 Hello, welcome to module 1 of the course
on Application of Spectroscopic Methods in
Molecular Structure Determination. In this
module, you will ask questions about what
is spectroscopy? What is a spectrum? What
is electromagnetic radiation? What are the
various regions of the electromagnetic radiations
and when matter is interacting with electromagnetic
radiation, what are the changes that take
place in the matter? What is the nature of
light? What is absorbance? What is transmission?
All these terminologies that are used in spectroscopy,
we will deal within this general module in
this particular presentation.
Now, spectroscopy in the broadest term is
defined as the study of interaction of electromagnetic
radiation with matter and when we talk about
matter we are dealing with atoms and molecules.
So, essentially the interaction of atoms and
molecules with electromagnetic radiation is
what constitutes the term spectroscopy. Spectroscopy
is a study of interaction of electromagnetic
radiations with matter as a function of frequency
because whenever we record a spectrum, we
are essentially scanning various frequencies
and recording the response of the material
that we study for the various frequencies
of the electromagnetic radiation. Spectroscopy
is the study of the exchange of energy between
electromagnetic radiation and matter. Here,
we are talking about the electromagnetic radiation
being the source of energy, the energy of
the photon of the electromagnetic radiation
gets transferred on to the atoms and molecule
and the changes that occur in the atoms and
molecule as result of the absorption of energy
is what is recorded as a spectrum.
Here is an electromagnetic spectrum dealing
with various frequencies and various energies
of the photons and the different regions of
electromagnetic spectrum. The regions of the
electromagnetic spectrum are labeled here
as radio waves, micro waves, infrared waves,
visible region of the electromagnetic spectrum,
ultraviolet region of the electromagnetic
spectrum, the soft and the hard x-rays. Finally,
the most powerful energetic gamma rays in
the right-hand side of the spectrum. Now,
the top scale corresponds to wavelength which
is expressed in meters which is a unit of
length. Just to understand what is this unit
of length actually means in terms of the dimensions
of commonly seen objects, we can for example;
correlate the wavelength of radio frequency
region corresponding to something like 10
meters to about 1000 meters corresponding
to the size of a building or size of a stadium
and so on. On the other hand, if you look
at the size of a molecule which is of the
order of a pico meter or so. We are talking
about the dimensions of the wavelength of
the x-rays or the gamma rays in this particular
region. Suppose, if we take a tiny dot on
a piece of paper which corresponds to about
a millimeter or less than a millimeter that
corresponds to the micro wave radiation region
of the electromagnetic spectrum. So, just
sort of gives you a perspective of what this
unit of dimension in terms of the length unit
corresponds to commonly observed objects on
our day to day life. In the bottom, the frequency
scale is mentioned. Frequency units are waves
per seconds, in other words per second is
the unit that is normally used for the frequency
scale. As you can see here the frequencies
starts from 10 to the power 6 per second to
about 10 to the power 20 per second in the
highest frequency region which is a gamma
ray region. As the frequency increases the
energy content also increases because frequency
is directly proportional to energy in terms
of E is equal to h nu, where nu is the frequency
of the electromagnetic radiation.
The last scale corresponds to energy per photon
of the electromagnetic region, spectral regions.
For example, we are talking about something
like 10 to the power minus 9 electron volt.
Electron volt is the energy unit that in which
this is this particular scale is expressed
10 to the power minus 9 corresponds about
to a nano electron volt is what we are talking
about a very low energy in terms of the energy
content of a photon of a radio frequency wave.
On the other hand, if you come to the gamma
rays it is about 10 to the power 6, one mega
electro volt is what we are talking about
and this is a high energy radiation, normally
we call it as ionizing radiation because they
ionize substances through which they pass
through.
So, we have a wide range of energy content
per photon of the various regions of the electromagnetic
radiation depending upon what kind of a radiation
that is being used. Different processes take
place in atoms and molecules and the response
of the atom and molecules for the various
frequency regions is what is recorded as a
spectrum.
In the electromagnetic spectrum, there is
a very small portion what is known as the
visible portion of the electromagnetic radiation
and this is the only portion to which human
eye is sensitive. In other words, we will
be able to perceive only the colors of this
particular region and not see the colors or
not see the other regions like for example,
the ultraviolet or the infrared spectroscopy.
They are invisible to the human eye. Human
eye is sensitive to only about 380 or 390
nanometer, which is the violet region of the
electromagnetic spectrum to about 760 or 780
nanometers, which is the red region of the
electromagnetic spectrum. This is very important
to know because when we talk about the UV
visible spectroscopy, you will be dealing
with this particular spectral region in the
UV visible spectroscopic technique.
Having defined spectroscopy as a study of
interaction of electromagnetic radiation with
matter, now we have to depending upon the
region of the electromagnetic spectrum that
is used, what are the various processes that
can take place in atoms and molecule? How
does one receive information in these processes?
And what are the spectroscopic techniques
that corresponds to various wavelength regions
of the electromagnetic spectrum?
This is actually defined in this particular
table. If you look at column number one, the
region of the electromagnetic radiation, energy
per photon is given starting from gamma rays
which are the powerful rays to the radio waves
which of the least energetic rays that we
are dealing with in this particular table.
Now, the processes that occur in molecules
and atoms corresponding to this particular
energy regime is what is mentioned on this
particular column and the spectroscopic technique,
corresponding spectroscopic techniques are
also given in red. Now, if we take for example,
gamma rays, gamma rays are very powerful high
energy photons and they can cause transitions
within the nucleus causing the change of nuclear
configuration and this essentially constitutes
the Mossbauer spectroscopy.
On the other hand, if we come to microwave
which are very, very low energy radiation
the transition among the rotational levels
of a molecule is what is happening in the
case absorption of the microwave radiation
and this essentially constitutes the rotational
spectroscopy of the electromagnetic spectrum
corresponding to the microwave radiation is
called the rotational spectroscopy. Now, if
we talk about radio wave frequency region
of the electromagnetic radiation then we are
talking about change in the nuclear or the
electron spin in the presence of the external
magnetic field. In the presence of an external
magnetic field, nuclear spins and electron
spins have different energies, ground state
and excited state and the cause of spin change
from the ground state to the excited state
is what is normally absorbed during the electron
and the nuclear spin resonance electron spectroscopy.
NMR and ESR are the techniques which are responsible
for this and we will deal with NMR in this
particular spectroscopic course in much more
detail.
This particular slide tells us something about
the responses upon interaction of electromagnetic
radiation. In other words, when a sample is
being excited by electromagnetic radiation,
what are the processes that can occur in the
sample? Now, let us consider a incident light
falling on the sample in this particular direction.
I(0) is the intensity of the incident light
that is falling on this substance. The substance
can absorb certain amount this light and then
transmit the remaining light into the other
side, I is the intensity of the transmitted
light.
Let us say for example, so I (0) minus I would
correspond to the absorbance or the intensity
that is being absorbed by the sample itself.
Instead of absorbing the light the sample
can simply scatter the light for example,
if it is scatters the light and if it is the
light of the same wavelength then we call
it as Rayleigh scattering. If it scatters
light of different wavelength, then we call
it as a Raman scattering. Raman spectroscopy
is based on the scattering phenomena and we
deal with Raman spectroscopy along with our
vibrational spectroscopy for the structural
elucidation purposes. Now, instead of the
absorbed energy instead of being scattered
and so on, the molecule can also emit light
for example, in the form of light emission
which corresponds to the emission spectroscopy.
We have fluorescence spectroscopy and phosphorescence
spectroscopy as emission spectroscopic techniques
corresponding to this. Now, let us define
certain terminologies that are used in the
area of spectroscopy. The ratio of I by I(0),
in other words the intensity of transmitted
light to the intensity of the initial light
is what is known as the transmittance or capital
T is the symbol that is used for this particular
terminology.
Now, logarithm scale of I(0) divided by I
[logI(0)/I] is what is known as the absorbance,
capital A is what is used for as the symbol
for this particular term and absorbance transmittance
are related to each other as you can see for
example, one is the logarithmic quantity which
is logarithm of 1 by T (log(1/T) is what is
known as the absorbance. Now, the type of
spectroscopic techniques that one can have
depends on the kind of phenomena that one
observes. If the absorbance of the light is
what is being observed, then we call it as
Absorption spectroscopy. If emission is what
is observed, then we call it as the emission
spectroscopy. If scattering is what is being
observed then we call it as scattering spectroscopy,
examples of Absorption spectroscopy are the
infrared, UV visible spectroscopy, rotational
spectroscopy and so on. Emission spectroscopy
corresponds to fluorescence and phosphorescence
spectroscopy and scattering spectroscopy for
example, corresponds to Raman spectroscopy.
Now, let us ask this question what is a spectrum?
A spectrum is essentially a plot of energy
in the x-axis and the response that is being
received from the sample as y-axis The energy
is the energy of the electromagnetic radiation
that is being applied in this particular case
and the response is the kind of response that
one records in terms of whether it is an absorbance
or transmittance or emission intensity or
scattering intensity is what is being recorded
on the y-axis and the peaks that are normally
seen in the spectrum are essentially due to
the intensities of absorption or transmittance
or intensities of emission or scattering is
what is plotted against the energy of the
electromagnetic radiation.
Certain laws are governing the quantitative
aspects of spectroscopy is important to understand.
One is called the Beer Lambert law, is a very
basic law dealing with the quantitative correlation
between absorbance and the concentration.
Now, this expression is what is known as the
Beer Lambert law, absorbance which is actually
logarithmic ratio of I(0) by I is directly
proportional to the concentration. In other
words, when light is passing through a medium
it depends on how many number of molecules
that it encounters, absorbance will be accordingly
more or less. In other words, absorbance is
directly proportional to the number of molecules
the light interacts on it is path. Now, the
proportionality constant is what is known
as extinction coefficient or the molar absorptivity
and extinction coefficient or the molar absorptivity
is a constant at a given wavelength for a
given substance and this is what makes the
absorption spectroscopy a quantitative tool
in order to find concentration of unknown
substances.
Now, let us have a look at the nature of light.
Light can be described to have dual nature
that is both the nature of wave as well as
particle. In the wave nature of light, it
is actually explained in the form of this
particle diagram that is shown here. If x
is the direction of propagation of the light
then we can define two fields, one is an electric
field another one is the magnetic field. The
blue one is the magnetic field and the red
one is the electric field. Thus, you can see
here the two fields, namely the electric field
and magnetic field lie orthogonal to each
other. In other words, they are perpendicular
to each other and light can be expressed in
the form of a sine wave consisting of two
sine waves, interacting sine waves for example,
one corresponding to the electric field, the
other one corresponding to the magnetic field.
The distance between the hump to the hump
is what is known as the Wavelength and this
wavelength is what we referred in terms of
the electromagnetic spectrum that earlier
we saw in terms of the defining the regions
of the electromagnetic spectrum with the different
Wavelengths and the arrows that are shown
here are the vectors of the intensity or the
amplitude of the electric field in this particular
case. The amplitude of the magnetic field
is shown in the blue color. So, this is essentially
the wave nature of light is being explained
in this particular manner and whenever molecules
interact with electromagnetic radiation in
spectroscopy like for example, a electronic
spectroscopy or infrared spectroscopy or rotational
spectroscopy it is supposed to be the electric
field that is interacting with the molecules
in those kinds of spectroscopy. Unlike for
example, in the case of magnetic resonance
spectroscopy it supposed to be the magnetic
field that is interacting with the magnetic
nuclei. So, it is in the case of NMR and ESR
essentially the magnetic field is what is
interacting with the molecules.
In order to explain the other nature namely
the corpuscular nature of the light one proposes
that light consists of particles called photons
with finite energy content. In other words,
E is equal to h x nu this is very famous expression
which deals with the energy of photon which
corresponds to the h which is the Planck's
constant and nu the frequency of the electromagnetic
radiation, which is correlated to the velocity
of light in vacuum namely c for example, divided
by lambda. In other words, E is equal to h
x nu which is equal to the h x c by lambda,
where lambda is the wavelength of the light
that we are dealing with.
In order to calculate the amount of energy
that is present in a photon and one can use
this expression very easily. Let us have an
example of how much of a photon energy is
present in a 500-nanometer wavelength region
which is the visible wavelength region for
example. So, all you have to do is plug in
the Planck’s constant which is available
readily which is 6.626 into 10 to the power
minus 34 joules per second and put in the
value of the velocity of the light which is
about approximately 3 into 10 to the power
8 meters per second divided by 500 nanometers,
which is 500 into 10 to the power minus 9
meters.
Now, the units which are the time units they
cancel out each other. The distance unit namely
the meters they cancel each other finally,
you are ending up with the energy which is
joules in this particular case. So, the energy
content of a photon of a 500-nanometer wavelength
light is about 3.98 into 10 to the power minus
19. Light of 297 nanometers which is in the
ultraviolet region has much higher energy.
In fact, this is about 400 kilo joules per
mole. This is per mole is what is expressed,
in other words, if you want to convert per
photon energy into a per mole energy you have
to multiple this quantity by per photon quantity
by Avogadro number which is 6.023 into 10
to the power 23. So, if you multiply the per
photon energy by Avogadro number you get the
energy per mole of the corresponding wavelength.
Now, according to the quantum mechanics, energy
levels are quantized. We are talking about
electronic energy levels or vibration energy
levels or rotational energy levels they are
quantized. What is meant by quantization?
The energy gap between the ground state and
the excited state is a finite one. When we
say it is a finite one then, the energy difference
and the energy of the incident photon matches
with the energy difference ground state and
the excited state one can expect the absorption
of the light to take place. In other words,
E1 let us say ground state energy and E2 is
the excited state energy of certain system
it could be a electronic energy or it could
be vibrational energy or it could be rotational
energy as the case may be. Now, the difference
in the energy is delta E and the delta E is
equal to h nu. So, when the nu corresponds
the electromagnetic radiation of certain frequency
corresponds to the difference in the energy
between the ground state and excited state,
absorption is going to take place.
Otherwise, if there is a different frequency
which does not match this particular criterion
of delta E, then the absorption will not take
place that is what is implied in this particular
diagram.
Now, once you say there are two energy levels,
there should be a population in this energy
level of molecules and there should be a population
in this energy level in this molecule. One
can easily calculate the difference in the
population or the ratio of the population
in the excited state to the ground state using
the Maxwell-Boltzmann distribution. Maxwell-Boltzmann
distribution is expressed by this particular
equation here, n2 by n1 is the ratio of the
number of molecules in the excited state n2,
n1 is the number of molecules in the ground
state and this ratio corresponds to the delta
E divided by kT in terms of exponential term
being added here. Now, in the Boltzmann distribution
one can calculate the ratio of the excited
state to the ground state using this particular
equation.
When you do that using the Maxwell-Boltzmann
distribution expression, one can show that
at equilibrium we are talking about thermal
equilibrium. Let us say at room temperature
less than one percent of the molecules are
present in the excited state vibrational level
at room temperature that is we are talking
about the vibrational spectroscopy. When we
say there is a ground state vibration and
in the excited vibration state. Nearly 99.9
percent of the molecules do exist in the lower
vibrational level, compared to the higher
vibrational level and when we come to electronic
energy states when we talk about ground state
electronic state to the excited state electronic
state because electronic states have much
higher energy gap practically all the molecules
will be present in the ground state compare
to the excited state and this is very important
to understand because this kind of population
difference is directly responsible for the
sensitivity of the technique that we are talking
about.
Now, let us look at the NMR spectroscopy or
ESR spectroscopy, where we are talking about
nuclear spin state, ground state and excited
state. The energy difference between the two
states are extremely small compared to the
energy states of the electronic spectroscopy
or the infrared spectroscopy. So, as a result
of that the population difference is extremely
low only parts per million difference is present
in the population difference is present between
the ground state and the excited state and
that makes the NMR spectroscopy as the least
sensitive spectroscopy techniques that one
can think of compare to for example, the electronic
spectroscopy or the vibrational spectroscopic
technique. So, the population difference is
directly responsible for the sensitivity because
the intensity of absorption is directly proportional
to the excess population that is present in
the ground state in comparison to the excited
state. So, when the entire population is in
the ground state this spectroscopic technique
will be very sensitive because we have a large
number of molecules to excite from the ground
state to excited state.
On the otherhand when you have already reached
equal population between the excited state
and the ground state there is very little
excess population present in the ground state
to be excited to the higher level. So, accordingly
the intensity of absorption will be low. So,
the sensitivity will be consequently low in
the case of NMR and other spectroscopy techniques.
Now, let us define another parameter what
is known as the natural spectral line width.
Let us say, if the energy levels are quantized
then the absorption and the emission should
occur in a precise frequency. In other words,
a monochromatic frequency is what would be
responsible for each absorption and emission
wavelengths that we are talking about, but
usually this does not happen. There is a measurable
width to any spectral line which is known
as the natural line width of the spectroscopy
and this is defined as shown in this particular
diagram. This is essentially a lambda plotted
against the relative intensity you can see
that the spectral line has a particular shape
usually at Gaussian shape and the if we take
half the intensity and measure the width of
the spectral line here, this is what is known
as the full-width half-maximum spectral width
and this is what is normally known as the
natural spectral line width.
Now, why do we have a natural spectral line
width instead of simply line kind of a spectrum?
The finite lifetime of the excited state is
what is responsible for the line width. In
other words, the finite lifetime of the excited
state and the consequent uncertainty in the
excited state energy is what gives the line
space for many of the spectroscopic techniques.
Now, the uncertainty principle is expressed
in a different format in this particular case.
Normally, uncertainty principle is expressed
in terms of momentum and position where you
can have either uncertainty in the momentum
or uncertainty in position. In this particular
case, this is expressed as the energy and
lifetime in other words this is could be the
uncertainty in the difference in the energies
or uncertainty in the energy and this will
be the lifetime of the system.
Now, when we talk about for example, lifetimes
of the excited state or the relaxation times,
if you are talking about ultraviolet spectroscopy
for example, the relaxation times are the
typically of the order of the pico seconds
or faster. So, there is a large uncertainty
associated with a excited state energy level
and this uncertainty in the excited state
energy level is what causes the line broadening
which leads to the natural line width of the
UV visible spectroscopy. On the other hand,
if you look at NMR spectroscopy, where the
relaxation times of the order of the seconds
much slower than the relaxation times of the
electronic spectroscopy, the uncertainty in
the energy of the excited spin state is very,
very low as the result of that you get sharp
peaks less than one hertz width natural line
width what we get in the case of NMR spectroscopy.
Now, so far we have been talking about spectroscopic
technique which have some commonality in terms
of either being absorbance or the emission
kind of a spectroscopy, where electromagnetic
radiation is used as a source of energy for
causing certain types of transitions. It could
be electronic transition, it could be spin
transition or it could be vibrational transition
and so on. The commonality of all the spectroscopic
that we talked about is very different when
we compare it with for example, Mass spectrometry.
First of all, mass spectrometry is not a spectroscopic
technique at all. It is a spectrometry technique
there is no interaction between electromagnetic
radiation and sample in mass spectrometry,
is a very special kind of technique. It is
an extremely useful technique for molecular
structure determination that is why it is
normally put under the spectroscopy many spectroscopy
books talk about mass spectrometry as well
because it is useful in structural elucidation
problems. In mass spectrometry, we are talking
about interaction between a sample that is
an atom or molecule and an electron which
is usually high energy electron of the order
of 70 electron volts or 100 electrons volts,
leading to ionization of the sample fragmentation
of the sample and so on. We will deal with
the mass spectrometry slightly later and just
wanted to bring out the difference between
the various spectroscopy and mass spectrometry
in terms of the, this spectroscopy being spectrometry
being a special case and it comes under the
spectroscopy essentially because of it is
useful not because of it is commonality with
the other spectroscopic methods.
Now, let us see what are the resources that
we need to follow this particular program.
At least one or more books need to be referred
to. This book which was published last year
that is in 2015 by Prof. D. N. Sathyanarayana,
who was a former professor in the Department
of Inorganic and Physical Chemistry in Indian
Institute of Science, Bangalore. This is a
very nice book, it deals with various spectroscopic
techniques right from radio waves to gamma
rays for example, in other words he is covering
the nearly the entire electromagnetic radiation
region in this particular book. It gives very
fundamental aspects of the many of the spectroscopic
techniques with suitable examples and so on.
I would recommend that you read this book
for clear understanding of the basics of these
spectroscopic.
This is another book by name it is Spectroscopy
by Pavia and others. There are four other
authors associated with this particular book.
This is essentially a spectroscopy book with
emphasis on problem solving, in other words
structural elucidation problem solving is
what is the emphasis that is shown in this
spectroscopy book. It is an excellent text
book it is available in Indian edition should
be possible for some of you to buy this book
and refer this particular book.
NMR Spectroscopy by Herald Gunther is a very
famous text book for example, it is very lucidly
presented text book in the terms of the concepts
and applications of NMR spectroscopy. So,
this spectroscopy book is also going to be
very useful for this particular course.
This particular book namely Spectroscopic
Identification of Organic Compounds by Silverstein
and Webster is actually used to be Silverstein
and Basler. Now, it is called as Silverstein
and Webster in terms of the authors names
and this is published in 2006. This is also
an excellent source of problem solving sessions
in the spectroscopy.
Finally, the Organic Spectroscopy by William
Kemp is also a good source of information
particularly, if you are dealing with organic
structural which we will be doing in this
particular course.
Now, I would like to thank you for patient
hearing. We will see you in the next module.
Thank you.
