Spectrophotometer

MATERIALS

Bromophenol blue, mixture of bromophenol blue and methyl orange solutios, micropipette, cuvette, test tubes, tips.

PROCEDURE

Part 1: Determination of Amax of bromophenol blue

The wavelength of spectrophotometer is set to 470nm

Distilled water is filled into a 1cm cuvette and the smooth surface of the cuvette is cleaned using a paper towel. The cuvette is placed into the spectrophotometer and was re-zeroed. This cuvette is used as a blank.

The cuvette containing distilled water is removed and replaced with another cuvette which contains bromophenol blue.

The absorbance is read and recorded in Table 1.1 and the experiment is repeated with the wavelength 500, 530, 560, 590, 620, 650 and 680nm. Each experiment is re-zeroed using the blank cuvette after setting each wavelength.

A graph of absorption spectrum is plotted (absorbance readings versus corresponding wavelength) and the wavelength with maximum absorbance reading from the graph is determined.

Part 2: The effect of concentration on absorbance of bromophenol blue solution

The mixture of tubes 1-6 is prepared and labelled according to Table 1.2a by using a micropipette and a vortex mixer to mix the contents.

The spectrophotometer is set to the Amax wavelength for bromophenol blue obtained in part 1. Tube 1 is used to zero the absorbance reading.

The absorbance reading of tubes 2-6 are measured by using the same cuvette used for tube 1.The absorbance readings are recorded in Table 1.2a The samples are returned to the original test tubes

The concentration of the bromophenol blue solutions in the test tubes 1-6 are calculated and the concentrations are recorded in Table1.2a

A standard concentration curve of absorbance versus concentration of bromophenol blue is plotted. The molar absorptivity of bromophenol blue at Amax of bromophenol blue (in unit L mg-1 cm-1 as described in Beer-Lambert’s Law) by using the standard concentration curve.

Part 3: Determination of the...