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Chapter 10: Spectroscopic Methods

Absorption, emission and scattering methods including UV/Vis, IR, atomic absorption, photoluminescence, atomic emission, turbidimetry and nephelometry

Neil Fitzgerald

on 27 August 2014

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Transcript of Chapter 10: Spectroscopic Methods

Chapter 10: Spectroscopic Methods
Overview of Spectroscopy
UV/Vis and IR Spectrometry
Atomic Absorption
Atomic Emission
Nephelometry and Turbidimetry
Wave Particle Duality
Electromagnetic Radiation
Example Spectroscopic Techniques
X-ray Florescence
UV/Vis Spectroscopy
Atomic Absorption
Atomic Emission
IR Spectroscopy
Microwave Spectroscopy
Nuclear Magnetic Resonance
Radiation can be treated as waves or particles (called photons)
Basic Components of Spectroscopic Instruments
: an atom or molecule absorbs radiation. Electrons move to a higher energy level (excited). Requires a radiation source
: Electrons in an excited atom or molecule relax releasing radiation. Atom or molecule can be excited by a radiation source (fluorescence or phosphorescence), thermally by a flame or plasma (e.g. ICP), or by a chemical reaction (chemiluminesence)
Wavelength selection
sources provide radiation over a wide wavelength range. Often produced by the electrical discharge of a high pressure gas e.g. deuterium lamp (for UV) or Xenon lamp (for fluorescence). Required for scanning spectrometers.
sources emit radiation at specific wavelengths. Often produced by the electric discharge of a low pressure atomic gas e.g hollow cathode lamp for AA.

Good for measurements at a single wavelength. Lasers produce very intense coherent radiation and are considered line sources
Sources of thermal energy for emission include flames and plasmas e.g. Flames Emission Spectroscopy of Inductively Coupled Plasma Atomic Emission Spectrometry
IR sources are often made from resistive material that emit IR radiation (heat) when an electric current is applied e.g. nichrome wire or Nernst Glower
Exothermic chemical reactions can also be used as a source of energy in spectroscopy e.g. chemiluminescence
It is usually necessary to select a narrow band of wavelengths (bandwidth) in order to perform a spectroscopic experiment. A larger bandwidth allows more radiation to pass (improved signal:noise) but poorer resolution
Fourier Transform
Absorption filters absorb radiation at high and low wavelengths to allow a narrow band to pass
Interference filters use constructive and destructive interference to select a narrow band of wavelengths
Simple (no moving parts) and cheap but limited to a single wavelength. Good for instruments dedicated to a specific analysis
Part of a spectrometer incorporating a dispersive element (usually a diffraction grating) that can be rotated, either manually or with a motor, to select different wavelengths Good for scanning spectrometers. Bandwidth depends on the width of the exit slit
Gratings are polished surface with regular grooves (like a compact disk) that disperse light due to constructive and destructive interference
Uses an interferometer to allow wavelengths to be measured in the time domain (the interferogram) and converted mathematically (using a Fourier Transform) into the frequency domain to obtain a spectrum.
Two major advantages:
Jacquinot's advantage: more radiation can pass (better signal: noise)
Fellgett's advantages: spectra collected quickly allowing replicate spectra to be averaged (also improves signal: noise)
Photon Transducers
Detector incorporating a photoemissive material
Photoemissive materials eject electrons when photons strike them e.g. a photodiode
Ejected electrons can be multiplied by dynodes to produce a larger current in a photmultiplier tube
When many photodides are arranged into an array (called a photodiode array), it is possible to collect a spectrum over a large wavelength range instantaneously.
Photmultipler tube
Dynodes at positive potential eject multiple electrons for each electron that strikes them
Advantages: Simple cheap and sensitive
Disadvantages: produces a "dark current" (signal with no light present). Can only measure one wavelength at a time
Thermal Transducers
For IR spectroscopy, thermal transducers are often used
Thermal transducers respond to a change in temperature for example thermistors change electrical resistance with temperature, pneumatic transducers responded to a change in gas volume as a function of temperature
Deuterium lamp: continuum Source
Hollow Cathode Lamp: line source
Beer's law
UV/Vis absorption involves promoting electrons to a higher electronic energy level due to the absorption of photons
IR photons have less energy and can only promote electrons to higher vibrational levels within a electronic energy levels
Limitations to Beer's Law
At high concentrations, analyte species can interact with each other such that the molar absorptivity changes and absorption is no longer directly proportional to concentration
Occurs when the analyte is involved in equilibria. For example, if the analyte is a weak acid, it is at equilibrium with its conjugate base which will have a different molar absorptivity causing an apparent deviation from Beer's Law
Deviation due to the assumption of monochromatic radiation.
Monochromatic radiation is not possible, only a narrow range of wavelengths.
Molar absorptivity depends on wavelength. Good agreement to Beer's law is possible for a narrow bandwidth at the peak maximum
Instrumental deviations can also occur due to the presence of stray light (Pstray) in the spectrometer
Instrument Designs
Sample cells
IR Spectroscopy
filter photometer
uses absorption or interference filters to select the wavelength. It uses a shutter to calibrate (shutter closed = 0%T
shutter open with blank solution = 100 %T).
Simple and cheap but must be set a specific wavelength. Cannot scan to obtain a spectrum
single beam instrument
uses a monochromator to select the wavelength. Different wavelengths can be selected by rotating the grating. Cannot scan to obtain a spectrum
double beam instrument
splits light to two directions, one passes through the sample and the other through a blank. Uses a motor to rotate the monochromator (scanning). Can obtain a spectrum
diode array spectrometer
uses a solid state detector (like a digital camera) to measure lots of wavelengths at once. Small and fast. Can obtain replicates quickly and signal average to improve signal to noise ratio. The signal to noise ratio improves as a function of the square root of the number of replicates
Sample cells have to be transparent to the source radiation. In the visible region any transparent material will work. In the UV region sample cells are made from quartz. For IR salt plates (e.g. KBr) are used. Cells should be clean (avoiding scratches and fingerprints) to reduces light scattering. According the Beer's law, larger pathlengths lead to greater absorbances, however, short pathlengths are usually sufficient (1 cm cells are common for UV/Vis)
IR spectroscopy can use filter photometer, double beam or fourier transform (FTIR) instruments. FTIR is now the most common design. For FTIR, a blank must be obtained and subtracted from sample spectra. Liquid samples can be sandwiched between salt plates. Solids are made in a 'mull' by mixing with a viscous oil (e.g. Nujol) or made into a pellet by mixing with salt and applying pressure. Another approach is to guide the beam through a crystal (known Attenuated Total Reflectance). Some of the IR radiation will 'leak' out of the crystal and can be absorbed by a sample in close contact with it.
Hollow Cathode Lamps
Nebulizer and
spray chamber
Atomic absorption requires the analyte to be present as free atoms. The most common methods to atomize a sample are using a flame or a graphite furnace
Sample introduction for flame AA is usually accomplished using a spray chamber and nebulizer. Liquid samples are made into a spray in the spray chamber. The spray chamber acts to break large droplets into smaller ones which pass through to the flame
In Graphite furnace AA, the sample is placed directly onto the surface of the graphite cuvet, usually by an autosampler
Sample Introduction
Graphite Furnace
In graphite furnace AA, a small volume of sample in place in a small graphite cuvet. The cuvet is heated by applying a high voltage (in an argon atmosphere to prevent combustion of the graphite). A heating program is used to dry, ash, and atomize the sample
Advantages of graphite furnace compared to flame AA are: lower detection limits (ug/L level compared to mg/L level for flame), small volume of sample used, and minimal sample preparation required
Disadvantages are: slow analysis time (1-2 minutes per measurement), difficult to measure volatile elements (requires the use of a chemical modifier to prevent loss of analyte during the ashing stage)
An analyte is said to be exited when an electron is promoted to a higher energy state. The excited analyte will quickly return to its lower energy state (relaxation)
When excitation is caused by absorbing a photon, it leads to fluorescence or phosphorescence
Inter system crossing is when an electron is converted from a singlet to a triplet state
Excited triplet state is when the electrons are not paired
In an excited singlet state, the electrons are spin paired
Internal conversion. Electron transitions from a higher to lower level without emitted radiation
Electrons in an analyte can also be excited thermally. Higher temperature causes more analyte atoms or molecules to become excited and a greater emission intensity. The fraction of analyte species in the excited state is given by the Boltzmann equation
The ability of an exited analyte to emit a photon is defined by the florescent quantum yield, , which is the fraction of excited state molecules returning to the ground state. Florescence intensity is related to the quantum yield according to the equation:
RF coil
Plasma torch
A fluorescence instrument using filters is called a
. A fluorescence instrument using monochromators is called
has two monochromators and is capable is scanning the excitation wavelengths and/ or the emission wavelengths
Scanning the excitation wavelengths and monitoring a fixed emission wavelength produces an excitation spectrum. Scanning the emission wavelengths at a fixed excitation wavelength produces an emission spectrum
Sources are commonly either mercury discharge lamps for fluorimeters (line source) or xenon arc lamps (continuum source) for spectrofluorimeters
Florescence measurements are obtained at 90 degrees from the incidence light to avoid incident radiation (fluorescence emission occurs in all directions)
Good fluorophores tend to have large delocalised electron systems e.g. nonheterocylic aromatic compounds. Quantum yields are also influenced by temperature and solvent type
Phosphorescence is a so-called 'forbidden' process. It is a weaker process and occurs on a much longer timescale than fluorescence. Phosphorescent quantum yield is described by a similar equation to fluorescent quantum yield:
Phosphorescence spectrometers avoid fluorescence emission by making use of the longer lifetime for phosphorescent emission
The phosphorescence emission measurement is made when the excitation source is blocked
Relaxation by external conversion can be prevented by using the appropriate solvent and freezing in liquid nitrogen to produce an optically clear solid
Plasma Sources
The most common source for atomic emission is Inductively Coupled Plasma (ICP).
Purpose of source is to atomize and thermally excite the analyte atoms
ICP is very high temperature (10,000 K)
Plasma is formed by ionizing a stream of argon gas
Plasma is started (seeded) by a spark and maintained by a radio frequency generator
Heating is caused movement of electrons and ions under influence of oscillating rf field
ICP Torch
Consists of three concentric quartz tubes
Argon gas is introduced into each tube
Nebulizer gas; carries sample aerosol into the plasma
Plasma gas: forms the plasma
Auxillary or coolant gas; introduced tangentially to the outter tube to keep plasma in place
Sample introduction is achieved with a nebulizer and spray chamber
ICP Spectrometers
Modern ICP spectrometers have solid state detectors (similar to the photodoide array) and can measure multiple wavelengths simultaneously
Advantages of ICP are: good detection limits (ppb range), good linear dynamic range, few interferences, multiple element analysis
Disadvantages: more expensive that atomic absorbance instruments, more sample prep required compared to graphite furnace AA, high operating cost (uses lots of high purity argon), Graphite Furnace AA has lower detection limits for some elements
Brief Introduction to ICPMS
Not a spectroscopy technique (not measuring radiation) but uses the ICP
ICP used to atomize and ionize analyte
Analyte ions separated by mass and measured
Advantages: large element coverage (nearly whole periodic table), very low background signal (leads to low detection limits, ppt range), can measure isotopes
Disadvantages: high purchase and running cost, sample preparation required, few interferences but can be severe
Rayleigh (small particle) scattering
Large particle scattering
Origins of scattering
Two types:
-radiation is absorbed and emitted at the same wavelength (no loss of energy). Techniques include nephelometry and turbidity
- radiation absorbed and emitted at different wavelengths. Example technique is Raman spectroscopy
Elastic collisions classified as small (Rayleigh) or large particle scattering.
Rayleigh scattering occurs when particle's dimension is less than 5% of incident wavelength
Scattering intensity is proportional to incident radiation frequency to the fourth power
When detector is in line with the source, it is turbidimetry
When detector is an 90 degrees from the source, it is nephelometry
Instrumentation is similar to UV/vis spectrophotometry (for turbidimetry) and spectrofluorimetry (for nephelometry)
Turbidimetry or Nephelometry?
Depends on two factors:
Intensity of scattering radiation compared to incident radiation. Nephelometry is more appropriate for samples containing few scattering particles
Size of the scattering particle. More scattered radiation at 90 degrees occurs for small particles, therefore, nephelometry is more appropriate.
Calculating Concentration
In turbidimetry, transmittance (T) is a ratio of the transmitted intensity (It) divided by the incident intensity (I0):

The relationship between transmittance and concentration is similar to Beer's law:

In nephelometry, the concentration can be found by:
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