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![Page 1: R O Y G B V - Faculty Server Contact | UMass Lowellfaculty.uml.edu/david_ryan/84.514/MolSpecLecture3-2005.pdf · 2005-10-28 · Beer-Lambert Law (or Beer’s Law) I o A ... Assumptions](https://reader031.vdocuments.site/reader031/viewer/2022030818/5b2ea62b7f8b9af0648cf21c/html5/thumbnails/1.jpg)
R O Y G B V
GammaRaySpectroscopy
X-RayAbsorption,Fluorescence
UV-visAbsorption,Fluorescence
InfraredAbsorptionSpectroscopy
MicrowaveAbsorptionSpectroscopy
NMREPR
NuclearTransitions
Inner ShellElectrons
Outer ShellElectrons
MolecularVibrations
MolecularRotations
SpinStates
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Quantitative Aspects of AbsorptionBeer-Lambert Law (or Beer’s Law)
IoA = log ---- = ε b CI
IT = ---- %T = T x 100
IoIo = measured source intensity
I = measured intensity after absorption
Intensity change does not change absorbance
Absorbance
Transmittance
molar absorptivityconcentration
path length
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Effects other than absorption that reduce source intensity (i.e., scattering, reflection) may also be measured as absorbance and must be accounted for when measuring I & Io
IncidentBeam
ReflectionLoses
ReflectionLoses
Cuvette
Transmitted Beam
Light loses occur due to:1) reflection at boundaries2) scattering by molecules
or particles
3) absorption which is process of interest
scatter
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• Absorbance & Transmittance are unitless• If C is mol/L & b is in cm then ε is L/mol-cm• To minimize the effect of light loses from
reflection the procedure followed in UV-visspectrophotometry is to measure Io with a reference blank of pure solvent in the light path & then measure I under the same conditions – cuvettes should be optically matched if using 2 & clean, free of scratches, lint, fingerprints, etc.
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Beer’s Law applies to all absorption processes
Assumptions made in deriving Beer’s Law:1) Only interaction between radiation (light)
and the absorber (sample) is absorption. This breaks down if reflection and scattering are not compensated for. Also breaks down if the absorbed radiation is reemitted as fluorescence (not normally a problem) or if stray light in the instrument reaches detector
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Assumptions made in deriving Beer’s Law:2) Monochromatic radiation – in reality this
condition is only approximated, instrument measures a narrow band of radiation
ε varies with λ sothe best place tomeasure A is at 1where A is nearlyconstant with λ
Measurements at2 suffer from thevariation in ε overthe bandwidth
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Assumptions made in deriving Beer’s Law:3) Pathlength is the same over the volume
being measured – this becomes a problem with round cells
λ varies across the bandwidth so somewavelengths passthrough more solutionthan others (b varies)
The consequenceis the same as forpolychromaticradiation = curved response
Roundcell
Lightbeam detector
Different components of the incident beam are absorbent with different efficiencies
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4) The nature of the absorber does not change with concentration – a variety of effects can cause this assumption to break down, e.g. dimerization, acid-base or complexation equilibria
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All of the above mentioned deviations from Beer’s Law (or instrumental deviations) are really only deviations in the sense that the experimental conditions deviate from the conditions that have been assumed in deriving Beer’s Law
Beer’s Law always holds
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Types of Spectroscopy
AbsorptionAtomic – AA - not coveredMolecular1) UV-vis – electronic2) IR – vibrational3) Microwave – rotational4) NMR (radiowave, MHz) – nuclear spin5) ESR/EPR (GHz) – electron spin
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Types of Spectroscopy
EmissionAtomic – AE & AF - not coveredMolecular1) Fluorescence2) Phosphorescence Luminescence3) Chemiluminescence (UV-vis region)
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Types of Spectroscopy ?
ScatteringRaman spectroscopy – infrared regionTurbidimetry – UV-vis regionNephelometry – UV-vis region
Index of RefractionRefractometryOptical Rotatory Dispersion
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Ultraviolet – Visible – Infrared Instrumentation
• Focus our attention on measurements in the UV-vis region of the EM spectrum
• Good instrumentation available• Very widely used techniques• Longstanding and proven methods• IR instrumentation will be considered from
time to time particularly when there are similarities to UV-vis
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Absorption measurements require:1) source of radiation2) device for dispersing radiation into component wavelengths3) a means of putting sample into the optical path, i.e., cell4) Detector to convert the EM to an electrical signal5) readout device or circuitry, i.e., meter, computer, recorder, integrator, etc.
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Block diagram of instrument for absorption
Light SourceWavelengthSelector
SampleHolder Detector
SignalProcessing
ReadoutDevice
The location of thesecan be reversed
Range of λ’s
Narrow Band of λ’s � Io
Transmitted Intensity � I
Here the wavelengthof interest isselectedfirst, thenpassedthrough the sample
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Block diagram of instrument for absorption
Light SourceWavelengthSelector
SampleHolder Detector
SignalProcessing
ReadoutDevice
Range of λ’s
TransmittedI at all λ’s
Selected λBand � I
Here all wavelengths pass through thesample together, then the wavelength ofinterest is selected and detected
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Emission measurements require:1) means of exciting emission i.e., way of populating upper energy level which spontaneously emits2) device for dispersing radiation into component wavelengths3) a means of putting sample into the optical path, i.e., cell4) Detector to convert the EM to an electrical signal5) readout device or circuitry, i.e., meter, computer, recorder, integrator, etc.
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Block diagram of instrument for emission i.e.,& fluorescence
phosphorescence
Light Source
WavelengthSelector
SampleHolder Detector
SignalProcessing
ReadoutDevice
Range of λ’s
Emitted λSpectrum � I
Selected λBand � I
WavelengthSelector
Selected λ Band for Excitation� Io
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The requirements for the various components used in different instruments change with the type of spectroscopy as well as for different kinds of measurements within a type of spectroscopy
We will consider the components separately then combine them to make the overall instrument
And finally look at the measurements with regard to theory and practice
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Sources – important characteristics1) Spectral distribution i.e., intensity vs. λ
(continuum vs. line sources)2) Intensity3) Stability – short term fluctuations
(noise), long term drift4) Cost5) Lifetime6) Geometry – match to dispersion device
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I) CONTINUUM SOURCES1) Thermal radiation (incandescence) –
heated solid emits radiation close to the theoretical “Black Body” radiation i.e., perfect emitter, perfect absorber
Behavior of Black Body- Total power ~ T4 therefore need constant
temperature for stability when using incandescent sources
- Spectral distribution follows Planck’s radiation law