Chapter 13

An Introduction to
Ultraviolet/Visible Molecular
Absorption Spectrometry

Terms & Symbols

Beers 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.

Beers Law
Limitations to Beers Law
Real Limitations to Beers 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)

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

Double-Beam
Instrumetents
Double-Dispersing
Instruments
Diode Array
Instruments

References

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/mod10-03.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.pdf