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Diode-Array Spectroscopy for Educational Applications Wieland Schafer and Jiirgen Klunker Martin-Luther-UniversitatHalle-Wittenberg lnstitut fur Physikalische Chemie, Hoher Weg 7, D 06120 Halle, Germany Basics Although the fundamental design principle of the optical spectrometer was suggested by the founders of spectral analysis, KIRCHOFF and BUNSEN, as early as 1860 ( I ) ,it is still used i n numerous devices, albeit with different applications. Figure 1illustrates the construction principle of a single beam W M S spectrometer. The continuum emitted from the radiation source undergoes spectral decomposition by means of a n optical dispersion element (prism or difa b c e f Ll fraction grating). A slit masks most of the spectrum, allowing light of only slightly varying wavelengths to pass (monochromatic radiation). As the beam passes Figure 1. Construction principle of conventional single-beam spectrometers. (a) through the sample, i t is absorbed partially. The radiation source; (b) dispersion element; (c) slit; (d) monochromator; (e) samamount of ahsorution denends on the outical Droner- Pie; (f) detector; (g) recorder. ties of the substance. Thk decrease in light i i t e i s i t y is measured by the detection unit. To receive a n ahsorption spectrum, i t is necessary to vary the wavelength of the monochromatic light continually by mechanically moving t h e monochromator. T h u s , wavelength becomes a function of time. Fourier spectrometers (21, developed i n the late 60's, use a completely different method of measurement. Polychromatic light irradiates the sample and reaches a n interferometer, generating a n interference spectrum that contains all the spectral information (see Fig. 2). Because the data is unreadable, i t first must he calculated by means of a Fourier transformation. Because the development of purchasable Fourier spectrometers required computers able to perform this function, they have only been available on the market i n the last few decades. Another method of measurement was made available in 1979 with the introduction of the first commercially produced diode-array spectrometer and the subsequent appearance of a diode-array detector for HPLC i n 1982. Diode-array spectrometers differ fun- L damentally from spectrometers (3' 4)' Figure 2.Fourier spectrometer. (a)radiation source; (b)sample; (c) beam splitIn diode-array spectrometers2 a large number of ra- ter; (d)fixed mirror; (e) moving mirror: (f) detector; (g)computer; (h) Michelson diation receivers are arranged side by side to form a interferometer photodiode-array. As shown in Figure 3, the beam passes through the sample and the polychromator The diode-array spectrometer offers considerable advanwhere the wavelength becomes a function of the locus. tages over its conventional counterparts: When i t reaches the diode-array, the radiation is measured in one step. A computer reads the intensity of radiation a t High scanning speed saves time. each diode sequentially. Superior constancy of wavelengths occurs because Today, diode-array spectrometers a r e available for a there is no mechanical movement of optical components. spectral range of 180 nm to 1100 nm. The performance of a No mechanical wear or abrasion occurs. diode-array spectrometer depends on the sensitivity and Instant generation of data files for computing, storing, resolution of the silicon diode-array. The sensitivity of the and presentation are available. diode-arrays used in modern spectrometers i s comparable to that of photomultipliers. Resolution is limited only hy Diode-array spectrometers are useful especially for the the number of discrete diodes. measurement of spectra a t short intervals. As a result, one Depending on its application, a diode-array typically of the primary applications of diode-arrays is the HPLC (5, consists of between 128 and 1024 diodes. 6)that requires highly sensitive detectors that also have
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good selectivity and short detection intervals. Diode-array detectors also a r e suitable for kinetic studies and fluorescence emission spectrometry (7). Furthermore, the relatively simple design of diode-array photospectrometers allows one to build a fully 1 functioning model for educational use a t a reasonable cost. With a n ADC (analog to digital converter), a pera b c d e sonal computer can receive and plot Figure 3. Diode-array spectrometer: (a) radiation source; (b) sample; (c)polychromator; (d) diodethe spectra. array; (e)computer. Optical Assembly A diode-array for demonstration and experimentation i n educational settings allows one to take spectra from the visible portion of the spectrum. I t can he mounted on an optical bench a s shown in Figure 4. The continuum from the source (a) is parallelized by a condenser (b). The intens i t y c a n he a d j u s t e d w i t h a n i r i s diaphragm (c). The beam is then dispersed by a diffraction grating (D which, depending on its nature, produces the spectrum either by reflection or transmission. The s ~ e c t r u mis L
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projected On the diode-amay (g) at a Figure 4. Assembly of the diode-array spectrometer on an optical bench. distance that the red light (a) radiation source; (b) condenser lens; (c) diaphragm; (d) convex lens; (e) sample; (f) diffraction (775-800 n m ) a n d t h e violet light grating; (g) diode.array ( 4 0 0 4 2 5 nm) to fall on the outer diodes. The cuvette (el with the sample solution can be placed randomly i n the beam. In the installation suggested here, i t is located directly i n front of the diffraction grating. The convex lens must focus the beam into the s a m ~ l eA . low-voltage halogen bulb s e r w s 3s thc hght source. I)us t#l the low sensitivin ol'thr ohotdiodcs. oartirulilrlv i ~ ineher t tieauenrirs. tge bulb siould be 50 w or more: The lamp must he fed with smoothed direct current in order to re vent oscillation of the light intensity. If the power supply does not deliver smoothed voltage. r in ~ a r a l - . a 100 uF c a ~ a c i t o wired lel also can be used effectively. The filament of the bulb must be placed in the focus of the condenser. We used a one-lens condenser with a focal length of 120 nm. The demands made on the diffraction grating in demonstration applications are not very great. The sueeested " matine has 300 lines oer millimeter. but a greater number of lines produces hriihter spectra. prisms 5. Wirina scheme of the diode-arrav model. Fioure " cannot be substituted because. unlike diffraction matinas. R , - R16 ao s t a o e reslslors (500 KC)) do not show tKe 1in;ar the spectra produced by PDI - PD.6 511con pholodloacs ditribution of wavelengths. IN, - IN.6 ADC npLt cnannels ~~~~~
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The Diode-Array
Purchasable diode arrays are integrated circuits with microphotodiodes on their surface. Modern diode-array spectrometers usually have two diode-arrays (one for the W, one for the visible region) that often are cooled for hetter performance. The dlode-array for t~duci~tional applications is :isst,mbled with 16 silicon vhutodiodes saunre-sh:lotd, without filter). This arrangement is not only'chea~erandeasier to handle than a diode-array IC, i t is educationally advantageous, a s the viewer can see the different colors (wavelengths) of light on discrete, physical diodes while the spect r o m e t e r i s working. T h e diodes a r e mounted close together i n a single line on a printed circuit board. To supress reflections, the adjacent lateral faces of the diodes are painted black. 538
Journal of Chemical Education
I Figure 6. 3D-plot from the HPLC model experiment.
fitted with an external or internal 16-channel ADC should be able to d o t the s ~ e c t r a . The software we have written for the suggested experimental arrangement r u n s on a n IBMcompatible PCIAT with a t least 25 MHz working rate. I t requires an EGANGA card and a n ADCcard t y p e SC-1202 (Singular Technology Corp., Taipei). This economically priced ADC has 16 AD inputs (16 channels) and can convert voltages into 12-bit values. I t also features a softwarecontrolled, stabilized voltage output. The software performs the following functions: dark current offset 100% offset real time plotting of three selectable values
Figure 7. Sectional planes to Figure 7. .. .
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versus wavelength 2-Dplot for kinetic studies - 3 - D olot t o demonstrate the
transferring on-screen plots to disk (PCX format) controlled voltage output to supply the diode-array (this is optional, an external source also can be used) Taking the Measurements Needless to say, the room must be com~letelvdark to take the measurements. Before taking the sDectra, a dark current offset a n d maximum offset m u s t be conducted. The computer calculates the difference for each diode and records the infomation. Figure 8. Kinetic studies with the experimental diode-array spectrometer. The influence of strav must " lieht " be considered when conducting the dark current offset. A reliFigure 5 shows the wiring scheme. Each diode i s conable method for doing this is to block the beam just behind nected i n series to a resistor and biased with 5V. This voltthe cuvette with a sheet of black paper. The program reage can be supplied by the computer or by a n external quires a key to be pressed and then reads the values for the source. One hundred kR resistors are used. Adjustable redark current. The next step is to conduct the 100% current sistors with 500 kR are a better choice because they allow offset, that is, when 100% intensity strikes the diodes. one to adiust the sensitivitv of a single diode to receive the Thus, a reference cuvette must be filled with pure solvent corresponding wavelength. Each r(4stor is connected in paralkl to a n input of the AIIC. A;, soon as light strikes a and the beam released. Striking any key ends the procediodt:, the voltage at the resistor increases. This voltage is dure whereupon the values are recorded. While conducting recorded bv the AIM. and sent to the computer. Because the the offset, the computer displays the current values. The wavelength does not have to be highly accurate, calibratintensity of the light source must be adjusted until the difing the diode-array is relatively simple. The spectrum ference between dark current and maximum current is 100 must be projected on the array a s described above. One can or more a t each diode or the spectra will show too much fine-tune the array by using a substance with well-known "noise." After replacing the reference cuvette with the one absorption bands. At any selected measuring range, the with the sample in question, the spectra can be taken. An spectral resolution is determined by the number of diodes. adequate computer should be able to plot the spectra in In our example, using 16 diodes yielded a resolution of 25 real time and flicker-free by continually switching between nm. two pages of the video memory. The program allows one to plot intensity, absorption, and absorbance versus waveMethod of Measurement length. 1; addition, the software makes i t possible to plot the The voltaee a t the resistor that is connected to the diode -~~~ suectra one on too of another a t reeular intervals i n order appears a s a n analog signal that must be converted to a to study the kinetics of a reaction. digital signal by the ADC. Most powerful microcomputers ~~~~
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To demonstrate the principle of diode-arrav detection for HPLC, i t is necessaryto the spectra atdefined intervals in 3D. An option also is available to allow one to switch to an intensity Gersus time plot for any wavelength. Applications It is not the aim of this article to explain the theory of U V N I S spectroscopy. There i s considerable literature available providing a n overview of the theory and suggestions for experiments; e.g., Denny and Sinclair (8).For demonstration purposes. the diode-arrav can be used a s a detector for coiukn ch~omotographyh e a d of HPLC. Thus, a mixture of dvestnffs can be separated. However, during lectures and l&sons, this was too time consuming; and we opted for a simple model of chromotography. The model consists of a glass tube with a stopcock (buret). The lower p a r t contains a dense liquid (e.g., glycerin) and serves a s a "flow cell" that is placed i n the light beam instead of the cuvette. The upper part of the tube contains a number of liquids i n layers of decreasing density, alternating between colored a n d clear. When t h e stopcock i s opened, the liquids pass through the beam in a relatively short time. The resulting 3-D plot is similar to a n HPLC plot. Fieure 6 is a n examole of such a d o t . The tube cont a i n e d u t h e following liquids: carbon tetrachloride, methylene blue (2.5 mg in 100 mL of water), toluene, and neutral red (10 mgin 100 mL of methanol). The offset must be conducted usiua the colorless carbon tetrachloride in the botttom part ofthe tube before the stopcock is opened. Figure 7 shows the conrresponding absorbance versus time plots fo; the absorption maxima. The diode-array spectrometer provides a n excellent way to study the kinetics of fast reactions. The change in color of bromothymol blue, for instance, has been observed successfully by means of the experimental diode-array. Auseful reaction i s the saponification (hydrolysis) of tert-hutyl
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Journal of Chemical Education
chloride catalyzed by NaOH. The reaction produces tertbutanol and HC1 (NaCI). Acetone is used to reduce the concentration of water, making the reaction somewhat slower. Best results have been achieved with a large 100-mL cuvette. The offset can be conducted with pure water. After that, the cuvette is filled with a mixture 0>40 mL water, 40 mL acetone. 2 mL 0.05 molar NaOH. and 0.5 mL solution of 1 g bromothymol blue in 100 mL acetone. Tbe cuvette is placed in the beam. Then 20 mL tert-hutyl chloride are added, and the mixture i s stirred. When the solution is steady, the spectrometer is started. Even a slow computer should have enough time to calculate spline approximations and smooth the curves within three seconds. Figure 8 shows the resulting plot. At the beginning of the saponification of tert-butyl chloride, the indicator, bromothymol blue. a m e a r s blue (a stronr! band a t about 620 nm) because uf ;hr hnsicaoluriun sth he amount ofHCI i11crm.e~. C Th(! indicator beromes ello ow. and the pH V ~ U dccrea.ies. a weak band is formed a t 425 nm while the strong band weakens. The point a t which all spectra show the same intensity (about-470 nm) is called-the isobestic point. The interval of time before the indicator begins to change color and the duration of the change depends on the temperature. Literature Cited 1. Kirchhon, G.: Bunsen, R.
Chamische Annlyse durch Speciralhobachiungen lo sf^ wolds Klosrihrr der erakten Wis.wnrchnpen): Wilhelm Engelmann: Leipsig, 1860; Vol 72. 2. Marshall. A G.; Verdun. F R. Fourkr %)nnsfonns in NMR, Opfimi and Mess S p e r tmmelry: Elsevier: New York, 1990. 3. Milano, M. J.:Lam, S.;Sauonis,M.:Pautler,D. 8.:Pav,d.W,:Omshka.E. J ChrnmoLogr 1978, 149.599-614.
1. George. S. A : Maute. A. Chrnmvtographio 1982.15.419-425. 5. Fell. A E: Scott, H. P: Gill. R. ; Moffaf, A. C. J. C h m m o r w 1983,282, 123-140. 6. Poppe, H . Chromalogrnphio 1987.24.2542. 7. A d m n e s in Slrrndord mnd Mdhaiology in Speclromoiry, Burges. C.: Mielenr. K D.. Eds.: Elsevier: New York. 1987. 8 . Denny R. C.;Sindair, R. Virihieand Ultmuiolel Specfrnsmpy;Wiley: NeuVork, 1991.
