Chapter 29

Write a protocol for using a spectrophotometer. After reading this chapter, prepare a detailed stepwise protocol explaining how to use one of the spectrophotometers in your department. Ask another student or a tutor to evaluate your protocol and provide you with feedback.

Use the Beer-Lambert relationship in quantitative spectrophotometric analysis. Calculate the following (express your answer to 3 significant figures): (a) The concentration \(\left(\mu \mathrm{g} \mathrm{mL}^{-1}\right)\) of pentachlorophenol in a test solution giving an absorbance at \(300 \mathrm{~nm}\left(A_{300}\right)\) of \(0.57\) in a cuvette of path length \(5 \mathrm{~mm}\), based on an absorptivity of \(20 \mathrm{~L} \mathrm{~g}^{-1} \mathrm{~cm}^{-1}\). (b) The amount (ng) of pentachlorophenol in a \(50 \mu \mathrm{L}\) sub-sample from a test solution where \(A_{300}=0.31\) in a cuvette of path length \(1 \mathrm{~cm}\), based on an absorptivity of \(20 \mathrm{~L} \mathrm{~g}^{-1} \mathrm{~cm}^{-1}\).

Determine the molar absorptivity of a substance in aqueous solution. A solution of \(p\)-nitrophenol containing \(8.8 \mu \mathrm{g} \mathrm{mL}^{-1}\) gave an absorbance of \(0.535\) at \(404 \mathrm{~nm}\) in a cuvette of path length \(1 \mathrm{~cm}\). What is the molar absorptivity of a \(p\)-nitrophenol at \(404 \mathrm{~nm}\), expressed to 3 significant figures? (Note: \(M_{r}\) of \(p\)-nitrophenol is \(291.27\).)