Double-beam Instruments

Double-beam spectrophotometers. Most modern general-purpose ultraviolet/ visible spectrophotometers are double-beam instruments which cover the range between about 200 and 800 nm by a continuous automatic scanning process producing the spectrum as a pen trace on calibrated chart paper. 

Instrumental correction for background absorption using a double beam instrument or a continuum source has already been discussed (p. 325). An alternative is to assess the background absorption on a non-resonance line two or three band-passes away from the analytical line and to correct the sample absorption accordingly. This method assumes the molecular absorption to be constant over several band passes. The elimination of spectral interference from the emission of radiation by the heated sample and matrix has been discussed on page 324 et seq. 

When using a single-beam spectrometer, I0 is measured when a reagent blank is used to zero the absorbance scale. The value of I is then measured when the sample is inserted into the spectrometer. On the other hand, when using a double-beam instrument both the reagent blank, 70, and the sample, I, are measured continuously and the appropriate ratio is determined electronically. 

In practical situations the absorbance of a sample is determined by making two measurements, the first to determine 70 and the second to determine I. The determination of I0 is used to cancel a large number of experimental factors that could affect the result. When measuring I0 the sample container must closely match the unknown container in all ways except for the analyte content. The cuvettes should be a matched pair if a double beam instrument is used and the same cuvette can be used for both the blank and sample with a single beam instrument. The blank solution filling the cuvette should be identical to the solvent that the sample is dissolved in, except for the sample itself. If done correctly, the least-squares line for the calibration graph will come very close to the 0,0 point on the graph. 

A double beam instrument splits the electromagnetic radiation into two separate beams, one for the reagent blank, and the other for the sample. There are two ways to do this. The first method uses a mirror that is half silvered and half transparent. As shown in Fig. 5.17 this results in a continuous beam of light for both the sample and reagent blank. 

Fig. 5.17. Essentials of a double beam instrument in space. This is characterized by two continuous beams of light.

The second type of double-beam instrument is one where the light source is divided into two beams by a rotating sector mirror that alternately reflects and transmits the light. This results in a chopped beam of light that alternately passes through the reagent blank and the sample as shown in Fig. 5.18. 

Figure 3.1 Schematic diagram of an AAS spectrometer. A is the light source (hollow cathode lamp), B is the beam chopper (see Fig. 3.2), C is the burner, D the monochromator, E the photomultiplier detector, and F the computer for data analysis. In the single beam instrument, the beam from the lamp is modulated by the beam chopper (to reduce noise) and passes directly through the flame (solid light path). In a double beam instrument the beam chopper is angled and the rear surface reflective, so that part of the beam is passed along the reference beam path (dashed line), and is then recombined with the sample beam by a half-silvered mirror.

Some spectrophotometers are single-beam instruments, and some are double-beam instruments. In a double-beam instrument the light beam emerging from the monochromator is split into two beams at some point between the monochromator and the detector. The double-beam design provides certain advantages that we will discuss shortly. 

Let us begin with the instrumentation. Dispersive IR instruments, similar to the double-beam instruments described for UV-VIS spectrophotometry, have been used in the past but have become all but obsolete. While some laboratories may still use these instruments, we will not discuss them here. 

What features of a single-beam spectrophotometer differentiate it from a double-beam spectrophotometer In what situations is it used over a double-beam instrument ... 

Imagine an experiment in which the molecular absorption spectrum of a particular chemical species is needed. Which instrument is preferred—a single-beam or double-beam instrument Why ... 

How does a single-beam atomic absorption instrument differ from a double-beam instrument What advantages does one offer over the other ... 

A double-beam instrument is preferred for rapid scanning because adjustments for intensity changes after each wavelength can be made immediately before a sample is read. With a singlebeam instrument, there is a delay. 

A double-beam instrument is preferred for the reasons expressed in the answer to question 20. 

Double-beam instruments involve two light paths, one to measure the sample and the other a reference or blank. 

ISO 1600 1990 Plastics - Cellulose acetate - Determination of light absorption on moulded specimens produced using different periods of heating ISO 13468-1 1996 Plastics - Determination of the total luminous transmittance of transparent materials - Part 1 Single-beam instrument ISO 13468-2 1999 Plastics - Determination of the total luminous transmittance of transparent materials - Part 2 Double-beam instrument ISO 14782 1999 Plastics - Determination of haze for transparent materials... 

Because a CCD is a two-dimensional array of pixels, it is possible to create a double-beam instrument with a single detector. Coupled in an optical design with an imaging grating, the output of two fibers may be imaged simulfaneously and separately onto the CCD and both spectra readout by the electronics. 

In practice, double-beam instruments are used where the absorption of a reference cell, containing only solvent, is subtracted from the absorption of the sample cell. Double beam instruments also cancel out absorption due to the atmosphere in the optical path as well as the solvent. 

Measured extinction spectra for aqueous suspensions of polystyrene spheres—the light scatterer s old friend—are shown in Fig. 11.19. Water is transparent only between about 0.2 and 1.3 jam, which limits measurements to this interval. These curves were obtained with a Cary 14R spectrophotometer, a commonly available double-beam instrument which automatically adjusts for changing light intensity during a wavelength scan and plots a continuous, high-resolution curve of optical density. To reproduce the fine structure faithfully, the curves were traced exactly as they were plotted by the instru-... 

Figure 10.12—Sequence of events necessary to obtain a pseudo-double beam spectrum with a Fourier transform IR spectrometer. The instrument records and stores in its memory two spectra representing the variation of lu (blank) and / (sample) as a function of wavenumber (emission spectra 1 and 2 above). Then, it calculates the conventional spectrum, which is identical to that obtained on a double beam instrument, by calculating the ratio T — /// — f(A) for each wavenumber. Atmospheric absorption (CO2 and H20) is thus eliminated. The figure illustrates the spectrum of a polystyrene film.

Some analysts still consider that dispersive double beam instruments are preferable to FTIR instruments for quantitative analysis because they are the only ones that compare simultaneously the transmitted intensities in both reference and sample beams. 

Figure 11.14—Optical path between the monochromator exit and the detector for two double beam instruments (rotating mirror model and semi-transparent mirror model). Instruments with rotating mirrors are similar to those used in IR spectrophotometers. However, the light beam from the source goes through the monochromator before it hits the sample. This minimises photolytic reactions that could occur if the sample is exposed to the total radiation from the source. The optics of instruments with two detectors are simpler and only one mirror, semi-transparent and fixed, is necessary to replace the delicate mechanisms of synchronised, rotating mirrors.

Figure 12.9—OpticaI scheme of a spectrofluorimeter having two detectors, one of which is used to control the intensity of the light source. A fraction of the incident beam is reflected by the beam splitter and monitored by a photodiode to control the intensity of the incident beam. Comparison of the signals obtained from both detectors allows the elimination of any drift in the light source. This procedure, for single beam instruments, gives approximately the same stability as with double beam instruments. (Model F4500 reproduced by permission of Shimadzu.)...

Besides the double beam instrument that eliminates background due to light fluctuations from the source by measuring the background radiation from the flame, a second radiation source can be used to determine the absorption of the matrix. 

The second set-up uses an alternating on/off field and a fixed polariser with an orientation that suppresses detection of the tt component. This system is equivalent to a double beam instrument with a common path. The atoms are affected by the Zeeman effect but particles in suspension are not (Fig. 14.14). 

Sample chambers for spectrometers come in two varieties—those holding only one cuvette at a time (single-beam) and those holding two cuvettes, one for a reference, usually solvent, and one for sample (double-beam). In a double-beam instrument, the sample spectrum is continuously corrected by subtraction of the reference spectrum. In the past, single-beam instruments were usually less expensive but more cumbersome to use because reference and sample cuvettes required constant exchange. However, modern singlebeam instruments with computer control and analysis can be programmed to correct automatically for the reference spectrum, which may be stored in a memory file. The use of both types of instruments is outlined in the applications section. 

In order to determine the chlorophyll and carotenoid content of your extract, you must measure the absorbance at several wavelengths, 661.6, 644.8, and 470 nm. If you are using a single-beam spectrophotometer, use a cuvette of acetone to zero the instrument at each wavelength. A double-beam instrument should have a cuvette of acetone in the reference beam and the acetone extract solution in the sample beam. If desired, it is instructive to obtain a complete spectrum of the extract in the range 400-700 nm. This can be compared to the spectrum obtained for each of the individual pigments in part D. 

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