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Principles of
Instrumental Analysis
Chapter 14
Applications of Ultraviolet-Visible
Molecular Absorption
Spectrometry
FIGURE 6-21 Energy-level diagrams for (a) a sodium atom showing the source of a
line spectrum and (b) a simple molecule showing the source of a band spectrum.
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E = Eelectronic + Evibrational + Erotational
Excitation (of bonding electrons):
M + h → M* (excited)
Lifetime: 10-8~10-9 sec. followed with
relaxation (de-excitation) processes.
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FIGURE 6-24
Partial energy-level diagrams for a fluorescent
organic molecule.
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E = Eelectronic + Evibrational + Erotational
Excitation (of bonding electrons):
M + h → M* (excited)
Lifetime: 10-8~10-9 sec. followed with
relaxation (de-excitation) processes.
FIGURE 14-1 Ultraviolet absorption spectra for 1,2,4,5-tetrazine. In (a), the
spectrum is shown in the gas phase, where many lines due to electronic,
vibrational, and rotational transitions can be seen. In non-polar solvent (b) the
electronic transitions can be observed, but the vibrational and rotational structure
has been lost. In a polar solvent (c), the strong intermolecular forces cause the
electronic peaks to blend, giving only a single smooth absorption band.
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Bonding and Anti-bonding
Two 1s orbitals combine to form a bonding and
an anti-bonding MO.
Molecular Orbital Energy-Level Diagram for π
Bonding in Ethylene
Lone pairs in ammonia (A), water (B) and
Hydrogen chloride.
http://en.wikipedia.org/wiki/Lone_pair
http://2012books.lardbucket.org/books/principles-of-generalchemistry-v1.0/s13-04-polyatomic-systems-with-multip.html
http://www.sparknotes.com/chemistry/bonding/
molecularorbital/section1.rhtml
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Spectral nature depends on the
complexity, the physical state, and
the environment of the absorbing
species.
FIGURE 6-23
Some typical ultraviolet
absorption spectra.
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TABLE 14-1 Absorption Characteristics of Some common Chromophores.
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n—>*: 10-100 L/mol.cm
 —>*: 1000-15000 L/mol.cm
FIGURE 14-2
Absorption spectra for typical organic
compounds.
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TABLE 14-2 Absorption by Organic compounds Containing
Heteroatoms with Nonbonding Electrons
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transition between filled and
unfilled d-orbitals.
FIGURE 14-3 Absorption spectra of
aqueous solutions of transition
metal ions.
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Ions of Lanthanide & Actinide: 4f
and 5f electrons
FIGURE 14-4
Absorption spectra of aqueous
solutions of rare earth ions.
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Charge transfer absorption:
 > 10000
eD
A
Internal oxidation-reduction.
D: e-donor
A: e-acceptor
FIGURE 14-5
Absorption spectra of aqueous
charge-transfer complexes.
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The blood-red coloured complex
pentaaqua(thiocyanato-N)iron(III),
[Fe(NCS)(H2O)5]2+, indicates the presence
of Fe3+ in solution
Iron(2+),tris(1,10-phenanthroline
Fe(phen)32+
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14C Qualitative Applications
FIGUER 14-6
Effect of solvent on the absorption spectrum of acetaldehyde.
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FIGURE 14-3 Absorption spectra of aqueous solutions
of transition metal ions
cutoff wavelength
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FIGUER 14-7 Spectra for reduced cytochrome c at four spectral bandwidths.
(1) 20 nm, (2) 10 nm, (3) 5 nm, and (4) 1 nm.
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14C-3 Detection of Functional Groups (act as chromophores)
Hypsochromic (blue) shift: C=O groups in polar solvents.
Bathochromic (red) shift:
Auxochrome(助色團): shift chromophore peak to longer wavelengths or
increase its intensity.
TABLE 14-4 Absorption Characteristics of Aromatic Compounds
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Doppler red shift and blue shift
Blue shift in C=O
E1
E1
Solvent polarity ↑
h
E0
h’
E0 ’
Stabilization of ground state
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14D Quantitative Analysis by Absorption Measurements
Characteristics:
•Applications to Absorbing Species
•Applications to Non-absorbing Species
color-forming reagents (成色反應試劑): color-forming reaction should be
complete.
1) complexing agents for inorganic species:
2) organic chelating agents
Diphenylthiocarbazone:
(ligand for lead and mercury)
Diethyldithiocarbamate
(for copper)
1,10-phenenthrolene
(for iron)
Dimethylglyoxime
(for nickel)
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FIGURE 14-8 Spreadsheet to determine concentration of nitrite by multiple
standard additions.
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The total absorbance of a solution at
any given wavelength is equal to the
sum of the absorbance of the
individual components in the solution
A1 = M1bcM + N1bcN
A2 = M2bcM + N2bcN
FIGURE 14-9 Absorption spectrum of a two-component mixture (M+N) with
spectra of the individual components. Vertical dashed lines indicate optimal
wavelengths for determination of the two components.
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FIGUER 14-10 Comparison of a derivative
spectrum (a) with a standard absorption
spectrum (b).
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FIGURE 14-11 Absorption spectra of bovine
albumin: (a) ordinary spectrum, (b) firstderivative spectrum, (c) second-derivative
spectrum.
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14E Photometric and Spectrophotometric Titrations
FIGURE 14-12 Typical photometric titration curves.
Molar absorptivities of the analyte, the product,
and the titrant are given by εA,εP,εT, respectively.
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Figure 13-18.
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In its protonated form, Eriochrome Black T
is blue. It turns red when it forms a complex
with calcium, magnesium, or other metal
ions.
FIGURE 14-13(a) Photometric titration curves; (a) total hardness of water, (b) determination
of sulfate. In (a), total water hardness is obtained by titration with 0.10 M EDTA at 610
nm for 100 mL of a solution that contained 2.82 mmol/L total hardness. Eriochrome
Black T was the indicator.
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Turbidimetric titrations
FIGURE 14-13(b) In (b), 10.0 mL of a solution containing sulfate was titrated with 0.050 M
BaCl2 using Thorin as an indicator and a wavelength of 523 nm. The response shown is
proportional to transmittance.
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14F Spectrophotometric Kinetic Methods
Measurements are made under dynamic conditions in which the conc. of
reactants and products are changing as a function of times.
A+R
P
FIGURE 14-14 Change in concentration of analyte [A] and product [P] as a function of
time. Until time te the analyte and product concentrations are continuously changing.
This is the kinetic regime. In the equilibrium region, after te, the analyte and product
concentrations are static.
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FIGURE 14-15 Plot of initial rate of product formation as a function of substrate
concentration, showing the parts of the curve useful for the determination of
substrate and enzyme.
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FIGURE 14-16 Stopped-flow mixing
apparatus. To begin the experiment,
the drive syringes are filled with reagent
and sample and valves A, B, and C are
closed. The drive mechanism is then
activated to move the drive syringe
plungers forward rapidly. The reagent
and sample are mixed in the miser and
pass immediately into the observation
cell and stop syringe. When the stop
syringe fills, the plunger strikes the stop
block and the flow ceases almost
instantly with a recently mixed plug of
solution in the spectrophotometric
observation cell. For well-designed
systems, the time between mixing and
observation can be on the order of 2 to
4 ms.
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FIGURE 14-17 The predicative approach in kinetic methods. A mathematical model,
shown as the solid line, is used to fit the response, shown as the squares, during the
kinetic regime of a reaction. The model is then used to predict the equilibrium
absorbance, Ae, which is related to the analyte concentration. In the example shown,
the absorbance is plotted versus time and the early time data used to predict Ae, the
equilibrium value, shown as the circle.
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