Spectroscopy

THE ELECTROMAGNETIC SPECTRUM

 WHAT IS 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 (wavelengths absorbed
and to what extent) and/or emission (wavelengths
emitted and their respective intensities) are called
‘spectra’.
• The field of spectroscopy is concerned with the
interpretation of spectra in terms of atomic and
molecular structure (and environment).
Properties of ElectroMagnetic Radiation (Light)

1) Amplitude (A) – height of wave’s electric vector

2) wavelength (l) – distance (nm, cm, m) from peak to

peak
a) wave number (ሷ ) = 1/l (cm-1)
 3) frequency (n) – number of cycles or oscillations per second
a) hertz (Hz) or s-1.

4) velocity of propagation (vi) – rate of travel through space,

dependent on composition of medium
a) vi = nli
b) maximum velocity (c) – speed of light in a vacuum (3.00
x108 m/s)
c) slower in other media (~ 0.03% slower in air)
 UV-Visible Spectroscopy
- Also known as electronic spectroscopy
- Range is from 10nm-780nm (390-780nm vis)
- Involves electronic transitions.
- Used in analytical chemistry for the
quantitative determination of transition metal
ions, highly conjugated organic compounds
and biological macromolecules.
- Is commonly carried out in solutions but solids
and gases may also be studied.
 Compounds studied under UV-Visible
• Organic molecules such as conjugated
compounds and biomolecules
• Complex molecules.
• Electronic transitions in organic molecules
 Chromophores and Auxochromes
Chromophore: The molecule which is responsible
for the light absorption in the visible region.
-Common examples include retinal (used in the eye
to detect light), various food colorings, fabric dyes
(azo compounds), pH indicators, lycopene, β-
carotene, and anthocyanins.
Auxochrome: substituent which leads to shift of
the absorption maximum .
-Examples include the hydroxyl group (−OH), the
amino group (−NH2), the aldehyde group (−CHO),
and the methyl mercaptan group (−SCH3)
 UV- Visible Spectrometer
• Instrument is called a UV/Vis spectrophotometer.
• It measures the intensity of light passing through a
sample ( I), and compares it to the intensity of light
before it passes through the sample (Io). The ratio (I/Io)
is called the transmittance, and is usually expressed as
a percentage (%T).
Instrumentation
The absorbance is plotted against wavelength
 Quantitative analysis
• The basis for using spectrophotometric measurements
to quantitatively analyze a light-absorbing chemical
species, in solution is the Beer-Lambert law:

• A=εbc where A is absorbance at a given wavelength, ε is

the molar absorptivity (also known as molar extinction
coefficient), b is the distance the light travels through
the solution (called the pathlength), and c is the
concentration of the analyte in solution.

• The Beer-Lambert law simply states that absorbance is

directly proportional to the concentration of analyte in
the sample.
 Applications of UV-Visible
• Qualitative analysis
• Quantitative analysis
• Detection of auxochromes
Infrared Spectroscopy

- based on IR absorption by molecules.

- Molecules undergo vibrational and rotational transitions
during absorption of IR radiation.
rotational transitions
Potential Energy (E)

Vibrational transitions

Interatomic Distance (r)

Potential energy resembles classic Harmonic Oscillator

 IR radiation is in the range of 12,800 – 10 cm-1 or l = 0.78 – 1000 mm
- rotational and vibrational transitions often occur together

Typical IR spectrum for Organic Molecule

% Transmittance

Wavenumber (cm-1)
Types of Molecular Vibrations
Bond Stretching Bond Bending

symmetric In-plane rocking

asymmetric In-plane scissoring

Out-of-plane wagging

Out-of-plane twisting
 symmetric asymmetric In-plane scissoring

Out-of-plane twisting In-plane rocking Out-of-plane wagging

Instrumentation
Basic Design
- normal IR instrument similar to UV-vis
- main differences are light source & detector
Application of IR
Qualitative Analysis (Compound Identification)
- main application
- Use of IR, with NMR and MS, in late 1950’s revolutionized organic
chemistry
► decreased the time to confirm compound identification
10 -1000 fold
General Scheme
1) examine what functional groups are present by looking at group
frequency region
- 3600 cm-1 to 1200 cm-1
Group Frequency Region
- approximate frequency of many functional groups (C=O,C=C,C-H,O-H)
can be
calculated from atomic masses & force constants
- positions changes a little with neighboring atoms, but often in same
general region
- serves as a good initial guide to compound identity, but not positive proof.
Abbreviated Table of Group Frequencies for Organic Groups
Bond Type of Compound Frequency Range, cm-1 Intensity
C-H Alkanes 2850-2970 Strong
C-H Alkenes H 3010-3095 Medium
C C
675-995 strong

C-H Alkynes C C H 3300 Strong

C-H Aromatic rings 3010-3100 Medium

690-900 strong
0-H Monomeric alcohols, phenols 3590-3650 Variable
Hydrogen-bonded alchohols, phenols 3200-3600 Variable, sometimes broad
Monomeric carboxylic acids 3500-3650 Medium
Hydrogen-bonded carboxylic acids 2500-2700 broad
N-H Amines, amides 3300-3500 medium
C=C Alkenes 1610-1680 Variable
C=C Aromatic rings 1500-1600 Variable
C C Alkynes 2100-2260 Variable
C-N Amines, amides 1180-1360 Strong
C N Nitriles 2210-2280 Strong
C-O Alcohols, ethers,carboxylic acids, esters 1050-1300 Strong
C=O Aldehydes, ketones, carboxylic acids, esters 1690-1760 Strong
NO2 Nitro compounds 1500-1570 Strong
1300-1370
Fingerprint Region (1200-700 cm-1)
- region of most single bond signals
- many have similar frequencies, so affect each other & give pattern characteristics of
overall skeletal structure of a compound
- exact interpretation of this region of spectra seldom possible because of complexity
- complexity  uniqueness Fingerprint Region
2.) Quantitative Analysis
- not as good as UV/Vis in terms of accuracy and precision
► more complex spectra
► narrower bands (Beer’s Law deviation)
► limitations of IR instruments (lower light throughput, weaker
detectors)
► high background IR
► difficult to match reference and sample cells
► changes in e (A=ebc) common
- potential advantage is good selectivity, since so many compounds have
different IR
spectra
► one common application is determination of air contaminants.
Example 9: The spectrum is for a substance with an empirical formula of C3H5N. What is
the compound?

Nitrile or No aromatics
Aliphatic
alkyne group One or more
hydrogens
alkane groups
 NMR Spectroscopy
• When placed in a magnetic field, NMR active nuclei (such
as 1H or 13C) absorb electromagnetic radiation at a
frequency characteristic of the isotope.
• The resonant frequency, energy of the radiation absorbed,
and the intensity of the signal are proportional to the
strength of the magnetic field.
• For example, in a 21 Tesla magnetic field, hydrogen atoms
(commonly referred to as protons) resonate at 900 MHz. It
is common to refer to a 21 T magnet as a 900 MHz magnet
since hydrogen is the most common nucleus detected,
however different nuclei will resonate at different
frequencies at this field strength in proportion to their
nuclear magnetic moments.
• A spinning charge generates a magnetic field that results in
a magnetic moment proportional to the spin. In the
presence of an external magnetic field, two spin states exist
(for a spin 1/2 nucleus): one spin up and one spin down,
where one aligns with the magnetic field and the other
opposes it. The difference in energy (ΔE) between the two
spin states increases as the strength of the field increases,
but this difference is usually very small, leading to the
requirement for strong NMR magnets (1-20 T for modern
NMR instruments). Irradiation of the sample with energy
corresponding to the exact spin state separation of a
specific set of nuclei will cause excitation of those set of
nuclei in the lower energy state to the higher energy state.
• For spin 1/2 nuclei, the energy difference between the two
spin states at a given magnetic field strength is proportional
to their magnetic moment. However, even if all protons
have the same magnetic moments, they do not give
resonant signals at the same frequency values. This
difference arises from the differing electronic environments
of the nucleus of interest. Upon application of an external
magnetic field, these electrons move in response to the
field and generate local magnetic fields that oppose the
much stronger applied field. This local field thus "shields"
the proton from the applied magnetic field, which must
therefore be increased in order to achieve resonance
(absorption of rf energy). Such increments are very small,
usually in parts per million (ppm).
Instrumentation
 Mass spectrometry

• Mass spectrometry (MS) is an analytical

technique that ionizes chemical species and
sorts the ions based on their mass-to-charge
ratio. In simpler terms, a mass spectrum
measures the masses within a sample.
• Mass spectrometry is used in many different
fields and is applied to pure samples as well as
complex mixtures.
• These spectra are used to determine the
elemental or isotopic signature of a sample,
the masses of particles and of molecules, and
to elucidate the chemical structures of
molecules, such as peptides and other
chemical compounds.
 Atomic Absorption Spectrometry
• is a spectroanalytical procedure for the quantitative
determination of chemical elements using the
absorption of optical radiation (light) by free atoms in
the gaseous state.

• In analytical chemistry the technique is used for

determining the concentration of a particular element
(the analyte) in a sample to be analyzed. AAS can be
used to determine over 70 different elements in
solution, or directly in solid samples via electrothermal
vaporization,[citation needed] and is used in
pharmacology, biophysics and toxicology research.
• The technique makes use of absorption
spectroscopy to assess the concentration of an
analyte in a sample. It requires standards with
known analyte content to establish the
relation between the measured absorbance
and the analyte concentration and relies
therefore on the Beer-Lambert Law.
Instrumentation