Spectroscopy Absorbance Transmittance Beer-Lambert’s
Law Electromagnetic Spectrum Wavelength, Frequency
is a general term for the science that deals with the interactions of various types of radiation with matter. The interactions of interest were between electromagnetic radiation and matter
light, X rays, microwaves, radio waves and so forth are all different kinds of electromagnetic radiation The spectrum is divided into regions, with the visible region accounting for only a small portion, from 380 nm to 750 nm This visible region is flanked by the infrared and ultraviolet regions.
Electromagnetic radiation is said to have a dual behavior: particle and wave Therefore, it is characterized by wavelength, frequency, and amplitude The wavelength, λ (Greek lambda), is the distance from one wave maximum to the next The frequency, ν (Greek nu), is the number of waves that pass by a fixed point per unit time, usually given in reciprocal seconds (s-1) or hertz, Hz (1 Hz = 1 s-1)
The amplitude is the height of a wave, measured from the midpoint to the peak Multiplying the wavelength of a wave in meters (m) by its frequency in hertz (s-1) gives the speed of the wave in meters per second (m/s) : v λ = speed The rate of travel of all electromagnetic radiation in a vacuum is a constant value, “speed of light”, denoted by c, and has an exact value of 2.99792458 x 108 m/s, often rounded to 3.00 x 108 m/s
In this experiment, absorption spectroscopy is used. At certain wavelengths, a normally transmitting medium will absorb light of some wavelength and transmit the others. The photon’s energy is converted into an atomic or molecular process: excitation of electrons, vibration, rotation etc.
molecule’s energy is increased with exactly the same amount of energy of the photon it absorbed. M + hv M* M* is the molecule’s excited state
A photon whose energy is not equal to the difference between 2 energy states will not be absorbed
To prepare the absorbance spectrum of colored species; To determine the concentration range for maximum precision; and, To determine by spectroscopy the concentrations of colored species in a mixture.
One hundred milliliters each of 0.001 M KMnO4 and 0.001 M K2Cr2O7 were prepared.
10-mL solutions were prepared from the 0.001 M KMnO4 using the following dilution ratio of H2O:KMnO4: 10:0, 9:1, 8:2, 7:3, 6:4, 5:5, Prior3:7, usage, the 4:6, to 2:8, 1:9 spectrophotometer was warmed for 20 mins and then, the wavelength was adjusted to 545 nm.
The cuvette containing reference liquid was inserted and the absorbance was zeroed and the transmittance was set to 100%.
The absorbance of the solutions containing different H2O:KMnO4 ratio were recorded. The concentration in molarity and in ppm were computed. The concentrations were plotted against absorbance and %transmittance. The concentration range for max precision was determined. The absorbance of the unknown at 545 nm was also measured..
A 2:8 K2Cr2O7:H2O solution was prepared. The absorbance of this was measured at 440 nm and at 545 nm.
The absorption of the unknown and the 2:8 KMnO4:H2O solution at 440 nm were also measured.
The concentrations of Mn and Cr in the unknown were computed from the readings.
Dilution 1:9 2:8 3:7 4:6 5:5 6:4
MnO4 1 x 10-4 2 x 10-4 3 x 10-4 4 x 10-4 5 x 10-4 6 x 10-4
ppm Mn 5.494 10.988 16.402 21.976 27.470 32.968
A 0.208 0.489 0.749 1.019 1.285 1.541
T 0.6194 0.3243 0.1782 0.09572 0.05188 0.02877
%T 61.94 32.43 17.82 9.572 5.188 2.877
7 x 10-4
8 x 10-4 9 x 10-4
Concentration Range for Maximum Precision (ppm)
10 – 22 ppm 0.9989 0.449 9.698
Correlation Coefficient, r Absorbance of Unknown Concentration of Unknown (ppm Mn)
Concentration vs Absorption
Concentration Mn (ppm)
Figure 1. Plot of concentration of Mn (in ppm) versus the absorbance
Concentration vs Transmittance
80 60 40 20 0 0 10 20 30 40 50 60
Concentration Mn (ppm)
Figure 2. Plot of the concentration of Mn (in ppm) versus the transmittance (in %)
Sample Computations: A = εbc where slope=εb Based from the linear regression of the data, εb = 0.0463
Concentration of the unknown: 𝐴 0.449 𝑐 = = = 9.698 𝑝𝑝𝑚 𝑀𝑛 𝑏 0.0463
Spectrophotometric Determination of Mn and Cr in a Mixture
A at 545 nm A at 440 nm 0.489 0.031
Mn + Cr
Concentration of Mn (ppm) Concentration of Cr (ppm)
Sample Computations A440 = A440,Mn + A440, Cr A440 = εbcCr + εbcMn A540 = A540,Mn + A540,Cr A540 = εbcCr + εbcMn
0.031 𝑎440,𝑀𝑛 = = 0.002821 10.988 𝑝𝑝𝑚 0.089 𝑎440,𝐶𝑟 = = 0.004279 20.798 𝑝𝑝𝑚 0.489 𝑎545,𝑀𝑛 = = 0.04450 10.988 𝑝𝑝𝑚 0.016 𝑎545,𝐶𝑟 = = 0.0007693 20.798 𝑝𝑝𝑚
Plug in the known values to the equations (b=1cm, 𝑎 and absorbance A) and solve for the concentration of Mn and Cr. The experimental results were: CMn = 9.471 ppm CCr = 35.82 ppm
a and ε are both absorptivity. ε is molar absorptivity, used when c is in M and b is in cm.
Absorption Spectroscopy Transmittance (T)
Attenuation of a beam of radiation by an absorbing solution Due to interactions of photons and absorbing particles, the power of the beam is attenuated from Po to P.
Transmittance of the solution is the fraction of the incident radiation transmitted by the solution. It is given by, T = P/Po %T = (P/Po)x100 Absorbance is given by, A = -log T = log(1/T) = log(Po/P)
Absorbance is directly proportional to path length b and concentration c, and is a log function of T; *as seen in in the plot of %T vs. conc, the graph is logarithmic: as concentration increases, %T decreases *as seen in the plot of A vs. conc., the graph is linear: as concentration increases, absorbance increases
Other equation for Absorbance (A) A = abc where a is a proportionality constant called absorptivity
When b is in cm and c in molarity (mol/L), absorptivity is in L mol-1cm-1 and denoted by 𝜀 The equations A = abc and A= 𝜀bc are the statements of Beer-Lambert-Bouger Law or simply Beer’s Law
For mixtures, Beer’s Law can still be used. A = 1bc1 + 2bc2 + 3bc3 + … + nbcn where 1, 2,…,n corresponds to the different composition of the mixture.
In the experiment, an unknown mixture of KMnO4 and K2Cr2O7 was subjected to absorption spectroscopy to measure its absorbance at 545nm and 440 nm.
The absorbance measured was compared to the absorbance of the standard 0.001 M K2Cr2O7 and 0.001 M KMnO4 The absorptivity (a) of the 2 components were calculated a = A/concentration (a = varies on different wavelengths)
This was substituted to the two equations, one for 440 nm and another for 545 nm
A440 = εbcCr + εbcMn A540 = εbcCr + εbcMn System of equations was used to calculate for the concentrations of KMnO4 and K2Cr2O7 in the mixture Also, Cramer’s Rule can also be used
The violet color of the Mn is observed because it absorbs the color green ( = 500560 nm). This would explain why Mn has higher absorbance at = 545 nm. Chromium absorbs the color blue (accounting for its yellow color) which occurs at = 435-480 nm; therefore, Cr should register a higher absorbance reading at = 440 nm than at = 545 nm.
A substance exhibits a color which has a wavelength opposite to the wavelength of the color it absorbs
Spectroscopy is an efficient method that can be used in both quantitative and qualitative analyses It is more efficient when used in organic compounds because all organic compounds absorb EM radiation Many inorganic compounds do not absorb EM radiation, and they first need to be transformed into absorbing species
Beer-Lambert’s Law relates Absorbance to concentration of the analyte, path length and absorptivity. Absorbance is dependent on concentration (directly proportional) Absorptivity is constant for a certain wavelength and different in another wavelength The concentration range for max precision is estimated by getting the most linear portion of the graph
Keep the KMnO4 in a colored container to avoid its photodecomposition Make sure that the measurements to be done would be as accurate as possible (i.e. weighing etc.)
How does absorbance vary with concentration? As concentration increases, absorbance increases linearly. If plot A against C is extrapolated, will it intersect the point of origin? Yes because the absorbance was zeroed at the initial concentration (blank). How does % transmittance vary with concentration? As concentration increases, % transmittance decreases, logarithmically.
How does curve A vs C compare with that obtained by plotting %T vs C? Explain the difference.
In Figure 1, it was established that as concentration increases, absorbance also increases linearly. From the equation, A = -logT. This would explain the logarithmic decrease of the transmittance as the concentration of the substance is increased.
A 20.0ppm solution of a colored compound gives a 70.0% transmittance in a 1.00cm cell. Find the absorptivity of the solution.
c = 20.0 ppm T = 0.700 b = 1.00 cm 𝐴 = 𝑎𝑏𝑐 = − log 𝑇 1 𝑐𝑚 (20.0 𝑝𝑝𝑚) = − log(0.700) 𝑎 = 𝟎. 𝟎𝟎𝟕𝟕𝟓/𝒄𝒎 ∙ 𝒑𝒑𝒎
A solution of a colored species gives 25% transmittance. What will be the % transmittance of the solution whose concentration is 3 times that of the given? Assume linearity of response over the given concentration range.
= − log 0.25 = 𝑎𝑏𝑐 −log(0.25) 𝑐 = 𝑎𝑏 − log 0.25 𝐴 = 𝑎𝑏 × 3 = 3 × − log 0.25 𝑎𝑏 𝑇 = 10−𝐴 = 10−3log(0.25) = 0.015625 𝑻 = 𝟎. 𝟎𝟏𝟓𝟔 𝒐𝒓 %𝑻 = 𝟏. 𝟓𝟔%
Bhondwe, A. "Molar Absorptivity." Buzzle Web Portal: Intelligent Life on the Web. 3 Jan. 2011. Web. 03 Mar. 2011. <http://www.buzzle.com/articles/molarabsorptivity.html>. Hargis, L. (1988). “Analaytical Chemistry”. Prentice-Hall. New Jersey. Skoog, D. et al. “Introduction to Spectrochemical Methods”. Fundamentals of Analytical Chemistry, 8th ed. 2004. Brooks/Cole – Thomson Learning: California. pp 710, 715-716, 718, 720, 730.
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