In the Laboratory edited by
Cost-Effective Teacher
Harold H. Harris University of Missouri—St. Louis St. Louis, MO 63121
A Simple Method for Rapidly Obtaining Absorption Spectra with a Spectronic-20D+ Spectrophotometer
W
Jonathan E. Thompson Department of Chemistry, University of Nebraska at Kearney, Kearney, NE 68849;
[email protected] Use of Spectronic-20 spectrophotometers spans high school level to senior-level undergraduate curriculum. The low cost of these instruments has enabled many school districts and universities to purchase numerous Spec-20’s, and this has given nearly every student in a class the opportunity to gain hands-on experience with a spectrophotometer. In light of the popularity of Spec-20’s, various methodological improvements for their use have been suggested in the literature. Both square (1) and temperature-thermostated (2) cuvettes have been described as well as methods for interfacing and monitoring signals from the Spectronic 20 with personal computers (3, 4). While these improvements tackle certain limitations of the instrument, they do not address what is often the most common student complaint about using the instrument: the tedious process of determining an absorption spectrum for a substance of interest. Recording a spectrum over a 200-nm range with 5-nm resolution requires close to an hour of repetitive laboratory work even for experienced operators. The time required can easily double if students are not familiar with the proper operation of the instrument. Thus, this time load often represents a significant portion of a scheduled laboratory period and presents a burden to both students and educators. A closer analysis reveals the rate-limiting step of a typical procedure is often the time spent filling and washing the cuvette alternatively with blank and sample and performing the two-point calibration (0 and 100% transmittance) at each wavelength. If these steps could be modified, reduced, or eliminated, a large time savings would be realized. Towards this goal, I have developed a simple method for sequentially logging signal and wavelength data for both the sample and blank to a personal computer and modified the method for collecting an absorption spectrum. In this method two channels of the popular LabPro interface (Vernier Software and Technology, $220) were used with Logger Pro 3 software (Vernier Software and Technology, $149 for site license) to independently monitor voltages associated with the transmittance signal and measurement wavelength. Virtually all models of the Spectronic 20 feature an analog output on the bottom of the instrument to allow the user to measure a voltage that is proportional to the detector signal. The exact magnitude and range of output voltages is dependent upon the spectrophotometer model. Regardless of the signal magnitude, this voltage can be sampled by a data acquisition device and used to determine the intensity of light reaching the detector. Ultimately, this voltage can be used to determine the transmittance of the sample if the signal is measured for both the sample and the blank at a given wavelength, the measured signal scales linwww.JCE.DivCHED.org
•
early with light intensity, and both the sample and blank share a common zero point (0% transmittance). In addition to the signal data, it was also desired that the wavelength of measurement could be logged to a computer and correlated with the signal data. It was found that a small modification of Spec 20’s with digital front panel wavelength indicators could be carried out to accomplish this. Briefly, the cover of the Spec 20 was removed and wires are spliced at pins 1 and 3 of jumper J7 on the front of the Spec 20’s board assembly. These wires are routed through the body of the instrument and exit the instrument body through a small gap adjacent to the RS-232 port. These wires are then used to measure the voltage drop across a variable resistor that tracks movement of the wavelength dial of the Spec 20. The voltage drop can be correlated to the measurement wavelength through calibration, and this provides a convenient analog signal for wavelength determination. Once the modifications have been made to the Spec 20 and the LabPro is set up, an absorption spectrum can be collected in just a few minutes. In a typical experiment, the user adjusts to the starting wavelength and sets the 0% transmittance. The blank is then placed in the spectrophotometer and a data point is recorded through the data acquisition interface and software (additional details of the interface and a copy of the experiment file can be found in the Supplemental MaterialW). The user then adjusts the monochromator to the next desired wavelength and again records data. This process is repeated for the blank until signal versus wavelength data has been recorded for the entire spectral region of interest. The monochromator is then adjusted to the original wavelength setting, the blank is replaced by the sample, and a new scan initiated with measurements made at the same wavelengths as in the blank scan. The data for both the blank and sample scans are then imported into a spreadsheet program and the ratio of the signal at each wavelength for the sample as compared to the blank (e.g., sample signal versus blank signal) is calculated. This ratio represents the transmittance of the sample at each wavelength and can be converted to absorbance or plotted within the spreadsheet program. Note that the procedure does not involve setting the 100% transmittance with the blank at each wavelength, but rather involves simply measuring the detector signal (voltage) at the spectrophotometer’s analog output line for each wavelength. The percent transmittance on the front panel readout may vary anywhere from 10–200% during a scan owing to differences in detector sensitivity and emission intensity from the lamp with wavelength. Rather than accounting for these differences at each individual wavelength by setting the 100% transmittance as is normally done, this
Vol. 83 No. 6 June 2006
•
Journal of Chemical Education
913
In the Laboratory
to continually wash and fill the cuvette with sample and blank and set the 100% transmittance is eliminated. The method has been validated by comparing absorption spectra of a 0.19 mM aqueous solution of potassium permanganate obtained by the traditional approach and the method described within. The results of this study are illustrated in Figure 1. As can be observed in the figure, both methods produce similar absorption spectra. The wavelengths of maximum absorbance (λmax) coincide at 530 nm for both methods and the general shape of the spectrum and value of maximum absorbance coincide. However, the revised method provides an advantage as each spectrum required only ∼5 minutes to acquire and plot as compared to approximately 50 minutes required for the traditional approach.
A 0.6
Absorbance
0.5 0.4 0.3 0.2 0.1 0.0 400
500
600
Wavelength / nm B 0.6
Absorbance
0.5
Hazards
0.4 0.3
Meter terminals for older vacuum tube instruments can float at voltages > 100 V above ground (3). Care should be taken to reduce the possibility of electric shock, and a multimeter should be used to probe voltages prior handling bare wires. Removal of the Spectronic 20 cover should only take place after the instrument is unplugged from the wall jack.
0.2 0.1 0.0 400
500
600
Wavelength / nm Figure 1. Direct comparison between absorption spectra of potassium permanganate collected using the method described (A) and traditional scanning of the Spec-20 (B). The graph shown in (A) consists of 6 individual absorption spectra that have been overlaid on the plot to assess reproducibility. Collection of a single spectrum in (A) required approximately 5 minutes compared with approximately 50 minutes for the collection of the spectrum in (B).
W
Supplemental Material
Instructions for modifying the Spectronic-20D+ spectrophotometer and information about setting up the Vernier LabPro and software are available in this issue of JCE Online. Literature Cited
method accounts for these differences when the sample’s signal is normalized to the blank’s signal at each wavelength. While this difference may seem like a minor experimental detail, it leads to a considerable decrease in the time necessary for collection of an absorption spectrum since the need
914
Journal of Chemical Education
•
1. Aronson, J. N. J. Chem. Educ. 1975, 52, 800. 2. Thompson J. E.; Ting, J. J. Chem. Educ. 2004, 81, 1341. 3. Amend, J. R.; Morgan, M. E.; Whitla, A. J. Chem. Educ. 2000, 77, 252. 4. Nagel, E. H. J. Chem. Educ. 1990, 67, A74.
Vol. 83 No. 6 June 2006
•
www.JCE.DivCHED.org
