Download Ultraviolet and visible molecular absorption spectroscopy

Chapter 26
Molecular Absorption
Spectrometry
26 A Ultraviolet and visible molecular absorption spectroscopy
Ultraviolet and visible radiation absorption by molecular species can be
used for qualitative and quantitative analyses.
UV-visible absorption is used to monitor titrations and to study the
composition of complex ions.
Absorption generally occurs in one or more electronic absorption bands,
each of which is made up of many closely packed but discrete lines.
Each line arises from the transition of an electron from the ground state to
one of the many vibrational and rotational energy states associated with
each excited electronic energy state.
Absorption of radiation by organic molecules in the wavelength region between
180 and 780 nm results from interactions between photons and electrons that
either
participate directly in bond formation or that are localized about atoms.
The wavelength of absorption of an organic molecule depends on how tightly
its electrons are bound.
The shared electrons in carbon-carbon or carbon-hydrogen single bonds are
so firmly held that their excitation requires energies corresponding to
wavelengths in the vacuum ultraviolet region below 180 nm.
Electrons in double and triple bonds of organic molecules are not as strongly
held and are therefore more easily excited by electromagnetic radiation.
Unsaturated organic functional groups that absorb in the ultraviolet or visible
regions are known as chromophores.
Saturated organic compounds containing such heteroatoms as oxygen,
nitrogen,
sulfur, or halogens have nonbonding electrons that can be excited by
radiation in the 170- to 250-nm range.
Absorption occurs when electrons make
transitions between filled and unfilled dorbitals with energies that depend on the
ligands bonded to the metal ions.
The energy differences between these dorbitals
(and thus the position of the corresponding
absorption maxima) depend on the position
of
the element in the periodic table, its
oxidation
state, and the nature of the ligand bonded to
it.
Charge-transfer absorption is important for
quantitative analysis because
molar absorptivities are unusually large
(e > 10,000 L mol 1 cm 1), which leads to
high
sensitivity.
Many inorganic and organic complexes
exhibit this type of absorption and are
therefore called charge-transfer
complexes.
A charge-transfer complex consists of an
electron-donor group bonded to an
electron acceptor.
In most charge-transfer complexes
containing a metal ion, the metal serves as
the
electron acceptor.
Qualitative Applications of Ultraviolet/ Visible Spectroscopy
Spectrophotometric measurements with UV radiation are useful for detecting
chromophoric groups.
However, ultraviolet spectra do not have sufficient fine structure to permit an
analyte to be identified unambiguously.
UV qualitative data must be supplemented with other physical or chemical
evidence such as infrared, nuclear magnetic resonance, and mass spectra as
well as solubility and melting- and boiling-point information.
UV spectra for qualitative analysis are usually measured using dilute
solutions
of the analyte.
For volatile compounds, however, gas-phase spectra are often more useful
than liquid-phase or solution spectra.
A solvent for ultraviolet/visible spectroscopy must be transparent in the
region
of the spectrum where the solute absorbs.
Also, the analyte must be sufficiently soluble in the solvent.
Solvent polarity often influences the position of absorption maxima.
For qualitative analysis, analyte spectra should thus be compared to spectra
of known compounds taken in the same solvent.
The Effect of Slit Width
Small slit widths are used for qualitative studies to preserve maximum spectral
detail.
Peak heights and peak separation are distorted at wider bandwidths.
Stray radiation occasionally causes false peaks to appear when a spectrophotometer
is being operated at its wavelength extremes.
The important characteristics of spectrophotometric and photometric
methods are:
1.
2.
3.
4.
5.
Wide applicability
High sensitivity
Moderate to high selectivity
Good accuracy
Ease and convenience
Molecular absorption measurements are applicable in every area requiring
quantitative information:
1. Applications to absorbing species
1. Applications to Nonabsorbing species - Many nonabsorbing analytes can
be determined photometrically by causing them to react with
chromophoric reagents to produce products that absorb strongly in the
ultraviolet and visible regions.
Procedural Details
Wavelength Selection: To realize maximum sensitivity, spectrophotometric
absorbance measurements are usually made at the wavelength of maximum
absorption.
Variables that influence absorption: solvent, the pH of the solution,
temperature, high electrolyte concentrations, and the presence of interfering
substances.
The relationship between absorbance and concentration: The calibration
standards should approximate as closely as possible the overall composition
of the
actual samples and should encompass a reasonable range of analyte
concentrations.
The Standard Addition Method: The composition of calibration standards
should
approximate the composition of the samples to be analyzed.
The multiple-additions method is often chosen for photometric or
spectrophotometric analyses.
Several increments of a standard solution are added to sample aliquots of the
same size.
Each solution is then diluted to a fixed volume before measuring its
absorbance.
When the amount of sample is limited, standard additions can be carried out by
successive addition of increments of the standard to a single measured aliquot
of the unknown.
The measurements are made on the original solution and after each addition of
standard analyte.
Assume that several identical aliquots Vx of the unknown solution with a concentration cx are transferred to volumetric flasks having a volume Vt.
To each of these flasks is added a variable volume Vs mL of a standard solution of
the analyte having a known concentration cs.
If the chemical system follows Beer’s law, the absorbance of the solutions is
described by:
As 
bVs cs
Vt

bVx c x
Vt
 kVs c s  kVx c x
A plot of As as a function of Vs should yield a straight line of the form
As  mVs  b
m  kcs
b  kVx c x
Where m is the slope and b is the intercept
The unknown concentration cx can then be calculated as follows:
kcs
m

b kVx c x
Then,
cx 
bcs
mVx
Assuming that the uncertainties in cs, Vs, and Vt are negligible, the relative
variance is
2
2
2
 sc

 cx

s  s 
   m    b 
m b

2
sc  c x
 s m   sb 
   
m b
2
The total absorbance of a solution at any given wavelength is equal to
the sum of the absorbances of the individual components in the solution.
Wavelengths should be selected so that the
molar absorptivities of the two components
differ significantly.
Thus, at 1, the molar absorptivity
of component M is much larger than that for
component N.
From the known molar absorptivities and path length,
A1 = M1bcM + N1bcN
A2 = M2bcM + N2bcN
The accuracy and precision of spectrophotometric analyses are often limited
by
the indeterminate error, or noise, associated with the instrument.
The relationship between the noise encountered in the measurement of T and
the resulting concentration uncertainty can be derived by writing Beer’s law in
the form
1
 0.434
c   log T 
ln T
b
b
c 
Taking the partial derivative, we get
c can be interpreted as the uncertainty in c
Dividing the two equations, results in
 0.434
T
bT
c 0.434  T 



c
log T  T 
c
c

0.434   T 


log T  T 
Concentration errors When T = k1.
For many photometers and spectrophotometers, the standard deviation
in the measurement of T is constant and independent of the magnitude of
T.
Concentration errors when T = k2T2 + T
This error originates in the shot noise that causes the output of
photomultipliers and phototubes to fluctuate randomly abut a mean value.
Concentration errors when T = k3 T
The relative standard deviation in concentration from this type of uncertainty
is inversely proportional to the logarithm of the transmittance.
This type of uncertainty is important at low absorbances (high
transmittances) but approaches zero at high absorbances.
Photometric and Spectrophotometric Titrations
Titration Curves
A photometric titration curve is a plot of absorbance (corrected for volume
change)
as a function of titrant volume.
Instrumentation
Photometric titrations are usually performed with a spectrophotometer or a
photometer that has been modified so that the titration vessel is held
stationary in the light path.
Application
Photometric titrations often provide more accurate results than
a direct photometric determination because the data from
several measurements are used to determine the end point.
The presence of other absorbing species may not interfere
since only a change in absorbance is being measured.
The photometric end point has also been used to great
advantage in titrations with EDTA and other complexing
agents.
Spectrophotometry is a valuable tool for determining the composition of
complex
ions in solution and for determining their formation constants.
The three most common techniques used for complex-ion studies are
(1) the method of continuous variations,
(2) the mole-ratio method, and
(3) the slope-ratio method.
In the method of continuous variations, cation and ligand solutions with
identical
analytical concentrations are mixed such that the total volume and the total
moles of reactants in each mixture are constant but the mole ratio of reactants
varies
systematically.
The absorbance of each solution is then measured.
The corrected absorbance is plotted against the volume fraction of one
reactant, that is, VM /(VM + VL), where VM is the volume of the cation solution
and VL is the volume of the ligand solution.
In the mole-ratio method, a series of solutions is prepared in which the analytical
concentration of one reactant (usually the metal ion) is held constant while that
of
the other is varied.
A plot of absorbance versus mole ratio of the reactants is then prepared.
If the formation constant is reasonably favorable, two straight lines of different
slopes that intersect at a mole ratio that corresponds to the combining ratio in
the complex are obtained.
The slope-ratio approach is particularly useful for weak complexes but is
applicable only to systems in which a single complex is formed.
The method assumes
(1) that the complex-formation reaction can be forced to completion by a
large excess of either reactant,
(2) that Beer’s law is followed under these circumstances, and
(3) that only the complex absorbs at the wavelength chosen.
26 B Automated photometric and spectrophotometric methods
Instrumentation
Sample and Reagent Transport System
Sample Injectors and Detectors
Sample sizes for flow-injection analysis range from 5 to 200 mL, with 10 to
30 mL
being typical for most applications.
For a successful determination, it is important to inject the sample solution
rapidly as a plug, or pulse, of liquid; in addition, the injections must not
disturb the flow of the carrier stream.
The most useful and convenient injector systems are based on sampling
loops similar to those used in chromatography.
Advanced Flow-Injection Techniques
Flow-injection methods have been used to accomplish separations,
titrations, and
kinetic methods.
These methods include flow reversal FIA, sequential injection FIA, and labon-avalve technology.
26 C Infrared absorption spectroscopy
Infrared spectroscopy is a powerful tool for identifying pure organic and
inorganic
compounds because almost all molecular species absorb infrared radiation.
However, it is a less satisfactory tool for quantitative analyses than its
ultraviolet and visible counterparts because of lower sensitivity and frequent
deviations
from Beer’s law.
Additionally, infrared absorbance measurements are considerably less
precise.
The energy of infrared radiation can excite vibrational and rotational
transitions, but it is insufficient to excite electronic transitions.
Instruments for Infrared Spectrometry
Three types of infrared instruments:
1. Dispersive spectrometers (spectrophotometers),
2. Fourier transform spectrometers, and
3. Filter photometers.
The first two are used for obtaining complete spectra for qualitative
identification, while filter photometers are designed for quantitative work.
Fourier transform and filter instruments are nondispersive, that is, neither
uses a grating or prism to disperse radiation into its component
wavelengths.
Dispersive infrared instruments are similar in general design to the doublebeam (in time) spectrophotometers excepting the location of the cell
compartment with respect to the monochromator.
Fourier transform infrared (FTIR) spectrometers offer the advantages of high
sensitivity, resolution, and speed of data acquisition.
Fourier transform instruments contain no dispersing element, and all
wavelengths
are detected and measured simultaneously using a Michelson
interferometer.
In order to separate wavelengths, it is necessary to modulate the source
signal and pass it through the sample in such a way that it can be recorded
as an interferogram.
The interferogram is subsequently decoded by Fourier transformation, a
Qualitative Applications of Infrared Spectrometry
An infrared absorption spectrum contains an array of sharp peaks and
minima that are useful for the identification of functional groups are
located in the shorter-wavelength region of the infrared.
Quantitative Infrared Spectrometry
Quantitative infrared absorption methods differ from their ultraviolet and visible
counterparts because of the greater complexity of the spectra and the narrowness
of the absorption bands.
The intensity of the radiation passing through the sample is compared with that of
the unobstructed beam.
Alternatively, a salt plate may be used as a reference.
The resulting transmittance is often <100%, even in regions of the spectrum where th
sample is totally transparent.
Applications of Quantitative Infrared Spectroscopy