spectroscopy

SPECTROSCOPIC TECHNIQUES

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SPECTROSCOPY

Atoms and molecules interact with electromagnetic radiation (EMR) in a wide variety of ways.
Atoms and molecules may absorb and/or emit EMR.
Absorption of EMR stimulates different types of motion in atoms and/or molecules.
The patterns of absorption or emission are called ‘spectra’.
The field of spectroscopy is concerned with the interpretation of spectra in terms of atomic and molecular structure (and environment).

INFRARED SPECTROSCOPY

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INFRARED SPECTROSCOPY
Infrared radiation stimulates molecular vibrations.
Infrared spectra are traditionally displayed as %T (percent transmittance) versus wave number.
Useful in identifying presence or absence of functional groups.

¨IR range- 10cm- - 12800cm-

¨Sub regions
¨Near IR – 12500 – 4000cm-
¨Middle IR – 4000 – 667cm-
¨Far IR – 667 – 50 cm-
Finger print region – 900 – 1400cm- (identify the compound)

Water is a
-bent- non linear – triatomic molecule
Formula used for finding vibrations in non-linear molecule – (3N-6)
Carbon-di-oxide is a
– Linear – triatomic molecule
Formula used for finding vibrations in non-linear molecule - (3N-5)

INFRARED ABSORPTION BANDS

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INFRARED ABSORPTION BANDS

POSITION -REDUCED MASS- BOND STRENGTH (STIFFNESS)
LIGHT ATOMS HIGH FREQUENCY
STRONG BONDS HIGH FREQUENCY

STRENGTH -CHANGE IN ‘POLARITY’
STRONGLY POLAR BONDS GIVE INTENSE BANDS

WIDTH-HYDROGEN BONDING
STRONG HYDROGEN BONDING GIVES BROAD BANDS

INFRARED (VIBRATIONAL) SPECTROSCOPY

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POLARITY
Only vibrations that cause a change in ‘polarity’ give rise to bands in IR spectra.

Which of the vibrations for CO2 are infrared active?
Symmetric stretch
Asymmetric stretch
Bending (doubly degenerate)
Two types of stretching vibrations
Symmetric stretch
Asymmetric stretch

Four types of Bending Vibrations
¨Scissoring
¨Rocking
¨Twisting
¨Wagging

IR INSTRUMENTATION

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Source- Incandescent material

Other sources – Nernst glower , Globar , Carbon arc, Heated Nichrome

Monochromator – Quartz – prism

Detectors – Photo conductivity cell, Thermocouple, Pyroelectric detectors

APPLICATIONS OF IR SPECTROSCOPY

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—Identification of a substance (penicillin and peptides)
—Study of geometrical isomerism
—Characteristic of organic substances
—Study of photosynthesis and reparative in plants
—Assessment of purity of a sample
—Ideal and rapid method

NMR SPECTROSCOPY

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NMR spectroscopy is used to study a wide variety of nuclei:

1H
13C
15N
19F
31P

PRINCIPLES OF NMR.

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PRINCIPLES OF NMR

i) The nuclei of all elements carry a charge.

ii) When the spins of the protons and neutrons comprising these nuclei are not paired, the overall spin of the charged nucleus generates a magnetic dipole along the spin axis, and the intrinsic magnitude of this dipole is a fundamental nuclear property called the nuclear magnetic moment, µ.

NMR Signals, Tetramethyl silane

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NMR Signals

•The number of signals shows how many different kinds of protons are present.
•The location of the signals shows how shielded or deshielded the proton is.
•The intensity of the signal shows the number of protons of that type.
•Signal splitting shows the number of protons on adjacent atoms.

TETRAMETHYL SILANE

TMS is used as the internal standard
Solvents used are CDCl3,DMSO,CD3OD

Carbon-13

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Carbon-13

•12C has no magnetic spin.
•13C has a magnetic spin, but is only 1% of the carbon in a sample.
•Signals are weak, getting lost in noise.
•Hundreds of spectra are taken, averaged.

UV-Visible Spectroscopy

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Why should we learn this stuff? After all, nobody solves structures with UV any longer!

* Many organic molecules have chromophores that absorb UV
* UV absorbance is about 1000 x easier to detect per mole than NMR
* Still used in following reactions where the chromophore changes. Useful because timescale is so fast, and sensitivity so high. Kinetics, esp. in biochemistry, enzymology.
* Most quantitative Analytical chemistry in organic chemistry is conducted using HPLC with UV detectors
* One wavelength may not be the best for all compound in a mixture.
* Affects quantitative interpretation of HPLC peak heights.

Ultraviolet (UV) Spectroscopy

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Ultraviolet (UV) Spectroscopy – The Output

The output from a UV scanning spectrometer is not the most informative looking piece of data!! It looks like a series of broad humps of varying height.
Absorbance has no units – it is actually the logarithm of the ratio of light intensity incident on the sample divided by the light intensity leaving the sample.

There are two particular strengths of UV
(i) it is very sensitive
(ii) it is very useful in determining the quantity of a known compound in a solution of unknown concentration. It is not so useful in determining structure although it has been used in this way in the past.
The concentration of a sample is related to the absorbance according to the Beer Lambert Law.
Beer Lambert Law A = e.c.l
A = absorbance
c = concentration in moles l-1
l = pathlength in
e = molar absorptivity (also known as extinction coefficient) which has units of moles-1 L cm -1.

INSTRUMENTATION OF UV-VISIBLE SPECTROSCOPY

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INSTRUMENTATION

Radiation source
Wavelength selectors
Prisms – visible – glass prisms
ultraviolet – silica, quartz prisms
Sample holder – cells or cuvette
visible – glass
ultraviolet – fused silica cells
Solvent – ethyl alcohol

MASS SPECTROSCOPY

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COMPONENTS OF MASS SPECTROSCOPY

—1. Ionization chamber
—2. Analyzer
—3. Detector

MASS SPECTROSCOPY

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ESSENTIAL REQUIREMENTS OF MASS SPECTROSCOPY

Production of ions in gas phase = Cations (removing e-) and Anions (Adding e-)
* Acceleration to specific velocity
* Projection into suitable mass analyzer
* Detection of ions
Efficiency of cation depends - 1. Different Ionisation energy
2. Electron affinity

Uses of Mass spectroscopy

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Uses of Mass spectroscopy

1. Ionizing any material (in gas phase)
2. Investigation of any biological compounds

ELECTRON IMPACT IONISATION

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ELECTRON IMPACT IONISATION

1. Commonly used and simple source
2. Used for metabolic studies, drug studies

CHEMICAL IMPACT IONISATION

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CHEMICAL IMPACT IONISATION

Determine molecular mass
Production of high intensity molecular or pseudo molecular ions.
Generation of ions gives species like CH3+.
Suitable reagent gas (methane) used
Uses: study of drugs

Isotope Ratio Mass Spectrometry

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IRMS

Used in biological field
Used for metabolic studies
Used for the analysis of Human Breath
EI source, magnetic sector, mass analyser & Faraday cup detector
m/Z = B2R2/2V

FAST ATOM BOMBARDMENT

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FAST ATOM BOMBARDMENT

•FAS
•Introducing ionising beam into materials
•Liquid matrices used
•Solid surfaces damaged by high intensity beam
•Collision occur between fast moving ions & slow moving ions
•Use- Fragmentation of peptides.

PYROLOSIS MASS SPECTROMETRY

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PYROLOSIS MASS SPECTROMETRY

Materials subjected to high T
Ejection of volatile substances
EI source, mass analyzer
Certain T allowed
Studying the effects of different growth conditions, composition of cell walls.

ANALYSERS

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ANALYSERS USED IN MASS SPECTROMETERS

1. MAGNETIC SECTOR ANALYSER
2. ELECTRIC SECTOR ANALYSER
3. QUADRUPOLE MASS FILTERS

DETECTORS

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VARIOUS DETECTORS

1. FARADAY CUP DETECTORS
2. ELECTRON MULTIPLIER
3. ARRAY DETECTOR