Title of the course:

Advanced
Pharmaceutical Analyses
 Reference text:

• 1. Spectrometric Identification of Organic Compounds by Silverstein,

Bassler and Morrill;
• 2. Applications of absorption spectroscopy of organic compounds by
Dyer JR. 3. Organic Chemistry by McMurry; 5thed; Thomason
learning CA, USA 2000.
 Lecture title

• UV / visible spectroscopy; Sample handling and instrumentation;

Characteristic absorption of organic compounds; Rules for calculation of
lambda max and application; Application of UV/visible; spectroscopy;
Problems and solutions.
 Spectroscopy
• Spectroscopy is the study of the absorption and emission of light and
other radiation by matter.
 Electromagnetic radiation(EMR)

is a form of energy whose • .

behavior is described by the
properties of both waves and
particles
 Spectrophotometry
• Spectrophotometry is a method to measure how much a chemical substance
absorbs light by measuring the intensity of light as a beam of light passes
through sample solution.
• In spectrometric methods, the sample solution absorbs electromagnetic
radiation from an appropriate source, and the amount absorbed is related to
the concentration of the analyte in the solution.
 Interaction of Electromagnetic Radiation
with Matter
• The most interesting types of interactions in spectroscopy
involve transitions between different energy levels of
chemical species.
• Molecules absorb energy and this energy can bring about
translation, rotational, or vibration motion or ionization of
the molecules depending upon the frequency of
electromagnetic radiation they receive. Excited molecules
are unstable and quickly drop down to ground state again
giving off the energy they have received as electromagnetic
radiation.
 There are three basic processes by which a molecule
can absorb radiation:
First:- Second:- Third:-
The molecule rotates the atoms or groups of atoms the outer shell electrons
about various axes, the within a molecule vibrate of a molecule, valence
energy of rotation being relative to each other, and the
at definite energy levels, energy of this vibration occurs
electrons, may be raised
so the molecule may at definite quantized levels. The to a higher electron
absorb radiation and be molecule may then absorb a energy, corresponding
raised to a higher discrete amount of energy and to an electronic
rotational energy level, in be raised to a higher vibrational transition.
a rotational transition energy level, in a vibrational
transition.
 Interaction of Matter with radiation
• Absorption: A transition from a lower level to a higher level with transfer of energy from
the radiation field to an absorber, atom, molecule, or solid.
• Emission: A transition from a higher level to a lower level with transfer of energy from
the emitter to the radiation field. If no radiation is emitted, the transition from higher to
lower energy levels is called nonradioactive decay.
• The data that is obtained from spectroscopy is called a spectrum.
• A spectrum is a plot of the intensity of energy detected versus the wavelength (or mass or
momentum or frequency, etc.) of the energy. A spectrum can be used to obtain
information about atomic and molecular energy levels, molecular geometries, chemical
bonds, interactions of molecules, and related processes. Often, spectra are used to identify
the components of a sample (qualitative analysis).
• Electromagnetic radiation possesses' a certain amount of energy. The energy of a
unit of radiation, called the photon, is related to the frequency or wavelength by

• h= blank constant =6.626×10ˉ34 J.s

• Speed of light is 3.00 × 108 m/s
• The wavelength(ʎ) • The frequency (v)
• Is the distance between equivalent points • Is the number of cycles of
on the wave radiation passing a point in
train
space
• Wavelengths in the ultraviolet and visible per second. It is expressed as s-
regions are on the order of
nanometers. In the infrared region, they 1, or hertz (hz).
are micrometers, but the reciprocal
of wavelength is often used (wave
numbers, in cm-1
• The definitions show that the relation between these quantities is:
• C=vʎ

sometimes the wavenumber,

, is used where
 Example :
• The wave length of sodium d-line
is 589 nm . calculate the frequency
and wave number of line .
 UV-VIS Spectroscopy
• The ultraviolet region extends from about • The visible (VIS) region is actually a
10 to 380 nm, but the most analytically very small part of the
useful region is from 200 to 380 nm, called electromagnetic spectrum, and it is
the near-ultraviolet or quartz UV region. the region of wavelengths that can be
• Below 200 nm, the air absorbs and so the seen by the human eye, that is, where
instruments are operated under a vacuum; the light appears as a color. The
hence, this wavelength region is called the visible region extends from the near-
vacuum-ultraviolet region. ultraviolet region (380 nm) to about
780 nm.
 UV-VIS Spectroscopy
• Studies the electronic transitions of molecules as they absorb light in the UV and
visible regions of the electromagnetic spectrum the data is used to produce
absorbance spectra.
• (U.V region 10-400)
• (VIS. region 400-800)
• Organic molecules and other molecules
the electronic transitions in organic compounds and some other compounds can be
determined by ultraviolet-visible spectroscopy, provided that transitions in the
ultraviolet (UV) or visible range of the electromagnetic spectrum exist for this
compound.
 Spectrochemical methods
• Spectrochemical methods
visible spectrometry is one of the most widely used methods of analysis. It is
very widely used in clinical chemistry and environmental laboratories because
many substances can be selectively converted to a colored derivative .
 Spectral colors
• Colours that can be produced by visible light of a narrow band of
wavelengths (monochromatic light) are called pure spectral colours. This
"visible light" corresponds to a wavelength range of 400 - 800 nanometers
(nm) and a colour range of violet through red. The human eye is not capable
of "seeing" radiation with wavelengths outside the visible spectrum. The
visible colours from shortest to longest wavelength are: violet, blue, green,
yellow, orange, and red. Each colour has a specific wave length and frequency
.as shown in the table.
• A solution containing copper ions is blue because it absorbs the
complementary color yellow from white light and transmits the remaining
blue light.
• When polychromatic light (white • This color is complementary to the
light), which contains the whole absorbed colors. In a similar manner,
spectrum of wavelengths in the visible opaque objects will absorb certain
region, is passed through an object, the wavelengths, leaving a residual color to
object will absorb certain of the be reflected and "seen."
wavelengths, leaving the unabsorbed As an example, a solution of
wavelengths to be transmitted. These potassium permanganate absorbs light
residual transmitted wavelengths will in the green region of the spectrum
be seen as a color. with an absorption maximum of 525
nm, and the solution is purple.
Beer’s LAW
 Lambert
beer’s Law
Qualitative Analysis
Quantitative Analysis
 Quantitative calculations
•
The fraction of radiation absorbed by a solution of an absorbing analyte
can be quantitatively related to its concentration.
Beers law — relating the amount of radiation absorbed to concentration
the amount of monochromatic radiation absorbed by a sample is
described by the' beer - bouguer – lambert law, commonly called beer ' s
law.
Ex:
Example 2:
A 7.25 x 10-5 M solution of potassium permanganate has a transmittance of 44.1%
when measured in a 2.10cm cell at a wavelength of 525 nm.
(A)calculate the absorbance of this solution.
(b) the molar absorptivity of KMnO4.

• (a)
• A= - log T
• A = - log 0.441 = 0.355
• (b) Ɛ = A/bc = 0.355 / (2.10 cm x 7.25 x 10-5 mol L-1)
Ɛ = 2.33 x 103 L mol-1 cm-1
Example
 Spectrophotometer
spectrometer or spectrophotometer is an instrument that will resolve
polychromatic radiation into different wavelengths A block diagram of
spectrometer is shown below. All spectrometers require

1- a source of continuous radiation over the wavelength of interest.

2-monochromater for selecting a narrow band of wavelengths from the source
spectrum.
3- A sample cell
4- A detector or transducer for converting radiant energy
5- A device to read out the response of the detector.
• In single-beam systems, monochromatic
light from a monochromator (only a sample
beam) enters the sample compartment and
hits the detector directly. In a double-beam
system, however, the monochromatic light
from the monochromator is split into a
sample beam (S) and reference beam (R)
before entering the sample compartment
and hitting the detector. Each of these
designs is illustrated here. The single-beam
configuration has a simpler design because
it does not need a mechanism for splitting
the beam into sample and reference beams.
• Q: what is monochromator?
Monochromatic: electromagnetic radiation of a single wavelength.
Polychromatic: electromagnetic radiation of more than one wavelength.
• What is the difference between single beam and double beam ? A single
beam spectrophotometer has only one beam of light, while a double beam
spectrophotometer has two beams of light, one passing through a
reference solution and one passing through the sample
The Electronic Transition