Physical Chemistry Lab If Experiment No. 4 SPECTROPHOTOMETRY : Objectives 1. Examination of the essential components in a simple spectrophotometer- the Jenway 6100 & Perkin Elmer Lambda 40 . 2. Determination of absorption spectrum of a solution 3, Preparation of Beer’s Law Plot . Apparatus ~__ Spectrophotometer, Test tubes, Test tube rack Reagents Cobalt (II) Nitrate, Distilled water ‘ Procedure Operation of the Spectrophotometer In a few brief procedures, the spectral characteristics of the Spectrophotometer will be examined, and the visible spectrum of a simple substance will be recorded. It tnust be emphasized that handling of cuvettes are very important. Any variation in the cuvettes, such as stains, scratches, or changes in glass curvature will cause varying results, Thus it is essential to foilow several rules: 1. Donot handle the lower portion of the cuvette through which the light passes. -2. Always tinse the cuvette with several portions of the solution to be measured bofore taking measurements. 3. Wipe off any liquid drops or smudges with a clean tissue before placing the cuvette in the spectrophotometer. 4, When inserting a cuvette, always do so with the index mark facing the front of the instrument, and aftegthe cuvette is seated, line up the index marks exactly. A. The Visi Place a dry cuvette containing a strip of plane white paper into the instrument and rotate the cuvette until. he beam reflects upward to view (except for Jenway, cuvette is optional). Ohserve and record the color of the beam in 50 nm intervals from 350 to 650 nm. Adjust tke light control if necessary to see the beam. B. Visible Spectrum of a Solution Obtain a standard solotion of Cobalt ({1) Nitrate. Set the spectrophotometer to ‘a wavelength of 375 nm.arfftzadjust it to 100% transmittance when containing a distilled water cuvette. Be sire to use the same distilled water cuvette for all future readings. Fill a second cuvelte about one third full with the salt solution! and take the % transmittance and absorbance readings also at 375 nm. Change the School of Chemical Engineering and Chemistry ee | |Physical Chemistty Lab IT -wavelength dial to 400 nm. Again set 100% T reading for distilled water cuvette and take the % T and A readings of the same samples. C. Beer’s Law Plot Continue the procedure at the maximum absorbance of the solution. Set the ot reading to 100 sing diBilled water cuvette, Take the absorbances reading of a * 005M Cobalt (Il) Nitrate. Dilute the 0.05 M salt solution to 0.04M, 0.03M, 0.02M, DDIM, and 0.005M, and get the absorbance reading of Cobelt () ‘Nitrate solution oe nknown concentration from your instructor. Determine the concentration of the unknown from the Beer's Law plot. : ‘Treatment of Results 1. Plot the absorption spectrum, that is %6L and A versus wavelength of the Cobalt Nitrate solutions. Locate and record the ‘wavelength at which absorption maximum oceur. Also plot the absorbance reading versus the wavelength in a single graph. 2, Prepate a graph of the Beer’s Law using the data taken Prepare one graph of Yet versus concentration on a semilog paper, with 967 om the log axis, and third graph of absorbance versus concentration on the linear graph. On the absorbance versus concentration plot draw the best straight line through the points using the Ieast square method of curve fitting. Use the plot to determine the concentration of your unknown. await : | oateant af Chemical Engineering and ChemistryPhysical Chemistry Lab IT Group No. Date:_ Instructor: _ Name Signature Leader: Members: FINAL DATA SHEET A, Visible Light Wavelength Color 350 400 | ‘ 450 500 z 550 —- 600 650 ‘Absorbance | Wavelength _| Absorbance Absort 500 610 720 310 20 730, 320 60 740 330 640 750 1 $40 650 760 350 660) 770 560 670 __ 780 570 680 [790 380 0 300) 590 700 600 710 10 School of Chemical Engineering and Chemistry oer ARALC. Beer’s Law Plot Wavelengt! Concentration, M Absorbance 0.05: 0.04 : 0.03 0.02 0.01 0.005 Unknown: Concentration of Unknown. * School of Chemical Engineering and Chemistry GRAM I Physical Chemistry Lab-IE