Talanta 77 (2009) 1155–1159

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Talanta

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A portable, inexpensive and microcontrolled spectrophotometer basedon white LED as light source and CD media as diffraction grid

Germano Verasa,b, Edvan Cirino Silvaa,∗, Wellington Silva Lyraa,Sófacles Figueredo Carreiro Soaresa, Thiago Brito Guerreiroa,Sérgio Ricardo Bezerra Santosc

a Universidade Federal da Paraíba, Departamento de Química, 58.051-970, João Pessoa, PB, Brazilb Universidade Estadual da Paraíba, Departamento de Química, 58.109-753, Campina Grande, PB, Brazilc Centro Federal de Educacão Tecnológica da Paraíba, 58.015-430, João Pessoa, PB, Brazil

a r t i c l e i n f o

Article history:Received 17 April 2008Received in revised form 18 August 2008Accepted 19 August 2008Available online 27 August 2008

Keywords:White LEDCD mediaProgrammable interrupt controller (PIC)

a b s t r a c t

A portable, microcontrolled and low-cost spectrophotometer (MLCS) is proposed. The instrument com-bines the use of a compact disc (CD) media as diffraction grid and white light-emitting diode (LED) asradiation source. Moreover, it employs a phototransistor with spectral sensitivity in visible region asphototransductor, as well as a programmable interrupt controller (PIC) microcontroller as control unit.The proposed instrument was successfully applied to determination of food colorants (tartrazine, sun-set yellow, brilliant blue and allura red) in five synthetics samples and Fe2+ in six samples of restorativeoral solutions. For comparison purpose, two commercial spectrophotometers (HP and Micronal) wereemployed. The application of the t-paired test at the 95% confidence level revealed that there are not sig-nificant differences between the concentration values estimated by the three instruments. Furthermore,a good precision in the analyte concentrations was obtained by using MLCS. The overall relative standard

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Visible spectrophotometerFood colorantsIron (II)

deviation (R.S.D.) of each analyte was smaller than 1.0%. Therefore, the proposed instrument offers an eco-nomically viable alternative for spectrophotometric chemical analysis in small routine, research and/or

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. Introduction

Since the seventies of the last century, when researchers aslaschka et al. [1] developed photometers based on light-emittingiode (LED), LEDs have been applied on spectrophotometric instru-entation as radiation sources having relatively small effective

andwidth [2,3]. However, the increasing demand for chemicalnalyses makes necessary to develop LED-based instruments toork in a wide range of wavelengths what requires multiple

adiation sources. The approach increases cost and complexityf the optical and electronic system. For example, optical fibersre usually employed to direct the radiation towards the detectornd a more elaborated hardware is necessary to control the LED

rift.

Nowadays, it is easy to acquire white LEDs that substituteshe complex optical system obtained when monochromatic LEDsre used as sources of several wavelengths in the visible region.

∗ Corresponding author.E-mail address: edvan@quimica.ufpb.br (E.C. Silva).

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039-9140/$ – see front matter. Published by Elsevier B.V.oi:10.1016/j.talanta.2008.08.014

ts components are inexpensive and of easy acquisition.Published by Elsevier B.V.

herefore, it is possible to develop simple spectrophotometerssing a single white LED with an adequate disperser of thehite light. For this purpose, the literature reports two works

nly that employ white LED as radiation source [4,5]. Shimazakit al. [4] were pioneers in the development of a white LED-ased spectrophotometer which was used in determination ofe2+ in river water by the o-phenantroline method. Li et al. [5]escribed a portable multi-function instrument using differentpectrophotometric techniques. As application, the spectropho-ometric determination of Cr6+ and Al3+ was carried out usingiphenylcarbazide and pyrocatechol violet as chelant and chro-ogenic reagents, respectively.Commercial spectrophotometric instrumentation commonly

mploys absorption or interference optical filters, prisms or diffrac-ion grids as radiation dispersers. In most of these instruments, theispersive device is a diffraction grid, an optical component con-

aining series of grooves traced on a glass plate or polished metal. Inhis context, a compact disc (CD) media could be used as diffractionrid due to its grooves. Despite the use of CD media as radiation dis-erser [6], no application in spectrophotometric instrumentationas been found.

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In the present work, a portable, microcontrolled and low-ost spectrophotometer (MLCS) having a white LED as radiationource and a CD media as diffraction grid for measurementssing the visible region was developed. In order to increase theignal-to-noise ratio in the spectrophotometric measurements, ahototransistor with maximum spectral sensitivity in the visibleegion was used. In the MLCS, pulsed radiation from the whiteED is dispersed by the CD media generating monochromaticadiation that is focalized in the phototransistor by a stepperotor. The control of the proposed instrument, the acquisition

nd treatment of data are accomplished by an electronic circuitased on a programmable interrupt controller (PIC) microcon-roller. Two analytical applications were elected to illustrate theerformance of the MLCS. The first is represented by individualeterminations of four food colorants (tartrazine, brilliant blue,llura red and sunset yellow). The second concerns to the deter-ination of Fe2+ in restoratives by using the 1,10-phenantrolineethod.

. Experimental

.1. Reagents, solutions and samples

Food colorants tartrazine (E-102), sunset yellow (E-110), alluraed (E-129), and brilliant blue (E-133) were purchased fromigma–Aldrich. Stock solutions of 1000 mg L−1 of each colorantere prepared dissolving its necessary amount in phosphate buffer

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ig. 2. Electronic layout of the MCLS detector. Legend: PT = phototransistor; C = polyethylemplifier; U3 = quadruple switches. Details in text.

Fig. 1. Schematic diagram illustrating the optical system of the MLCS.

olution at pH 7.00. Standard solutions of synthetic samples ofach colorant were prepared by the appropriated dilution of theespective stock solution. The standard solutions were prepared inuthentic triplicates at the concentrations 2.0, 4.0, 6.0, 8.0, 10.0,2.0, 14.0, 16.0, 18.0, 22.0, and 26.0 mg L−1.

FeCl3·6H2O was used to prepare stock solutions of Fe3+

000 mg L−1. Working solutions (Fe2+ 2.0–10.0 mg L−1) were pre-

ared from the stock solution after reduction of the Fe3+ usingscorbic acid 1.0% (w/v). Six samples of restorative oral solu-ions (containing Fe2+) from different marks were acquired inrugstores at João Pessoa city, Brazil. These pharmaceutical for-

ne capacitors; R = resistors; U1 = operational amplifier; U2 = quadruple operational

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ulations were diluted to fit in the linear range of the analyticalurve.

Recently distilled water was used to prepare all solutions.

.2. Description of the proposed instrument

Fig. 1 illustrates the optical components employed in the MLCSnd Fig. 2, the electronic layout of the detection unit. This instru-ent was assembled into a box with 30 cm front, 20 cm height and

0 cm length. The optical–mechanical system is composed by whiteED, focus lens to create a small light image (2 mm diameter) onhe diffraction grid (CD) and stepper motor with gear to move theiffraction grid and to promote the scan of the spectral region of

nterest. To increase the spectral resolution, the position of the step-er motor was adjusted so that the phototransistor can detect theadiation from the second overtone.

The instrument uses a PIC 16F877 programmable microcon-roller as control unit driven by a stabilized power supply. Theontrol software was elaborated with C language. The microcon-roller presents a 10-bit multi-channel analog to digital converterA/D), 8k × 14 words of flash program memory, 256 × 8 bytes ofEPROM data memory, 368 × 8 bytes of RAM, three timers andhree ports with eight channels (multiplexed pins) each, for gen-ral purpose, that can be configured for data acquisition. Theicrocontroller allows drastic reduction of the number of elec-

ronic components in the spectrophotometer and makes possibleo introduce or alter functionalities in the instrument without anyardware modification.

The white LED Control Module pulses the white LED radiationt 7 Hz. The module turns the white LED on for readings of the ana-ytical signals and off for determination of instrumental noise. Theignals are detected, filtered and amplified in the Detection Modulef the MLCS, showed in Fig. 2. In the layout, five operational ampli-ers are used for different purposes: to convert the photocurrent

nto voltage (U1), to filter the high frequency noise (U2:A) as a high-ass Butterworth filter; to amplify the analytical signal (U2:B), toetect the peak signal as a sample-hold unit (U2:C) and to fit thenalytical signal (U2:D) in the A/D scale (5 V) using CMOS switchesU3:B–D) to change the gain depending on the value of the input sig-al. C5 and R11 are used as low-pass filter and P1 is used for off-set.MOS switch U3:A is used to discharge the capacitor of the sample-old circuit. The switches are controlled by the PIC. The processedignal E is sent to the A/D converter of the PIC to be converted intobsorbance values. To calculate the absorbance values, the loga-ithm function used the measurement of blank (water) as reference.oth blank and sample measurements were registered turning theED on to consider maximum light power and off in order to con-ider the dark signal. After digitalization, the absorbance values areent by the microcontroller to the liquid crystal display (LCD) mod-le, a TECH2004D-FL-GBS-S character type LCD with two buses:ight bits for data and three bits for control. In this work, the com-unication between microcontroller and LCD was always carried

ut with six bits, four being used for data transmission and twoor data control. The Stepper Motor Control Module controls thengular displacement of the CD media.

A clock elaborated with a 4-MHz piezoelectric crystal is used forynchronization of the microcontroller internal functions.

. Results and discussion

.1. Calibration of the MLCS optical–mechanical system

An ocean optics spectrometer, model USB450, which substitutedhe phototransistor as detector, was used to calibrate the MLCS

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ptical–mechanical system. Since the slit for the light entrancen the probe and the sensitive area of the phototransistor havehe same dimensions (ca. 0.3 mm), the spectra registered by thecean optics spectrometer represent the profile of the radiation thateaches the phototransistor. In this process, the spectrometer reg-sters the reflected spectrum resulting from the radiation diffractedn the CD media. It was carried out five scans to obtain an averagepectrum for each step of the stepper motor. Average spectra weresed to build a calibration curve between the number of steps andhe wavelength of maximum emission of the spectrum. In this case,n equation was estimated by using the linear least-squares and theesult is presented below.

tep = wavelength × 0.50246 − 214.7837

The number of steps and peak wavelengths are linearly cor-elated in the spectral range 440–640 nm with a correlationoefficient of 0.99968. Therefore, practically the entire visiblepectral region can be exploited for spectrophotometric measure-ents by the MLCS. The overall standard deviation at the peakavelengths was 0.30 nm, indicating a good repeatability asso-

iated to the wavelengths obtained starting from the number ofteps.

The collected data were also employed to establish the effectiveandwidths at maximum wavelength for each step and the value3 nm was estimated in the range 440–640 nm.

The dispersion of the maximum signals measured for eachtep was also assessed and the overall relative standard deviationR.S.D.) was estimated as 2.6%. This result indicates a good repeata-ility of the signals associated to the radiant power of the whiteED.

.2. Determination of food colorants in synthetic samples

The performance of the MLCS was initially evaluated by ana-yzing food colorants in the synthetic samples. The absorbance

easurements were accomplished at the wavelengths of itsaximum absorption: 486 nm (sunset yellow), 500 nm (allura

ed) and 582 nm (brilliant blue). However, in the case of tar-razine solutions, the measurements were performed at 440 nmue to the low sensibility of the MLCS in the wavelength ofaximum absorption of this colorant (426 nm). For referenceeasurements, the food colorants were also analyzed by an HP

iode-array, model 8453, and a Micronal spectrophotometer, model34211.

In order to investigate the maximum concentration for linearesponse (the limit of linearity—LOL), ANOVA (analysis of variance)nd F-test for lack of fit were applied to the models elaboratedith the concentration range 2.0–26.0 mg L−1, for each colorant.

he results showed no lack of fit up to the LOL as presented inable 1. An F-test for the regression significance revealed that theerformed linear regressions are highly significant up to the LOLalues.

The values of limits of detection (LOD) and limits of quan-ification (LOQ), which were estimated according to IUPACecommendations [7], are also shown in Table 1. It is worth not-ng that LOL, LOD and LOQ present similar values for the threenstruments and for the four food colorants.

After the study above, the analytical curves were constructedmploying the following concentration ranges: 2.0–12.0 mg L−1 for

llura red and 3.0–18.0 mg L−1 for the other colorants. In Table 2 areresented the results for the colorants determinations in syntheticamples.

As can be seen, there is a good agreement between the results ofoncentration determined by the proposed instrument and those

1158 G. Veras et al. / Talanta 77 (2009) 1155–1159

Table 1Values of LOD, LOQ and LOL obtained for the four colorants

Colorant Instrument LOD(10−2 mg L−1)

LOQ(10−2 mg L−1)

LOL(mg L−1)

Allura redMicronal 3.1 9.4 14.0HP 0.7 2.2 14.0MLCS 0.1 0.2 12.0

Sunset yellowMicronal 3.2 9.7 18.0HP 0.7 2.1 18.0MLCS 13.0 39.4 18.0

TartrazineMicronal 2.6 7.8 18.0HP 0.8 3.6 18.0MLCS 8.1 24.7 18.0

Brilliant blueMicronal 5.5 16.6 18.0HP 7.8 23.8 18.0MLCS 3.1 9.3 18.0

Table 2The average concentration values of food colorants and confidence intervalsobtained in the analyses of synthetic samples

Colorant Sample Expectedvalue (mg L−1)

Estimated value (mg L−1)

Micronal HP MLCS

Allura Red

01 3.0 2.9 ± 0.1 2.9 ± 0.2 2.9 ± 0.102 5.0 4.7 ± 0.0 4.8 ± 0.1 4.7 ± 0.203 7.0 6.9 ± 0.1 7.0 ± 0.2 7.0 ± 0.304 9.0 8.9 ± 0.0 8.7 ± 0.1 8.9 ± 0.105 11.0 11.0 ± 0.1 11.0 ± 0.2 11.0 ± 0.4

Sunset Yellow

01 4.0 4.0 ± 0.0 4.1 ± 0.1 4.1 ± 0.202 8.0 7.8 ± 0.1 7.8 ± 0.1 7.9 ± 0.203 10.0 10.1 ± 0.1 10.0 ± 0.1 10.0 ± 0.304 14.0 13.9 ± 0.1 13.9 ± 0.1 13.8 ± 0.505 16.0 15.9 ± 0.1 15.8 ± 0.1 15.6 ± 0.2

Tartrazine

01 4.0 4.0 ± 0.1 3.9 ± 0.1 4.1 ± 0.102 8.0 8.0 ± 0.1 7.9 ± 0.1 7.7 ± 0.103 10.0 10.2 ± 0.5 10.2 ± 0.1 10.2 ± 0.204 14.0 13.9 ± 0.5 13.9 ± 0.1 14.0 ± 0.605 16.0 15.7 ± 0.0 15.8 ± 0.0 15.8 ± 0.1

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01 4.0 4.1 ± 0.1 4.3 ± 0.1 4.2 ± 0.002 8.0 8.0 ± 0.0 7.8 ± 0.1 8.0 ± 0.0

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Tabela 4Recovery percentage obtained for Fe2+ in six samples of restorative oral solutions

Sample Added Fe2+ (mg L−1) Recovery (%)

Micronal HP MLCS

013.0 99.31 100.34 97.185.0 98.86 100.19 100.567.0 98.98 97.99 96.41

023.0 99.65 100.00 97.185.0 98.67 99.24 98.057.0 96.81 97.27 97.25

033.0 96.19 94.56 93.855.0 96.01 98.48 99.867.0 98.55 99.28 99.05

043.0 101.39 98.30 98.725.0 99.62 100.19 105.297.0 99.27 98.28 98.10

053.0 104.51 103.74 97.695.0 97.91 99.05 97.507.0 99.42 101.72 100.84

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rilliant Blue 03 10.0 9.9 ± 0.0 9.9 ± 0.1 10.1 ± 0.104 14.0 13.9 ± 0.1 13.7 ± 0.1 13.8 ± 0.105 16.0 15.9 ± 0.1 15.8 ± 0.1 15.9 ± 0.1

btained by using the commercial spectrophotometers. In fact, thepplication of the t-paired test at the 95% confidence level revealedhat there are not significant differences between the concentra-ions estimated by the three instruments.

Concerning to the precision of results, in Table 2 is shown thathe confidence intervals are narrow and similar for the concen-ration values obtained by the three instruments. Moreover, theverall R.S.Ds. associated to the results from the three instruments

abela 3verage values of Fe2+ concentration and confidence intervals obtained in the anal-ses of samples of restorative oral solutions

ample Estimated values (mg L−1)

Micronal HP MLCS

01 4.9 ± 0.1 4.9 ± 0.0 4.9 ± 0.12 5.8 ± 0.1 5.7 ± 0.0 5.8 ± 0.13 4.2 ± 0.1 4.1 ± 0.1 4.3 ± 0.14 6.3 ± 0.1 6.3 ± 0.1 6.2 ± 0.15 4.5 ± 0.0 4.4 ± 0.0 4.5 ± 0.06 9.0 ± 0.1 8.9 ± 0.1 8.9 ± 0.0

.S.D. 0.8% 0.5% 0.7%

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63.0 104.51 95.58 103.855.0 97.91 97.92 94.167.0 99.42 99.43 97.68

ere smaller than 1.0%. Such finding indicates a good precision inhe estimative of the colorants concentrations by using the threenstruments.

.3. Determination of Fe2+ in restoratives oral solutions

In these analyses, the measurements of absorbance were carriedut at 515 nm in the three instruments. Restorative oral samplesf six different manufacturers were analyzed by using analyticalurves built in the concentration ranges 2.0–10.0 mg L−1, whichere validated by using ANOVA and F-test for lack of fit and for

egression significance at the 95% confidence level.Table 3 presents the results in terms of average values of Fe2+

oncentration and the confidence intervals estimated at 95% level.he results show that good agreements of the results were obtainedmploying the three instruments and it is corroborated by thepplication of the t-paired test at the 95% confidence.

Table 3 also presents the values of overall R.S.Ds., which indicategood precision in the estimation of the Fe2+ concentrations by the

hree instruments.In spite of the t-test to reveal absence of systematic error (bias)

t does not guarantee its actual absence in the results. In ordero check the eventual presence of bias, a recovery test was car-ied out according to the procedure described elsewhere [8]. Forhis purpose, known quantities of the analyte were added into theestorative samples and the measured recovery taxes are presentedn Table 4. As can be seen, satisfactory recovery taxes were obtainedor all fortified samples. Therefore, this result evidences the absencef bias associated to the estimated values of Fe2+ concentrationmploying the three instruments.

. Conclusions

A portable, inexpensive and microcontrolled spectrophotome-er was developed in this work. Such advantageous characteristicsere achieved due to the incorporation of a CD media as diffraction

rid and a white LED as radiation source. Other relevant char-cteristic of the proposed instrument concerns to the use of ahototransistor with spectral sensitivity in the visible region ashototransductor.

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The MLCS was successfully applied to the determination of foodolorants (sunset yellow, tartrazine, allura red and brilliant blue)n synthetic samples and Fe2+ in restorative oral solutions. In allpplications, the proposed instrument lead to results similar to thebtained by the commercial instruments employed for compari-on. Therefore, the proposed instrument offers an economicallyiable alternative for spectrophotometric chemical analysis in small

outine, research and/or teaching laboratories.

cknowledgement

The authors thank the Brazilian agency CNPq for scholarship.

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M.C.U. Araújo, Quim. Nova 28 (2005) 1102.4] Y. Shimazaki, S. Watanabe, M. Takahashi, M. Iwatsuki, Anal. Sci. 16 (2000)

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