliance of the as-signed value with the acceptance criteria is defined byEq. (1) and it was evaluated on the basis of acceptedor rejected.

xn � xrj jffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiU 2

n þ U 2a

q < TL ð1Þ

where xn is the nominal value, xr is the reference value,Un is the uncertainty of the nominal value, Ua is theuncertainty of the assigned value and TL tolerancelimits defined in ISO 1042 and ISO 835.

Volumetric glassware was accepted as reference stan-dard, when the assigned value and associated uncertaintywere within the acceptance limits. The glassware was des-ignated as WSVG when the assigned value plus or minusthe associated uncertainty lies outside one of the accep-tance limits. The glassware was rejected as RSVG, whenthe assigned value and associated uncertainty did not fallin the previous judgement of compliance. In order toprovide confidence of measurements, a calibration pro-gramme was developed and RSVG were calibrated oncea year by the NMI. The calibration results for time zero,years two and four were compared with the maximumacceptable deviation defined by Eq. (1), and used toestablish the external calibration intervals.

2.2. Calibration procedure

2.2.1. Traceability

The in-house calibration hierarchy used to achievetraceability to SI units was composed of: RSVG, WSVGand volumetric glassware. The reference value of capac-ity was obtained by a gravimetric process at the NMI,according to ISO 4787 (1984), ISO 835 (1981) and ISO1042 (1998). This was achieved applying a weighing pro-cedure consisting of 10 weighing cycles, where in eachcycle, the mass corresponding to volume capacity wasobtained by the difference between the filled vessel withdistilled water and the empty vessel. The equation ofproportionality that converts the average mass tovolume measurement is described as:

I. Castanheira et al. / Food Control 17 (2006) 719–726 721

V 20 ¼ ðIL � IEÞ �1

qw � qA

� �� 1� qA

qB

� �� ð1� cðt � 20ÞÞ ð2Þ

where V20 is the volume at 20 �C (reference tempera-ture), in ml; IL and IE, the weight of the vessel with waterand of the empty vessel, respectively, in g; qw, the den-sity of water, at calibration temperature, in g ml�1; qA

and qB, the density of air and of the mass pieces, ing ml�1 (reference values of 0.0012 g ml�1 and 8.0 g ml�1,respectively); c, the coefficient of cubic thermal expan-sion of the material, in �C�1 and t, the temperature ofthe water used in the calibration, in �C.

2.2.2. Measurement uncertainty

The uncertainty of the glassware calibration processis defined as a parameter associated with the result ofa measurement that characterizes the dispersion of thevalues that could be attributed to the measurand,according to EURACHEM Guide (2000). The expres-sion for the combined standard uncertainty (u(V20))was developed from Eq. (2) using the error propagationformula. The major contributions to the combined stan-dard uncertainty were estimated through a functionmodel, presented in Eq. (3), and considering seven build-ing blocks: repeatability mass; meniscus; water density;temperature; air density; density of mass species andthermal expansion of the material. The expanded uncer-tainty (U) defined ‘‘as an interval within which the valueof the measurand is believed to lie with a particular levelof confidence’’ EURACHEM (2000) was calculatedusing a coverage factor of k = 2, considering 95% asapproximate level of confidence.

uðV 20Þ ¼oV 20

om

� �2

u2ðmÞ þ oV 20

oqw

� �2

u2ðqwÞ"

þ oV 20

oqA

� �2

u2ðqAÞ þoV 20

oqB

� �2

u2ðqBÞ

þ oV 20

oc

� �2

u2ðcÞ þ oV 20

ot

� �2

u2ðtÞ þ u2 dV menð Þ#1

2

ð3Þ

where u(m) is the uncertainty of the weighing; u(qw), theuncertainty of water density; u(qA), the uncertainty of airdensity; u(qB), the uncertainty of the mass pieces; u(c),the uncertainty of the coefficient of cubic thermal expan-sion of the material; u(t), the uncertainty of the temper-ature and u(dVmen), the uncertainty of the meniscus.

2.3. Operator’s performance

The influence of the operator�s performance on thecapacity value of WSVG was evaluated through a cali-bration test, using the same experimental procedure de-

scribed above. The ‘‘blind’’ test was carried out by ananalyst with less experience in the calibration procedure,and consisted on the evaluation of results at the gradu-ation line under study, taking into account three param-eters: (a) reported value and associated uncertainty; (b)precision and (c) meniscus reading. The ‘‘blind’’ resultswere benchmarked to the ones given by a NMI operatorand adequate statistical tests were applied. The criteriaused to classify the operator�s performance were: poorand good performance. This was derived from the sumof the overall results. Each analyst result was consid-ered: satisfactory; questionable or unsatisfactory, if thevalue was respectively, below, equal or above the tabu-lated value of the statistical test. The operator�s perfor-mance was considered as good, if 75% of the values ofthe overall result were satisfactory. The remaining situ-ations were considered as poor performance. This proce-dure was applied to two types of volumetric glassware:pipettes and flasks of different capacities.

2.4. Temperature conditions

The effect of temperature on the nominal capacityof volumetric glassware was carried out at NMI follow-ing a three step experimental protocol: first, the glass-ware was calibrated according to the calibrationprocedure described above; secondly, the glasswarewas submitted to a chosen temperature during a periodof time and in the third step, the glassware was equili-brated at standard reference temperature and then sub-mitted to the calibration procedure again. The effect oftemperature is given by the difference between the vol-ume at graduation line obtained in the first and thirdstep.

Pipettes and flasks were withdrawn from daily rou-tine laboratory work, divided into three different groupsand submitted to this procedure. In group I, the glass-ware was submitted daily to a temperature of 4 �C dur-ing five days (120 h). In group II, the temperature wasset daily (24 h) to 50 �C during five days and the calibra-tion procedure was performed at day one, three and five.In group III, the glassware was maintained during fivedays at 50 �C and submitted to the calibration proce-dure at day five.

2.5. Testing for persistent bias

2.5.1. Determination of vitamin content

Vitamin determination was done by conventionalmethodology. This consisted on saponification for fat-soluble vitamins and hydrolysis for water-soluble vita-mins, followed by liquid–liquid extraction and HPLCanalysis according to the methods described in the liter-ature: Vitamins A (EN 12823-1), D (EN 12821), E (EN12822) and vitamins B1 (EN 14122), B2 (EN 14152), B6(EN 14164), niacin (CEN/TC 275/WG 9).

Fig. 1. Calibration results of volumetric glassware as referencestandards.

UAL

xr

Reference standard

Working standard

722 I. Castanheira et al. / Food Control 17 (2006) 719–726

2.5.2. Bias due to volumetric glassware

The potential bias of the analytical procedure, due tolaboratory volumetric glassware was studied using twoprotocols (A) and (B), and assuming that no othervariation occurred in the measuring system during thedetermination of vitamin content. In protocol (A), mea-surement results were obtained according to standardoperation procedure. In protocol (B) and for each stepof the analytical procedure, the ‘‘nominal value’’ ofvolumetric glassware was estimated by adding to thereference value, the worst value within experimentalvariability for the operator and temperature tests. Biaswas considered significant, if the relative differencebetween protocol (A) and (B) was larger than theexpanded uncertainty of this difference.

2.6. Statistical analysis

HPLC response characteristics were estimatedthrough calibration functions obtained by least-squaresregression analysis. The following parameters were con-sidered: polynomial coefficients and their uncertainties,residuals and regression coefficients. Mean and standarddeviations of analytical data for vitamins content werecalculated for a significance level of P < 0.05.

The following statistical tests were applied for dataanalysis: F-test was used to compare variances of thereplicate analysis, En-score was used to compare twoindependent means with the same number of observa-tions and single outlier selection was done by Dixon�stest (Miller & Miller, 1988).

0 2 4

Time (years)

LAL

Fig. 2. Examples of calibration results with reference value andassociated uncertainty: xr, reference value; UAL, upper acceptablelimit; LAL, lower acceptable limit.

3. Results and discussion

An overview of calibration results for RSVG is givenin Fig. 1. It can be seen that for different types ofvolumetric glassware, the percentage of glassware cali-brated that failed the acceptance criteria varied from0% up to 40%. Cylinders are in agreement with theacceptance criteria defined above; 10% of the flasks cal-ibrated do not fulfill this criterion; 20% of burettes can-not be labeled as reference standards and for pipettes,40% of the candidate standards were not compliant withthe specifications of the acceptance criteria as referencestandards (BIPM, 1993). Calibration results issued forpipettes as RSVG are presented graphically in Fig. 2,for time zero, year two and four, revealing that appro-priate care of volumetric glassware was taken. The samegraphic profile was obtained for flasks. These results,indicating that nominal capacity remained constantovertime is probably due to consistent storing and clean-ing conditions as described by accreditation bodies(UKAS, 2002). For working standards more than 90%of pipettes and flasks passed the defined acceptancecriteria.

An indirect calibration procedure (Rodrıguez, Gra-cia, Lopez, & Sanchez, 2001) which consists on the con-version of the mass into volume and comparison of thedeliverable water of RSVG and WSVG, on a periodi-cally time schedule of six months and under the assump-tion that measurement uncertainty remains constant,was also used.

The establishment of in-house metrological hierarchybased on an indirect calibration procedure of WSVG,with more rigorous specifications than the manufacturetolerances, allows regular checking of volumetric glass-ware for stability and integrity of daily used glassware

Fig. 3. Schematic representation of sources and main components ofmeasurement uncertainty, for the calibration procedure, performedagainst national standards.

I. Castanheira et al. / Food Control 17 (2006) 719–726 723

without lost of traceability links. This procedure is moreadvantageous than to refer all volumetric glassware toan external link (European Commission, 1998; EURA-CHEM, 2003).

Evaluation of the measurement uncertainty, in thecalibration procedure of reference standards, was basedon a model which consists on the contribution of severalstandard uncertainties, quantified in terms of experi-mental standard deviation times the sensitivity coeffi-cient of each source, as described in Eq. 3. Thisempirical model was evaluated by the Ishikawa (1976)diagram and based on type A (experimental) and typeB (estimated) uncertainties inputs which combinesintralaboratory repeatability and reproducibility withparameters (e.g.: water density; thermal expansion coef-ficient) whose contributions derive from tabulatedvalues published in the literature (Spieweck & Bettin,1992). All the values obtained for the expanded uncer-tainty were below the predicted uncertainties of 0.1%;0.2%; 0.3% (UKAS, 2002). The values of referencestandards were well within the variation claimed bythe manufacture (Noack, 1998) and in agreement withthe results of Euromet intercomparisons (Euromet,2004). The uncertainty budget provides a mean to com-pare different contributions to the combined standarduncertainty (Fig. 3). The largest contribution came fromexperimental sources: mass determination, meniscusreading, temperature, precision of each determinationwhich includes the operator�s performance and whichwas found to be the highest source. Therefore, the oper-ator�s performance was accounted for and was subject toa more detailed study.

The acceptable operator�s performance was evaluatedapplying appropriate statistical tests to identify the maincauses of error. Table 1 summarizes the results obtainedby analysts from the National Metrology Institute(ANMI) and from a Food Analysis Laboratory (AFAL)for flasks and pipettes, and for the same calibration pro-cedure. Outliers were not found when Dixon�s test wasapplied to ANMI or AFAL results.

Significant differences between the results given bythe two analysts ANMI and AFAL were detected for bothtypes of volumetric glassware. jEn-scorej > 1 was ob-served for flasks and pipettes indicating that the results

Table 1Overview of statistical tests, applied to evaluate operator�s results

Flask En-score F-test P

20 ml 50 ml 20 ml 50 ml

#1 �22.5 �26.4 9.00 4.00 ##2 14.8 15.6 1.00 1.00 ##3 3.1 �7.4 4.00 1.00 ##4 �22.6 10.7 1.00 4.00 ##5 �4.5 �9.9 4.00 1.00 #

Criteria used to classify operator�s results: jEn-scorej > 1, unsatisfactory; jEn

provided by AFAL are different from the assigned value(Armishaw, King, & Millar, 2003). The analyst�s preci-sion (ANMI versus AFAL) was evaluated using F-testand the results were compared with the tabulated values(Miller & Miller, 1988) demonstrating that AFAL hasbeen working with a different level of precision. Theevaluation of operator�s performance was grounded ona measurement theory (Mari, 2003), implemented withtwo kinds of tests: pair comparison and magnitude esti-mation using a set of reference values in all points ofscale (Rossi, Crenna, & Codda, 2003). The judgementof evaluation occurs in three steps: (a) single measure-ment through outliers tests; (b) error propagation dueto statistical evaluation of mean of results (c) estimationdegree of inaccuracy by null hypothesis. This approachseems to be very useful to evaluate the analyst skills,to identify the cause of error and to decide on furthertraining programmes. This is helpful not only duringchecking and calibration procedure, but also in the over-all analytical procedure, since a large number of vita-mins analysis measurements are based on volumetricmeasurements (Klejdus et al., 2004; Sharpless, Margolis,& Thomas, 2000).

ipette En-score F-test

2 ml 5 ml 2 ml 5 ml

1 1.3 �6.7 1.00 4.002 2.5 �2.4 9.00 1.003 4.2 �3.1 4.00 2.254 – �2.8 – 1.005 – �0.6 – 9.00

-scorej = 1, questionable; jEn-scorej < 1, satisfactory; F0.95(9,9) = 3.18.

724 I. Castanheira et al. / Food Control 17 (2006) 719–726

Deviation from the reference value due to tempera-ture and exposure intervals is presented in Fig. 4. Thenominal capacity did not change when volumetric glass-ware was maintained at 4 �C. These results are expectedfor borosilicate material (Burfield & Hefter, 1987). Butat 50 �C, significant effects were noticed, depending onthe type of volumetric glassware, capacity and exposureperiod. The highest deviation from reference value wasobserved at day five for both types of volumetric glass-ware and the largest effect was observed for pipettes of1 ml (Fig. 5). The results in the present study show thattemperature effect is different according to temperaturevalue, period of time, type and capacity of volumetric

-0.014

-0.010

-0.006

-0.002

0.002

0.006

Time (days)

Dev

iatio

n fro

m re

fere

nce

valu

e (%

)

4 ºC

50 ºC

51 3

Fig. 4. Results of temperature tests for graduated pipettes as afunction of time.

0

0.1

0.2

0.3

0.4

Capacity of pipette (ml)

Dev

iatio

n fro

m re

fere

nce

valu

e (%

)

2 5 1

Fig. 5. Temperature effect on nominal capacity of graduated pipettesafter five days at 50 �C.

glassware. Burfield and Hefter (1987) has concluded thatfor oven drying borosilicate glassware, the calibrationvalue is not significantly affected. Our results are notin agreement with Burfield�s conclusion possibly becausethe calibration methods and the uncertainty calculationsapplied here are different. In this study, uncertainty cal-culations and the calibration process were determinedthrough a metrological approach which is more rigorousto investigate differences between or within measure-ments results (De Bievre & Taylor, 1997).

In daily use, machine washing and oven or air dryingof glassware at moderate temperatures (around 60 �C)are unavoidable, for volumetric glassware used withcomplex food matrices. Therefore, the glassware shouldbe submitted to a qualification performance test regu-larly (Bedson & Sargent, 1996; UKAS, 2002).

Fig. 6 shows the mean bias due to volumetric glass-ware in the determination of vitamins content. Persistentbias (Taverniers, De Loose, & Bockstaele, 2004), is de-fined as the difference between the expectation of the testresults (protocol B) and the reference value (protocolA). Potential bias due to volumetric glassware wasinvestigated in all steps of the analytical procedure forthe determination of vitamins content and the largestcontribution to bias was observed for the calibrationcurve and extraction step.

The operator�s performance and temperature effectson volumetric glassware are the factors that influencethe most the nominal capacity. This enabled us todemonstrate that lack of accuracy can overestimate thevalue of vitamins content, in the range of 1–11%, exceptfor Vitamin D where the results are unbiased. Thispattern of results strongly suggests the influence of

Fig. 6. Influence of bias due to volumetric glassware on reported valueof vitamin concentration. The largest source comes from low capacityglassware.

I. Castanheira et al. / Food Control 17 (2006) 719–726 725

systematic errors in the preparation of the calibrationcurve. Vitamin D was quantified using an internal stan-dard and for the other vitamins, the content was calcu-lated by regression analysis using a calibration curvewith five external standards, which were prepared by adilution procedure. The largest bias was observed forvitamin B1, perhaps due to the high amount of volumet-ric glassware used in the analytical procedure, especiallyduring the preparation of calibration curve. For vita-mins A, E and B2, bias is introduced by low capacityvolumetric glassware (pipettes and flasks) and for vita-min B6 bias is due to pipettes of 2 ml and 5 ml. Theresults obtained when a small number of flasks and pip-ettes are used did not differ significantly in both proto-cols, while those obtained when a high number of lowcapacity pipettes are used were systematically biased.From this study, we conclude that a cumulative effectof error in the dilution steps for the preparation of thecalibration curve is probably associated with the lackof fit of linearity and can contribute significantly to lessaccurate results. This study has implications concerningroutine work in the determination of vitamins content infood and was motivated by the need of traceability andcomparability of results. Differences ranging from 10%to 40% for the value of vitamin A, E, B1, B2 and B6content have been reported by food analysis laborato-ries and are largely connected with the expertise levelof participants (Part 1: Hollman et al., 1993a; Part 2:Hollman et al., 1993b; Mathiasson et al., 2002). In thedetermination of Vitamin A, in serum, the same conclu-sion was reported (Hulshof, Brouwer, Burema, & West,2002). Although variations in stability and integrity ofvolumetric glassware can be relatively small when com-pared to other sources of error, they need to be properlyevaluated as defined by item no. 5.9.1, in the newrevision of ISO 17025 (FDIS ISO 17025, 2004), andare necessary to improve collaborative tests results andinter-laboratory comparability. This work raises theproblem of correlating the nominal capacity variationwhich occurs in daily used volumetric glassware withthe overestimation content of vitamins. Therefore, ourresults can contribute to improve the evaluation oflaboratory performances and comparability of resultsas recommended by Codex Alimentarius (Codex, 2002).

4. Conclusions

The present methodology evaluates the factors affect-ing the accuracy of volumetric glassware. This approachis robust and feasible, thus, enabling its adoption infood analysis laboratories.

This study highlights the critical control points of lab-oratory volumetric glassware addressing the need ofperiodic check intervals and calibration in the measure-ment plan.

Furthermore, collaboration between food analysisand metrological laboratories, reveals to be very useful:(a) to provide support for education and training of per-sonnel; (b) to improve the traceability of working levelby increasing the quality of link to SI units of assignedvalues and associated uncertainties; (c) to support theorganization of inter-laboratory comparisons, a tool toassess the ability of food laboratories to generate dataquality that may be utilized in aims such as establish-ment of food/disease-health-status using differentsources of analytical data.

Acknowledgments

The authors would like to thank Dr. Maria AntoniaCalhau, Head of Department-INSA and to Engo Anto-nio Cruz, Director of IPQ/LCM for encouragement ofthis work, to Dr. Luısa Oliveira and Enga Eduarda Fil-ipe, MD for providing laboratory facilities and to Hel-ena Bartolo for vitamins analysis. This work waspartially supported by PIDDAC/IPQ/LCM/Universid-ades (1997–2004) and POCTI /CTA/95/2001.

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