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 LawReal 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 AnalysesTypes 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 ComponentsSourcesWavelengths selectorsSample containersRadiation detectorsSignal processors and readout devices

Instrumentation

Sources- Light sources

Deuterium and Hydrogen Lamps

Tungsten Filament Lamps

Tungsten Filament Lamps

Instrumentation

Types of InstrumentsSingle-beamDouble-beam in spaceDouble-beam in timeMultichannel

Instrumentation

Single-Beam Instruments

Instrumentation

Double-Beam Instruments

Instrumentation

Typical InstrumentsPhotometersVisible PhotometersProbe-type PhotometersUltraviolet Absorption PhotometersSpectrophotometers

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/Transmittance scale

InstrumentationVisible Region

Instrumentation Single-Beam Instruments for the Ultraviolet/Visible

Region

InstrumentationSingle-Beam Computerized

Spectrophotometers

Inside of a single-beam spectrophotometer 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