SPECTROSCOPY AND CHROMATOGRAPHY

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● QUESTION
1. Answer the ff in tabulate form. Refer to our reference book posted in moodle. Do not copy-paste the keypoints at
the beginning of each chapter.

Method Principle Parts and Application/Uses Advantage Disadvanta

function from other ge from
spectrometry other
spectrometr
y
Ultraviolet and The foundation of 1. The light sources – Use of UV/visible An Ultraviolet - The time
Visible spectrometry UV-Visible a deuterium lamp for spectrophotometry Visible Light required to set
Spectroscopy is the the UV region from to determine Spectrophotometer up a UV-VIS
absorption of 190 to 350 nm and a pKa values. 's (UV-Vis spectrometer
ultraviolet or visible quartz halogen or Pharmacopoeial spectrophotometer' for use is its
light by chemical tungsten lamp for the methods rely heavily s) advantage is principal
substances, which visible region from on simple analysis that it can perform drawback.
generates distinctive 350 to 900 nm. by UV/visible analyses quickly Setting up
spectra. The basis 2. The spectrophotometry and is simple to spectrometers
of spectroscopy is monochromator – to determine active use. An UV/VIS for UV-VIS is
the interaction of used to disperse the ingredients in spectrophotometer crucial. Any
light and matter. light into its formulations. aids astronomers outside light,
constituent UV/visible in their analysis of electrical noise,
wavelengths, spectrophotometry galaxies, neutron or other
 which are further is a standard stars, and other contaminants
selected by the slit. method for celestial that can affect
3. The optics – may determining the phenomena. the
be designed to split physico- spectrometer's
the light beam so that chemical properties reading must
the beam passes of drug molecules be removed
through two sample prior to formulation from the
compartments and for measuring location.
their
release from
formulations.
Infrared The foundation of IR dispersive the extent to which High Scan Speed: The lack of
spectrometry spectroscopy is the instruments, which IR In one second, substantial
idea that molecules use a spectrophotometry infrared infrared
absorb particular monochromator to can be used to spectroscopy can absorption in
frequencies that are select each elucidate gather data for the these modes
unique to their wavenumber in turn structures is limited. whole frequency results in poor
structure. All of the in The information range. In order to sensitivity to
atoms in molecules order to monitor its given in Figure 5.9 complete the scan chemical units
are continuously intensity after the is confined to the before the with modest
vibrating with radiation has passed more substance begins oscillatory
respect to one through the sample, easily recognisable to degrade, IR can dipoles during a
another at and bands in the IR be utilized to vibrational
temperatures higher Fourier transform spectra of evaluate a transition.
than absolute zero. instruments, which molecules; this is to substance that is
use an discourage the not particularly
interferometer. notion that IR is a stable.
technique used for
extensive structure
elucidation – in
pharmaceutical
analysis it is a
fingerprint
technique. The most
readily assigned
absorptions are
usually at > 1500
cm1
 . The bands < 1500
cm1 are in the
fingerprint
region of the
spectrum where the
absorption is very
complex and it is
difficult to
be confident in the
assignment of
absorptions to
particular functional
groups. Fuller
tables of the bands
in the fingerprint
region are given
elsewhere1 and the
present
treatment is focused
largely on the bands
> 1500 cm1
.
Atomic The fundamental 1.Light Source. A AAS is used in a Atomic Atomic
spectrometry tenet of atomic hollow cathode lamp number of limit tests spectroscopy's spectroscopy's
spectroscopy is that coated with the for metallic core premise is core premise is
unbound atoms in element being impurities, e.g. that unbound that unbound
their ground state analysed. magnesium and atoms can absorb atoms can
can absorb light of a 2. Flame. The flame strontium in calcium light of a particular absorb light of
specific wavelength. is usually acetate, palladium wavelength in their a particular
Each element's air/acetylene, in carbenicillin ground state. No wavelength in
absorption is providing a sodium, and lead in other elements their ground
unique; no other temperature ca bismuth absorb this state. No other
elements absorb 2500C. subgallate. It is also wavelength; each elements
this wavelength. Nitrous used to assay element absorbs it absorb this
oxide/acetylene may metals in a number exclusively. wavelength;
be used to produce of other each element
temperatures up to preparations: zinc absorbs it
3000C, in zinc insulin exclusively.
which are required to suspension and
 volatilise salts of tetracosactrin zinc
elements such as injection; copper
aluminium or and iron in ascorbic
calcium. acid; zinc in
3. Monochromator. acetylcysteine; lead
The monochromator in
is used to narrow bismuthsubcarbonat
down the width of the e; silver in
band of radiation cisplatinum;
being examined and lead in oxyprenolol;
is thus set to monitor aluminium in
the wavelength being albumin solution;
emitted by the hollow and calcium,
cathode lamp magnesium,
mercury and zinc in
water used for
diluting
haemodialysis
solutions.
Molecular 4. Detector The
emission intensity of the
spectrometry selected radiation is
then measured using
a
photosensitive cell
Nuclear Numerous nuclei The sample is placed An advantage of this NMR enables The inherent
magnetic have spin, and all in 3, 5, 10 millimetre method of users to gather insensitivity of
resonance nuclei are glass NMR tubes and quantitation is that a detailed structural the techniques
spectrometry electrically charged, is spun pure external information from is the major
according to the in the fixed magnetic standard for the intact molecules' drawback of
NMR underlying field in cases of drug is not required vibrations in their NMR
principle. Energy one-dimensional since the response natural spectroscopy
can go from the experiments at ca is purely environment. The and imaging un
base energy to a 30 revolutions/s by proportional to the process of comparison to
higher energy level means of an air number of protons gathering and other
if an external turbine, thus ensuring present and this can analyzing data is modalities. For
magnetic field is uniformity of the be measured made easier and practical
provided (generally magnetic field across against a pure faster by NMR purposes, the
a single energy the sample in a internal standard. spectrometers. NMR
gap). horizontal direction. Thus the purity Users can identify experiment's
The sample is of a substance can compounds using signal is weak
analysed be determined the known libraries and most
in solution in a without a pure of NMR closely
deuterated solvent to standard for it being spectrometers. correlated with
ensure there is no available. the nuclei
interference with the concentration in
signal from protons in the sample.
the relatively much
larger amount of
solvent.

Mass spectrometry A mass Mass spectroscopes Specific applications The MS/MS has Disadvantages
spectrometer consist of five basic of mass three major of mass spec
generates multiple parts: a high vacuum spectrometry advantages: The are that it isn't
ions from the system; a sample include drug testing capacity to study very good at
sample under handling system, and discovery, food numerous identifying
investigation, it then through which the contamination molecules hydrocarbons
separates them sample to be detection, pesticide regardless of that produce
according to their investigated can be residue analysis, whether they are similar ions and
specific introduced; an ion isotope ratio from the same it's unable to
mass-to-charge source, in which a determination, structural family or tell optical and
ratio (m/z), and then beam of charged protein not; The capacity geometrical
records the relative particles identification, and to highlight the isomers apart.
 abundance of each characteristic of the carbon dating. specific The
ion type. sample can be metabolites of a disadvantages
produced; an disease; It's an are
analyzer, in which the automated compensated
beam can be technique offering for by
separated into its the possibility of combining MS
components; and a large-scale with other
detector or receiver analysis techniques,
by means of which such as gas
the separated ion chromatograph
beams can be y (GC-MS).
observed or
collected.

Method Principle Parts Application/Uses Advantage from Disadvantage

and other from other
function spectrometry spectrometry
Gas Chromatography The GC works The Gas GC-MS can be GC-MS provides GC-MS provides
on the principle Chromatography/ used for the enhanced sample enhanced sample
that a mixture Mass bioanalysis of body identification, identification,
will separate Spectrometry fluids to detect higher sensitivity, higher sensitivity,
into individual (GC/MS) narcotics, an increased an increased
substances instrument barbiturates, range of range of
when heated. separates alcohols, and drugs analyzable analyzable
The heated chemical mixtures such as samples, and samples, and
gases are (the GC anticonvulsants, faster results, faster results,
carried through component) and anesthetics, which enable a which enable a
a column with identifies the antihistamines, whole new range whole new range
an inert gas components at a sedative hypnotics, of applications for of applications for
(such as molecular level and anti-epileptic GC-MS in several GC-MS in several
helium). As the (the MS drugs. It is also areas. areas.
separated component). It is useful in detecting
substances one of the most pollutants and
emerge from the accurate tools for metabolites in
 column opening, analyzing serum and in fatty
they flow into environmental acid profiling in
the MS. samples. microbes.
High-Performance The basic The basic The HPLC is used Speed, Efficiency The disadvantage
liquid principle of principle of HPLC in the quantitative and Accuracy of HPLC
HPLC is that it is that it separates determination of Compared to other HPLC can be a
Chromatography separates a a sample into its plasma levels of chromatographic costly strategy, it
sample into its constituent parts drugs and their techniques, such required countless
constituent parts based on the metabolites. This is as TLC, HPLC is costly organics,
based on the relative affinities of special extremely quick needs a force
relative affinities of distinct importance during and efficient. It supply and
of distinct molecules for the the development of uses a pump, ordinary support is
molecules for mobile phase and new drugs and for rather than gravity, required. It can be
the mobile the stationary monitoring therapy. to force a liquid muddled to
phase and the phase used in the The measurement solvent through a investigate issues
stationary phase separation. of concentrations of solid adsorbent or grow new
used in the certain biogenic material, with methods.
separation. substances helps in different chemical
diagnosis. components
separating out as
they move at
different speeds.
Thin-Layer The principle of Chromatography Chromatography is Precise It is a
Chromatograph TLC is the is based on the based on the separation, time-consuming
distribution of a principle where principle where analyses, and process for the
y
compound molecules in molecules in purification is separation of
between a solid mixture applied mixture applied possible using compounds.
fixed phase (the onto the surface onto the surface or chromatography. It is expensive as
thin layer) or into the solid, into the solid, and It requires very low higher quantities of
applied to a and the stationary the stationary fluid sample volumes. solvents are
glass or plastic fluid phase (stable phase (stable It works on a wide required.
plate and a phase) is phase) is range of samples The automated
liquid mobile separating from separating from including drugs, process becomes
phase (eluting each other while each other while food particles, complicated and
solvent) that is moving with the moving with the aid plastics, therefore costly.
moving over the aid of a mobile of a mobile phase. pesticides, air and It has a low
solid phase. phase. water samples, separation power.
and tissue
extracts.