Spectrophotometry Fundamentals Questions and Answers

Answer: A

A change in molar absorptivity alters the slope rather than the intercept, since it multiplies the concentration term. The other three all add a constant absorbance present even at zero analyte. Whether a defect changes a line's slope or its intercept is the first distinction to draw when a calibration looks wrong, and it halves the list of possible causes immediately.

Answer: B

The reference path carries a matched cell containing the blank, so solvent and cell absorption cancel along with any source or detector drift. An empty path would leave the cell and solvent uncorrected, a second sample would cancel the analyte too, and a fixed filter corrects nothing that varies. Arranging the reference to differ from the sample in exactly one respect is what makes the comparison meaningful.

Answer: C

A rotating mirrored disc directs the beam first through the sample and then through the reference, many times a second, so one detector sees both and their ratio is formed electronically. Ambient rejection is a useful side effect of the modulation rather than the purpose, and neither slit nor detector is switched. Using one detector for both paths removes any mismatch between two detectors, which would otherwise reappear as exactly the error the arrangement exists to remove.

Answer: D

Any difference in sensitivity or drift between two detectors would appear in the ratio as an error indistinguishable from absorbance, which is precisely what the double-beam arrangement is trying to eliminate. Speed and amplification are secondary, and cell matching is needed regardless. Removing a component from a comparison, rather than trying to match two of them, is nearly always the more robust choice, and the chopper is what makes it possible.

Answer: A

A split-beam design divides the light permanently and gives each path its own detector, so both are measured at once with no moving parts. The price is that the two detectors must be matched and must stay matched. Chopping trades mechanical complexity for freedom from detector mismatch, and splitting trades the reverse - which is a clean illustration of two designs solving the same problem by moving the difficulty to different places.

Answer: B

Without a matched cell in the reference path the sample cell's walls and the solvent would appear as absorbance, since only one path would contain them. Beam intensity, detector balance and chopper operation are not the reason. The double-beam arrangement cancels whatever is common to the two paths, so anything that must cancel has to be placed in both.

Answer: C

Subtracting the blank's own contribution:

\[ A = 0.65 - 0.05 = 0.60 \]

giving 0.60. In normal use the blank sits in the reference path and the subtraction happens automatically, which is exactly why the instrument reads 0.60 directly when set up properly. Understanding what the automatic correction is doing matters when a reading has to be checked or a blank is suspect.

Answer: D

Absorbance is the logarithm of a ratio, and the reference supplies the denominator, so a different reference gives a different number for the same sample. Detector gain, wavelength calibration and lamp output are unaffected. That a measurement is meaningless without stating what it was measured against is true of any ratio, and it is why a published absorbance is always accompanied by its solvent and conditions.

Answer: A

A bubble in the beam scatters light out of the path, and the instrument records the missing light as absorbance. It does shorten the liquid path, but the scattering dominates and pushes the reading up rather than down. A bubble outside the beam is indeed harmless, which is why cells are filled and tapped carefully. An instrument that measures attenuation cannot distinguish absorption from scattering, and every practical precaution in this topic follows from that limitation.

Answer: B

Small differences in how a cell sits in its holder change which part of the glass the beam passes through and how it is reflected, giving a spread of readings that a single measurement hides. Settling, detector recovery and lamp output are not the reason. Repeating a measurement after disturbing the thing measured is what reveals the reproducibility of the whole procedure rather than merely the stability of the electronics.