Chapter 13
An Introduction to
Ultraviolet/Visible Molecular
Absorption Spectrometry
Terms & Symbols
Beer’s Law
log Po/P = ebc = A
The derivation of this law assumes
a)
That the incident radiation is monochromatic.
b)
The absorption occurs in a volume of uniform
cross-section.
c)
The absorbing substances behave independently
of each other in the absorbing process.
Beer’s Law
Limitations to Beer’s Law
 Real Limitations to Beer’s Law
 Apparent Chemical Deviations
 Apparent Instrumental Deviations with
Polychromatic Radiation
 Instrumental Deviations in the Presence
of Stray Radiation
The Effects of Instrumental Noise
on Spectrophotometric Analyses
Types of Noise
 Shot noise – This noise is generated by current flowing across
a P-N junction and is a function of the bias current and the
electron charge. The impulse of charge q depicted as a single
shot event in the time domain can be Fourier transformed into
the frequency domain as a wideband noise.
 Thermal noise – In any object with electrical resistance the
thermal fluctuations of the electrons in the object will generate
noise.
 White noise- The spectral density of thermal noise is flat with
frequency.
 Burst noise – Occurs in semiconductor devices, especially
monolithic amplifiers and manifests as a noise crackle.
The Effects of Instrumental Noise
on Spectrophotometric Analyses
Types of Noise

Avalanche noise – Occurs in Zener diodes are reversed
biased P-N junctions at breakdown. This noise is considerably
larger than shot noise, so if zeners have to be used as part of
a bias circuit then they need to be RF decoupled.

Flicker noise – This noise occurs in almost all electronic
devices at low frequencies. Flicker noise is usually defined by
the corner frequency FL.
Sources of Noise

Case I: sT = k1

Case II: sT = k2(T2 + T) ½

Case III: sT = k3T
The Effects of Instrumental Noise
on Spectrophotometric Analyses
Effect of Slit Width on Absorbance
Measurements
Instrumentation
Instrument Components
 Sources
 Wavelengths selectors
 Sample containers
 Radiation detectors
 Signal processors and readout devices
Instrumentation
Sources- Light sources
Deuterium and Hydrogen Lamps
Tungsten Filament Lamps
Tungsten Filament Lamps
Instrumentation
Types of Instruments
 Single-beam
 Double-beam in space
 Double-beam in time
 Multichannel
Instrumentation
Single-Beam
Instruments
Instrumentation
Double-Beam Instruments
Instrumentation
Typical Instruments
 Photometers
 Visible Photometers
 Probe-type Photometers
 Ultraviolet Absorption Photometers
 Spectrophotometers
Instrumentation
Most common spectrophotometer:
Spectronic 20.
1. On/Off switch and zero
transmission adjustment
knob
2. Wavelength
selector/Readout
3. Sample chamber
4. Blank adjustment knob
5. Absorbance/Transmittanc
e scale
Instrumentation
 Visible
Region
Instrumentation

Single-Beam Instruments for the Ultraviolet/Visible
Region
Instrumentation
 Single-Beam
Computerized
Spectrophotometers
Inside of a
single-beam
spectrophot
ometer
connected
to a
computer.
Instrumentation

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Double-Beam
Instrumetents
Double-Dispersing
Instruments
Diode Array
Instruments
References
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http://www.anachem.umu.se/jumpstation.htm
http://userwww.service.emory.edu/~kmurray/mslist.html
http://www.anachem.umu.se/jumpstation.htm
http://userwww.service.emory.edu/~kmurray/mslist.html
http://www.anachem.umu.se/jumpstation.htm
http://www1.shimadzu.com/products/lab/spectro/uv3150.html
http://www.perten.com/product_range/diode_array/da_technology.html
http://repairfaq.ece.drexel.edu/sam/CORD/leot/course10_mod03/mod1
0-03.html
http://las.perkinelmer.com/catalog/Product.aspx?ProductID=L950
http://www.olisweb.com/products/upgrades/ir983.php
http://imagers.gsfc.nasa.gov/ems/visible.html
http://biology.easternct.edu/courses/spectwenty.htm
http://www.cairnweb.com/systems/prod_lamp.html
http://www.odyseus.nildram.co.uk/RFIC_Theory_Files/Noise_Tutorial.p
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