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Separation, Electroanalytical and Spectrometric Techniques
Separation, Electroanalytical and Spectrometric Techniques
Spectrochemical Techniques
Separation, Electroanalytical and
Spectrochemical Techniques
Notice
Table of Contents
I. Separation, Electroanalytical, and Spectrochemical Techniques________ 3
III. Time_____________________________________________________ 3
IV. Materials__________________________________________________ 3
VI. Content___________________________________________________ 4
6.1 Resume _____________________________________________ 4
6.2 Outline_ _____________________________________________ 5
6.3 Graphic Organizer______________________________________ 7
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III. Time
• Separation and Chromatographic Techniques 25 hours
• Electrochemical Techniques 15 hours
• Spectroscopy and Atomic Spectroscopic Techniques 20 hours
• Molecular Spectroscopy 1(UV and IR) 30 hours
• Molecular Spectroscopy 2 (NMR) 15 hours
• Mass Spectrometry 15 hours
IV. Materials
You will require the following tools and resources for completing this module
V. Module Rationale
Separation, Electro analytical and Spectroscopic Techniques are the basis of instru-
mental analysis widely applied in industry, chemistry, biochemistry, environment
and school science. These techniques are based on principles of chemistry taught at
school level. Therefore in this module we shall study the principles on which these
techniques are based and acquire the basic skills necessary to use the techniques.
Studying this area deepens the understanding of the underlying chemistry principles
making the learner better able to teach them at school level.
VI. Content
6.1 Resume
This module consists of three interrelated subject areas; Separation Techniques and
Chromatographic Techniques, Electro analytical Techniques and Spectroscopic
Methods.
The module will be taught in six learning units reflecting common concepts and
approaches.
Separation Techniques and Chromatographic Techniques unit will review elemen-
tary separation techniques that are usually taught in the school system, followed by
a discussion of Chromatography Techniques; these are covered by introducing the
general chromatographic theory, followed by its application in different techniques
of plane and column chromatographic techniques.
Electro Analytical Techniques will introduce principles on which potentiometry is
based, elaborate the common applications of potentiometry, this will be followed by
voltammetry, starting with polarographic techniques ending with cyclic and anodic
stripping voltammetry.
The unit on Spectroscopy and Atomic Spectroscopic techniques will review concepts
of energy matter interaction, energy levels in atoms and molecules, and the unit will
end with a discussion of atomic spectroscopic techniques.
Molecular Spectroscopy 1 will start with a discussion of the theory of UV-Visible
spectroscopy, how it arises and how it is used in qualitative and quantitative analysis,
instrumentation of the modern UV-visible spectrophotometer. The unit will end with a
discussion of infrared spectroscopy starting with how the spectra arises, the different
peaks exhibited by specific functional groups and how to apply IR in identification
of functional groups and compounds.
Molecular Spectroscopy 2 will introduce nuclear magnetic resonance phenomenon,
followed by a discussion of proton NMR, the relationship of chemical shift with the
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6.2 Outline
• Separation Techniques
o Solvent Extraction
o Distillation
• Chromatography
o Theory of Chromatography
o The Development Process
• Types of Chromatographic Techniques
o Plane Chromatography
o Plane Chromatography
o Liquid Chromatography
o Gas Chromatography
• Potentiometry.
o Ion Selective Electrodes
o pH Glass Electrodes
o Potentiometric Titrations
• Voltammetry
o Polarography
o Pulse Polarography
o Cyclic Voltammetry
o Anodic Stripping Voltammetry
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• Spectroscopy:
o Electromagnetic Radiation
o The Atom and Atomic Spectroscopy
o Beers law
• Atomic Spectroscopic Techniques
• Mass Spectrometry
o Fragmentation Patterns
o Finger Print Spectrum
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Pre assessment
1) A beryllium atom has 4 protons, 5 neutrons, and 4 electrons. What is the mass
number of this atom?
a) 4 b) 9 c) 8 D) 7
8) The hydrogen halides are all polar molecules which form acidic solutions. Which
of the following is the weakest acid?
a) HI
b) HF
c) HBr
d) HCL
10) In all electrochemical cells, the process that takes place at the anode is _______
__________ and the process that takes place at the cathode is ____________.
a) oxidation, reduction
b) reduction, reduction
c) reduction, oxidation
d) oxidation, oxidation
13) The equation that represents a reaction that is not a redox reaction is:
a) Zn + CuSO4 → ZnSO4 + Cu
b) 2H2O2 → 2H2O + O2
c) H2O + CO2 → H2CO3
d) 2H2 + O2 → 2H2O
14) A mole of electrons has a charge of 96,485 coulombs per mole of electrons. This
quantity is known to chemists as:
a) 1 watt
b) 1 Ampere
c) 1 joule
d) 1 faraday
15) Which of the following properties of water explains its ability to dissolve acetic
acid?
a) The high surface tension of water, which is due to the formation of hydrogen
bonds between adjacent water molecules
b) The ability to serve as a buffer, absorbing the protons given off by acetic
acid.
c) The ability to form hydrogen bonds with the carbonyl and the hydroxyl groups
of acetic acid.
d) None of the above
c. -log [H +]
d. ln [H +]
e. -ln [H +]
18) How many ml of a 0.4 M HCl solution are required to bring the pH of 10 ml of
a 0.4 M NaOH solution to 7.0 (neutral pH)?
a. 4
b. 40
c. 10
d. 20
e. 2
19) An atom of which of the following elements has the greatest ability to attract
electrons?
a) silicon
b) sulphur
c) Nitrogen
d) Chlorine
Answer Key
1. b
2. a
3. b
4. c
5. a
6. c
7. d
8. b
9. d
10. a
11. a
12. c
13. c
14. d
15. c
16. c
17. b
18. c
19. d
20. d
African Virtual University 17
X. Key Concepts
of the original mixture. Sample molecules are displaced by each other and by the
more strongly sorbed compound. The result is that the eluting sample solute zones
may be sharpened. Displacement techniques have been used mainly in preparative
EPLC applications.
External standards: A separate sample containing known quantities of the same
compounds of interest. External standards are used primarily for peak identification
by comparing elution times.
Hydrophilic: It is often referred to as water loving. It adverts both to water compatible
stationary phases, and to water soluble molecules. Most columns used to separate
proteins are hydrophilic in nature and should not sorb or denature protein in the
aqueous environment.
Injector: A mechanism for accurately injecting a predetermined amount of sample
into the mobile phase stream. The injector can be a simple manual device, or a so-
phisticated auto sampler that permits automated injections of many different samples
for unattended operation
Partition coefficient (K): The amount of solute in the stationary phase relative to the
amount of solute in the mobile phase. It can also be the distribution coefficient, KD.
Retention time (tR’): The time between injection and the appearance of the peak
maximum. The adjusted retention time tR’ adjusts for the column void volume.
Retention volume (VR):The volume of mobile phase required to elute a substance
from the column. This is where Vm is the void volume, KD the distribution coefficient,
and Vs the stationary phase volume.
Stationary phase: The immobile phase involved in the chromatographic process. The
stationary phase in liquid chromatography can be a solid, a bonded or coated phase
on a solid support, or a wall coated phase. The stationary phase used often character-
izes the LC mode. For example, silica gel is used in adsorption chromatography, an
octadecylsilane bonded phase in reversed-phase chromatography, etc.
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Spectroscopy
Mass Spectrometry
Reference 1
Reference 2
Reference 3
http://www.mhhe.com/physsci/chemistry/carey/student/olc/ch13ir,html
Summary: This link is part of a large course of chemistry, for purposes of this module
however, the learner is required to read only chapter thirteen. It presents the material
covered in the module in a simple and understandable way. This reference provides
an alternative approach to the study of the module, with theoretical concepts tutorials,
and worked examples.
Justification: The worked examples in this reference provide practice in solving
problems associated with the module and reinforcing understanding concepts
African Virtual University 27
http://ull.chemistry.uakron.edu/analytical/Spectrophotometry/
Summary: This site contains summarized notes on the concepts covered in the
module
Justification: It is good to facilitate the learner remember facts and key concepts.
African Virtual University 28
http://www.nd.edu/~smithgrp/structure/workbook.html
http://www.cem.msu.edu/~reusch/VirtualText/Spectrpy/spectro.htm#contnt
http://www.chromatography-online.org/topics/gas/chromatography/detectors.
html
http://www.cis.rit.edu/htbooks/nmr/bnmr.htm
Summary: This site offers a good treatment of the 13-Carbon NMR with frequent
explanations accompanied by spectrums.
Justification: The treatment tends to go beyond the requirements of this module
therefore the learner is advised to use it only for reference.
African Virtual University 32
http://weather.nmsu.edu/Teaching_Material/soil698/Student_Reports/
Spectroscopy/report.htm
http://www.chemguide.co.uk/index.html#top
Summary: The site was originally intended to meet the needs of UK A level chemis-
try students, but I have since been widening it to cover material on all the UK-based
syllabuses including A level, IB, Scottish Advanced Higher and Cambridge Interna-
tional. In fact it is now being used by people on equivalent (16 to 18 year old) courses
worldwide and by students at the beginning of university level courses.
Justification: This site contains very simplified explanations of the material covered
in this module Justification
African Virtual University 34
http://www.colby.edu/chemistry/NMR/H1pred.html
http://www.shsu.edu/~chemistry/primers/primers.html
Summary: This site contains a large collection of multi media resources across all
themes covered in this module.
Justification: This site should be used a source of multimedia resources
African Virtual University 36
Unit I
Separation and Chromatographic Techniques
Required Readings
Separation
http://en.wikipedia.org/wiki/Separation_process
http://en.wikipedia.org/wiki/Distillation#Laboratory_scale_distillation
http://en.wikipedia.org/wiki/Liquid-liquid_extraction
http://en.wikipedia.org/wiki/Separating_funnel
http://en.wikipedia.org/wiki/Distillation
Chromatography
http://en.wikipedia.org/wiki/HPLC
http://en.wikipedia.org/wiki/Chromatography
http://hplc.chem.shu.edu/HPLC/index.html
http://www.waters.com/watersdivision/ContentD.asp?watersit=JDRS-5LTGBH
http://www.chemguide.co.uk/analysis/chromatography/hplc.html
http://www.chemguide.co.uk/analysis/chromatography/column.html#top
http://www.chromatography-online.org/Principles/Introduction/rs1.html
http://antoine.frostburg.edu/chem/senese/101/matter/chromatography.shtml
African Virtual University 37
http://www.chemguide.co.uk/analysis/chromatogrmenu.html#top
http://www.chemguide.co.uk/analysis/chromatography/thinlayer.html#top
http://www.rpi.edu/dept/chem-eng/Biotech-Environ/CHROMO/be_types.htm
http://www.forumsci.co.il/HPLC/modes/modes3.htm
http://www.chem.ubc.ca/courseware/121/tutorials/exp3A/columnchrom/
http://teaching.shu.ac.uk/hwb/chemistry/tutorials/chrom/gaschrm2.htm
Separation Techniques
Solvent Extraction
Liquid- liquid extraction also known as solvent extraction and partitioning, is a method
to separate compounds based on their relative solubility in two different immiscible
liquids, usually water and an organic solvent. It is an extraction of a substance from one
liquid phase into another liquid phase. In this technique a solution ( usually aqueous)
containing a solute or solutes is brought into contact with a second solvent ( usually
organic) with the aim of transferring one or more of the solutes from the solution to
the to the second solvent. The solution is vigorously shaken to make intimate contact
with the solvent. The apparatus is allowed to stand to allow phases to separate.
Partition Coefficient
Partition or distribution coefficient (KD) is the ratio of concentrations of a compound
in the two phases of a mixture of two immiscible solvents at equilibrium hence these
coefficients are a measure of differential solubility of the compound between these
two solvents.
Normally one of the solvents chosen is water while the second is hydrophobic such
as octanol. Hence both the partition and distribution coefficient are measures of how
hydrophilic (“water loving”) or hydrophobic (“water hating”) a chemical substance
is.
Factors Affecting Solvent Extraction
Polarity of the solute and the polarity of the solvent: in general polar solvent will be
distributed into the more polar solvent and non polar solutes will be more soluble in
the organic solvents unless they incorporate sufficient number of hydrophilic func-
tional groups like hydroxyl, and sulphonic
Generally ionic compounds would not be expected to extract into organic compounds,
these can be extracted by reacting them with complexing agents to form large neutral
non polar entities.
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Distillation
Laboratory scale distillations are almost exclusively run as batch distillations. The
device used in distillation, sometimes referred to as a still, consists at a minimum of a
reboiler or pot in which the source material is heated, a condenser in which the heated
vapour is cooled back to the liquid state, and a receiver in which the concentrated or
purified liquid, called the distillate, is collected. Several laboratory scale techniques
for distillation exist
Simple Distillation
In simple distillation, all the hot vapours produced are immediately channelled into
a condenser which cools and condenses the vapours. Thus, the distillate will not be
pure - its composition will be identical to the composition of the vapours at the given
temperature and pressure, and can be computed from Raoult’s law.
Simple distillation therefore usually used only to separate liquids whose boiling points
differ greatly (rule of thumb is 25 °C) or to separate liquids from non volatile solids or
oils. For these cases, the vapour pressures of the components are usually sufficiently
different that Raoult’s law may be neglected due to the insignificant contribution of
the less volatile component. In this case, the distillate may be sufficiently pure for
its intended purpose.
Fractional Distillation
For many cases, the boiling points of the components in the mixture will be too close.
In this case fractional distillation is used in order to separate the components well by
repeated vaporization-condensation cycles within a packed fractionating column.
http://en.wikipedia.org/wiki/Fractional_distillation
More theoretical plates lead to better separations. A spinning band distillation system
uses a spinning band of Teflon or metal to force the rising vapours into close contact
with the descending condensate, increasing the number of theoretical plates.
Steam Distillation
Vacuum Distillation
Chromatography
Theory of Chromatography
Chromatography is a separation process that is achieved by distributing the compo-
nents of a mixture between two phases, a stationary phase and a mobile phase.
Those components held preferentially in the stationary phase are retained longer
in the system than those that are distributed selectively in the mobile phase. As a
consequence, solutes are eluted from the system at different times in order of their
increasing distribution coefficients with respect to the stationary phase; in this way
a separation is achieved
Samples may be gaseous, liquid or solid in nature and can range in complexity
from a simple blend of two enantiomers to a multi component mixture containing
widely differing chemical species. Furthermore, the analysis can be carried out, at
one extreme, on a very costly and complex instrument, and at the other, on a simple,
inexpensive thin layer plate. Chromatography is the basis of a large number of ana-
lytical techniques. This unit presents the most common chromatographic techniques
and their applications.
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A net amount of solute must now leave the stationary phase and enters the mobile
phase to re-establish equilibrium. Thus, the solute moves through the chromato-
graphic system as a result of solute entering the mobile phase at the rear of the peak
and returning to the stationary phase at the front of the peak. However, the solute is
always transferring between the two phases over the whole of the peak in an attempt
to attain or maintain thermodynamic equilibrium. Nevertheless, the solute band
progresses through the system as a result of a net transfer of solute from the mobile
phase to the stationary phase in the front half of the peak. This net transfer of solute
is compensated by solute passing from the stationary phase to the mobile phase at
the rear half of the peak.
A term called the retention factor, k’, is often used to describe the migration rate of
an analyte on a column. You may also find it called the capacity factor. The retention
factor for analyte A is defined as;
k’A = t R - tM / tM
tR and tM are easily obtained from a chromatogram. When an analytes retention factor
is less than one, elution is so fast that accurate determination of the retention time is
very difficult. High retention factors (greater than 20) mean that elution takes a very
long time. Ideally, the retention factor for an analyte is between one and five.
We define a quantity called the selectivity factor, α, which describes the separation
of two species (A and B) on the column;
α, = k ‘B / k ‘A
When calculating the selectivity factor, species A elutes faster than species B. The
selectivity factor is always greater than one.
Band Broadening and Column Efficiency
To obtain optimal separations, sharp, symmetrical chromatographic peaks must be
obtained. This means that band broadening must be limited. It is also beneficial to
measure the efficiency of the column.
The Theoretical Plate Model of Chromatography
The plate model supposes that the chromatographic column is contains a large num-
ber of separate layers, called theoretical plates. Separate equilibrations of the sample
between the stationary and mobile phase occur on these “plates”. The analyte through
the column by transfer of equilibrated mobile phase from one plate to the next.
Plates are a theoretical concept for measuring column efficiency, either by stating
the number of theoretical plates in a column, N (the more plates the better), or by
stating the plate height; the Height Equivalent to a Theoretical Plate (the smaller the
better).
If the length of the column is L, then the HETP is
HETP = L / N
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The number of theoretical plates that a real column possesses can be found by exa-
mining a chromatographic peak after elution;
Plane Chromatography
Planar chromatography are separation techniques in which the stationary phase is
present as or on a plane. The plane can be a paper, serving stationery phase (paper
chromatography) or a layer of solid particles spread on a support such as a glass plate
(thin layer chromatography).
Paper Chromatography
Paper chromatography is a technique that involves placing a small dot of sample
solution onto a strip of chromatography paper. The paper is placed in a jar contai-
ning a shallow layer of solvent and sealed. As the solvent rises through the paper it
meets the sample mixture which starts to travel up the paper with the solvent. Dif-
ferent compounds in the sample mixture travel different distances according to how
strongly they interact with the paper. This allows the calculation of an Rf value and
can be compared to standard compounds to aid in the identification of an unknown
substance.
Thin Layer Chromatography
Thin layer chromatography (TLC) is similar to paper chromatography. However,
instead of using a stationary phase of paper, it involves a stationary phase of a thin
layer of adsorbent such as silica gel, alumina, or cellulose on a flat, inert substrate
usually glass plates or plastic material. Compared to paper, it has the advantage of
faster runs, better separations, and the choice between different adsorbents. Different
compounds in the sample mixture travel different distances according to how strongly
African Virtual University 47
they interact with the adsorbent. This allows the calculation of an Rf value which
can be compared to standard compounds to aid in the identification of an unknown
substance.
Coloured substances can be seen directly when viewed against the stationary phase
while colourless species may usually be detected by spraying the plate with an ap-
propriate reagent which produces coloured areas in the regions which they occupy.
Some compounds fluoresce in ultraviolet light and may be located in this way. Al-
ternatively if fluorescing material is incorporated in the adsorbent the solute can be
observed as a dark spot on a fluorescent background when viewed under ultraviolet
light, (When locating zones by this method the eyes should be protected by wearing
special protective goggles or spectacles.) The spots located by this method can be
delineated by marking with a needle.
Column Chromatography
Column chromatography is a separation technique in which the stationary bed is
placed within a tube.
for the mobile phase in the middle part of the tube (open tubular column).
Liquid Chromatography
trument
Column - a stainless steel tube packed with silicon beads that separates what I’m
looking for (the caffeine) from other compounds (like sugar)
Detector - An optical sensor (usually) that detects changes in the characteristics of
the solvent stream
Data System - A means of controlling the system components and storing, processing
and displaying data
A high pressure pump is required to force the mobile phase through the column at
typical flow rates of 0.1-2 ml/min. The sample to be separated is introduced into the
mobile phase by injection device, manual or automatic, prior to the column.
Chromatography Scale:
HPLC can be operated for a number of purposes
Analytical - Just Data High Sensitivity
Semi-Preparative - Data and a small amount of purified analyte (gram)
Preparative - Larger quantities of purified analytes (Kilograms) [High Capacity]
Modes of HPLC Separation
Separation of analytes is based on a number of mechanisms and each of these me-
chanisms result in a different mode of application of HPLC
Normal phase chromatography
Normal phase HPLC separates analytes based on polarity. This method uses a polar
stationary phase and a nonpolar mobile phase, and is used when the analyte of interest
is fairly polar in nature. The polar analyte associates with and is retained by the polar
stationary phase. Adsorption strengths increase with increase in analyte polarity, and
the interaction between the polar analyte and the polar stationary phase (relative to
the mobile phase) increases the elution time.
Reversed phase chromatography
Reversed phase HPLC (RP-HPLC) consists of a non-polar stationary phase and an
aqueous, moderately polar mobile phase. One common stationary phase is a silica
which has been treated with RMe2SiCl, where R is a straight chain alkyl group such
as C18H37 or C8H17. The retention time is therefore longer for molecules which are
more non-polar in nature, allowing polar molecules to elute more readily. Retention
time is increased by the addition of polar solvent to the mobile phase and decreased
by the addition of more hydrophobic solvent.
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Gas Chromatography
is where a micro syringe is used to inject sample through a rubber septum into a
flash vaporizer port at the head of the column. The temperature of the sample port
is usually about 50oC higher than the boiling point of the least volatile component
of the sample. For packed columns, sample size ranges from tenths of a microliter
up to 20 micro-liters. Capillary columns on the other hand, need much less sample,
typically around 10-3 µL.
Columns
There are two general types of column, packed and capillary (also known as open
tubular). Packed columns contain a finely divided, inert, solid support material (com-
monly based on diatomaceous earth) coated with liquid stationary phase. Most packed
columns are 1.5 - 10m in length and have an internal diameter of 2 - 4mm.
Capillary columns have an internal diameter of a few tenths of a millimeter. They
can be one of two types; wall-coated open tubular (WCOT) or support-coated open
tubular (SCOT). Wall-coated columns consist of a capillary tube whose walls are
coated with liquid stationary phase. In support-coated columns, the inner wall of the
capillary is lined with a thin layer of support material such as diatomaceous earth,
onto which the stationary phase has been adsorbed. SCOT columns are generally
less efficient than WCOT columns. Both types of capillary column are more efficient
than packed columns.
Column Oven
To get reproducible results, column temperature must be controlled to within tenths
of a degree. The optimum column temperature is dependant upon the boiling point
of the sample. To maintain a reproducible temperature, the chromatography column
is maintained in an oven that can be set at different temperatures.
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Detectors
There are many detectors which are used in gas chromatography. Different detectors
will give different types of selectivity. A non-selective detector responds to all com-
pounds except the carrier gas. A selective detector responds to a range of compounds
with a common physical or chemical property and a specific detector responds to a
single chemical compound. Detectors can also be grouped into concentration depen-
dant detectors and mass flow dependant detectors. The signal from a concentration
dependant detector is related to the concentration of solute in the detector, and does
not usually destroy the sample. Dilution of with make-up gas will lower the detectors
response.
Mass flow dependant detectors usually destroy the sample, and the signal is related
to the rate at which solute molecules enter the detector. The response of a mass flow
dependant detector is unaffected by make-up gas.
Support Dynamic
Detector Type Selectivity Detectability
gases range
Flame ion- Hydrogen
Mass flow Most organic cpds. 100 pg 107
ization (FID) and air
Halides, nitrates,
Electron
Concentra- nitriles, perox-
capture Make-up 50 fg 105
tion ides, anhydrides,
(ECD)
organometallics
Sulphur, phospho-
Hydrogen
Flame rus, tin, boron,
and air
photometric Mass flow arsenic, germa- 100 pg 103
possibly
(FPD) nium, selenium,
oxygen
chromium
Aliphatics, aro-
matics, ketones,
esters, aldehydes,
Photo-ion- Concentra-
Make-up amines, hetero- 2 pg 107
ization (PID) tion
cyclics, organo-
sulphurs, some
organometallics
Formative Assessment
1.
i) Name major components of gas chromatographs
ii) Name two types of GC columns
iii) State the common carrier gases used in GC, Name one chemical property they
must all posses
iv) Name one type of samples that are not analysed by GC and explain why.
v) Clearly explain what a selective detector is
2. For a typical chromatographic separation giving just-resolved peaks (Rs = 1.5),
assume that N = 3600, k’ = 2, and α = 1.15. Sketch the effects of changing these
parameters one at a time to (a) N = 1600, (b) k' = 0.8, and (c) a = 1.10.
3. To decrease the plate height and yet increase the resolution, what courses of
action are available? What penalties may accrue for each approach?
4. The relative response factors for p-dichlorobenzene and p-xylene (relative to
the value for benzene, assigned unity) were found to be 0.624 ± 0.034 and 0.917
± 0.018, respectively. Upon integration of chromatographic peaks the results in
the table were obtained. Calculate the percent composition of each sample.
Peak Area
Sample Benzene p-xylene Toluene
African Virtual University 56
Unit II
Electroanalytical Techniques
http://en.wikipedia.org/wiki/Electroanalytical_methods
http://www.chem.vt.edu/chem-ed/echem/electroc.html
http://www.chem.vt.edu/chem-ed/echem/potentio.html
http://electrochem.cwru.edu/ed/encycl/art-a03-analytical.htm
http://ull.chemistry.uakron.edu/analytical/Voltammetry/
http://ull.chemistry.uakron.edu/analytical/index.html
Potentiometry
In potentiometry, the measuring setup always consists of two electrodes: the measu-
ring electrode, also known as the indicator electrode, and the reference electrode.
Both electrodes are half-cells. When placed in a solution together they produce a
certain potential.
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The potential as reflected above measures the activity of the ions rather than the
concentration. Where a=activity, γ=activity coefficient c= concentration. The acti-
vity of the measured ion a, which is also used in the Nernst equation, is linked to the
normally interesting analytical concentration c via the activity coefficient*:
a=γC
Where Eo is the potential difference of the two electrodes at unit activity and stan-
dard conditions
For dilute solutions with concentration cM 0.001mol/L, the activity coefficient
tends towards 1 and the activity of the ion corresponds to its concentration as a first
approximation. γ is a function of the total electrolyte content. The mathematical
relationship between the activity aM of a measuring ion in solution ions and the
potential measured between the reference electrode and the measuring electrode is
described by the Nernst equation.
2.303 xRXT a
E = Εo + Log oxid
ZXF a re
Where is the difference between the standard potential of the electrode and the
potential of the standard electrode. The indicator electrode usually contains one of
the forms of the desired ions or enabling it to measure the other form that is in the
solution.
By measuring E, it is therefore possible to measure the concentration of analyte is
known.
Ph Glass Electrodes
The glass membrane of a pH glass electrode consists of a silicate framework containing
lithium ions. When a glass surface is immersed in an aqueous solution then a thin
solvated layer (gel layer) is formed on the glass surface in which the glass structure is
softer. This applies to both the outside and inside of the glass membrane. As the proton
concentration in the inner buffer of the electrode is constant (pH = 7), a stationary
condition is established on the inner surface of the glass membrane. In contrast, if
the proton concentration in the measuring solution changes then ion exchange will
occur in the outer solvated layer and cause an alteration in the potential at the glass
membrane. Only when this ion exchange has achieved a stable condition, will the
potential of the glass electrode also be constant. This means that the response time of
a glass electrode always depends on the thickness of the solvated layer. Continuous
contact with aqueous solutions causes the thickness of the solvated layer to increase
continuously – even if only very slowly which results in longer response times. This
is why conditioning the electrode in a suitable electrolyte is absolutely necessary
to ensure an initial solvated layer condition that is as stationary as possible so that
results can be obtained that are as reproducible as possible.
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Potentiometric Titrations
In potentiometry titrations, ion sensitive electrode is used to monitor potential changes
in the titration vessel.
A cell as described before is made composed of an indicator electrode and a reference
electrode. As the titrant is added to the reaction vessel it reacts with the analyte and
consumes it. At equivalence point there is a large change in potential indicating total
consumption of the analyte.
A plot of the volume of the titrant with potential shows a large inflexion at the end
point or a plot of E vs. Volume.
Titrant Volume V
dE/dV
Voltammetry
Voltammetry refers to the measurement of current that result from the application of
potential. Unlike potentiometry measurements, which employ only two electrodes,
voltammetric measurements utilize a three electrode electrochemical cell. The use
of the three electrodes (working or micro, auxiliary, and reference) along with the
potentiostat instrument allows accurate application of potential functions and the
measurement of the resultant current.
The micro electrode is usually polarized i.e. the concentration of the ions at the surface
electrode is different from the concentration of the ions from the bulk of the solution.
Therefore the diffusion of the ions from the bulk of the solution to the micro electrode
becomes an important phenomenon. The total current I= Im+ Id
Im=Migration current
Id=diffusion current
The different voltammetric techniques that are used are distinguished from each other
primarily by the potential function that is applied to the working electrode to drive
the reaction, and by the material used as the working electrode. Common techniques
to be discussed in this module include;
• Polarography
• Normal-pulse polarography (NPP)
• Differential-pulse polarography (DPP)
• Cyclic voltammetry
• Anodic-stripping voltammetry
Polarography
In Polarography the micro electrode is a succession of mercury drops (falling slowly
from a capillary tube) and is usually the cathode. The anode (or counter Electrode) is
usually a pool of mercury. The electrolyte is the solution of the analyte which must
be electroactive material to which is added and excess of a supporting electrolyte
usually KCl.
This type of micro electrode is called a dropping mercury electrode (DME)
If a voltage is imposed on the DME an It current will flow that is composed of the
following
It=Id +Im +Ir
Residual current Ir
A small current will flow due to the capacitive charging of the mercury drops and
reducible impurities in the supporting electrolyte.
Migration Current Im
The electroactive material reaches the DME by two mechanisms; by migration and
by diffusion. If the concentration of the supporting electrolyte is high, more than
100 times the analyte, then all the migration current will be carried by the support-
ing electrolyte.
Id = 708nD1/2m2/3t1/6c
With the excess of the supporting electrolyte the electro active material will reach
the DME by diffusion. As the voltage at the DME is increased this diffusion current
increases until it reaches a limiting value Id
From theory
D is a constant from diffusion theory, n is the number of electrons involved in the
electrochemical reaction, m is the mass of mercury drops and t is the interval between
mercury drops
It can clearly be seen that the diffusion current is proportional to the concentration
of the electroactive analyte.
Id
Current
E 1/2 Voltage
African Virtual University 62
Pulse Polarography
Figure 14: The Applied Potential Wave Form for Normal pulse Polaraography
For this experiment, the polarogram is obtained by plotting the measured current vs.
the potential to which the step occurs. As a result, the current is not followed during
Hg drop growth, and normal pulse polarogram has the typical shape of a sigmoid. By
using discrete potential steps at the end of the drop lifetime (usually during the last
50-100 ms of the drop life which is typically 2-4 s), the experiment has a constant
potential applied to an electrode with nearly constant surface area. After the initial
potential step, the capacitive current decays exponentially while the faradaic current
decays as the square root of time. The diffusion current is measured just before the
drop is dislodged, allowing excellent discrimination against the background capacitive
current. The normal pulse polarography method increases the analytical sensitivity
by 1 - 3 orders of magnitude (limits of detection 10-7 to 10-8 M, relative to normal
dc polarography.
African Virtual University 64
Figure 15: The Applied Potential Wave Form for Differential Pulse Polarography
In this manner, the total waveform applied to the DME is very much like a combination
of a linear ramp with a superimposed square wave. The differential pulse polarogram
is obtained by measuring the current immediately before the potential step, and then
again just before the end of the drop lifetime. The analytical current in this case is
the difference between the current at the end of the step and the current before the
step (the differential current). This differential current is then plotted vs. the average
African Virtual University 65
potential (average of the potential before the step and the step potential) to obtain the
differential pulse polarogram. Because this is a differential current, the polarogram
in many respects is like the differential of the sigmoidal normal pulse polarogram.
As a result, the differential pulse polarogram is peak shaped.
Differential pulse polarography has even better ability to discriminate against ca-
pacitive current because it measures a difference current (helping to subtract any
residual capacitive current that remains prior to each step). Limits of detection with
Differential Pulse Polarography are
10-8 - 10-9 M.
Cyclic Voltammetry
Cyclic voltammetry (CV) is an electrolytic method that uses microelectrodes and
an unstirred solution so that the measured current is limited by analyte diffusion at
the electrode surface. The electrode potential is ramped linearly to a more negative
potential, and then ramped in reverse back to the starting voltage. The forward scan
produces a current peak for any analytes that can be reduced through the range of the
potential scan. The current will increase as the potential reaches the reduction potential
of the analyte, but then falls off as the concentration of the analyte is depleted close
to the electrode surface. As the applied potential is reversed, it will reach a potential
that will reoxidize the product formed in the first reduction reaction, and produce a
current of reverse polarity from the forward scan. This oxidation peak will usually
have a similar shape to the reduction peak. The peak current, ip, is described by the
Randles-Sevcik equation:
ip = (2.69x105) n3/2 A C D1/2 v1/2
Where n is the number of moles of electrons transferred in the reaction, A is the area
of the electrode, C is the analyte concentration (in moles/cm3), D is the diffusion
coefficient, and v is the scan rate of the applied potential.
The potential difference between the reduction and oxidation peaks is theoretically
59 mV for a reversible reaction. In practice, the difference is typically 70-100 mV.
Larger differences, or nonsymmetric reduction and oxidation peaks are an indica-
tion of a nonreversible reaction. These parameters of cyclic voltammograms make
CV most suitable for characterization and mechanistic studies of redox reactions at
electrodes.
Anodic Stripping Voltammetry
Anodic stripping voltammetry is an electrolytic method in which a mercury electrode
is held at a negative potential to reduce metal ions in solution and form an amalgam
with the electrode. The solution is stirred to carry as much of the analyte metal(s)
to the electrode as possible for concentration into the amalgam. After reducing and
accumulating the analyte for some period of time, the potential on the electrode is
increased to reoxidize the analyte and generate a current signal. The ramped potential
usually uses a step function, such as in normal-pulse polarography (NPP) or diffe-
rential-pulse polarography (DPP).
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where il is the limiting current during reduction of the metal, td is the duration of
accumulation, n is the number of moles of electrons transferred in the half reaction,
F is the Faraday constant (96,487 coulombs/mole of e-), and VHg is the volume of
the electrode. The expression for current produced by anodic stripping depends on
the particular type of Hg electrode, but is directly proportional to the concentration
of analyte concentrated into the electrode. The main advantage of stripping analysis
is the preconcentration of the analyte into the electrode before making the actual
current measurement. Anodic stripping can achieve detection of concentrations as
low as 10-10 M.
African Virtual University 67
Unit III
Spectroscopy And Atomic Spectroscopic Techniques
• http://en.wikipedia.org/wiki/Atomic_Orbital
• http://en.wikipedia.org/wiki/Energy_level
• http://en.wikipedia.org/wiki/Atomic_absorption_spectroscopy
• http://ull.chemistry.uakron.edu/analytical/Atomic_spec/
• http://www.chem.vt.edu/chem-ed/spec/atomic/aa.html
Spectroscopy
Spectroscopy is the study of the interaction between wave radiation or light, as well
as particle radiation and matter. Spectrometry is the measurement of these interac-
tions and an instrument which performs such measurements is a spectrometer or
spectrograph. A plot of the interaction is referred to as a spectrum.
Spectroscopy is often used in physical and analytical chemistry for the identification
and quantification of substances through the spectrum emitted from or absorbed by
them. This unit introduces in a simple way the concept of radiation matter interac-
African Virtual University 68
tions, the common terminology used in spectroscopy and Beer’s law which is widely
applied in the quantitative spectroscopy. The unit further discusses commonly used
atomic spectroscopic techniques.
Electromagnetic Radiation
position, and radiant energy equivalent to the amount of energy initially absorbed
in the excitation process will be emitted. The process is illustrated in Figure 18.
Note that in Step I of the process, the excitation is forced by supplying energy.
The decay process in Step 2 involving the emission of light occurs spontaneously.
Figure 18: Adapted from Perkin Elmer Corporation: Excitation and Emission
The wavelength of the emitted radiant energy is directly related to the energy of
electronic transition which has occurred. Since every element has a unique electro-
nic structure, the wavelength of light emitted is a unique property of each individual
element. As the orbital configuration of a large atom may be complex, there are many
electronic transitions which can occur, each transition resulting in the emission of a
characteristic wavelength of tight, as illustrated in Figure 2, E= hν
The process of excitation and decay to the ground state is involved in the three fields
of atomic spectroscopy; either the energy absorbed in the excitation process or the
energy emitted in the decay process is measured and used for analytical purposes.
Molecules and Molecular Spectroscopy
For a given excitation process the molecule or the atom absorbs only one discrete
amount of energy. This should correspond to one frequency being absorbed. Howe-
ver in practice a group of molecules exists in a number of energy states each state
differing from the other by a small amount of energy. Thus a group of molecules in
a sample gives rise to absorption over a small range of energy giving rise to a small
band or a peak.
Formative Question
Explain why absorption spectra for atomic species consist of discrete lines at specific
wavelengths rather than broad bands as for molecular species.
African Virtual University 72
Beer’s Law
When radiation passes through a region containing atoms or molecules the radiation
will be absorbed. The diagram below shows a beam of monochromatic radiation of
radiant power P0, directed at a sample solution. Absorption takes place and the beam
of radiation leaving the sample has radiant power P.
Po
P
The amount of radiation absorbed is dependent on the nature of the sample, the
concentration of the sample and the length of the sample.
The amount of radiation absorbed may be measured in by a number of parameters:
Transmittance, T = P / P0% Transmittance, %T = 100 T
Absorbance,
A = log10 P0 / P A = log10 1 / T A = log10 100 / %TA = 2 - log10 %T
The last equation, A = 2 - log10 %T, is worth remembering because it allows you to
easily calculate absorbance from percentage transmittance data.
The relationship between absorbance and transmittance is illustrated in the following
diagram:
So, if all the light passes through a solution without any absorption, then absorbance
is zero, and percent transmittance is 100%. If all the light is absorbed, then percent
transmittance is zero, and absorption is infinite.
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Electrons exist in energy levels within an atom. These levels have well defined en-
ergies and electrons moving between absorb or emit energy equal to the difference
between them.
In atomic spectroscopy, the energy absorbed to move an electron to a more energetic
level and/or the energy emitted as the electron moves to a less energetic energy level
is in the form of a photon (a particle of light). Because this energy is well-defined,
an atom’s identity (i.e. what element it is) can be identified by the energy of this
transition. The wavelength of light can be related to its energy. It is usually easier to
measure the wavelength of light than to directly measure its energy.
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Atomic spectroscopy can be further divided into absorption, emission, and fluores-
cence.
In atomic absorption spectroscopy, light is passed through a collection of atoms.
If the wavelength of the light has energy corresponding to the energy difference
between two energy levels in the atoms, a portion of the light will be absorbed.
The relationship between the concentrations of atoms, the distance the light travels
through the collection of atoms, and the portion of the light absorbed is given by the
Beer-Lambert law.
The energy stored in the atoms can be released in a variety of ways. When it is released
as light, this is known as fluorescence. Atomic fluorescence spectroscopy measures this
emitted light. Fluorescence is generally measured at a 90° angle from the excitation
source to minimize collection of scattered light from the excitation source, often such
a rotation is provided by a Pellin-Broca prism on a turntable which will also separate
the light into its spectrum for closer analysis. The wavelength once again tells you
the identity of the atoms. For low absorbances (and therefore low concentrations) the
intensity of the fluoresced light is directly proportional to the concentration of atoms.
Atomic fluorescence is generally more sensitive (it can detect lower concentrations)
than atomic absorption.
Atomic Emission
In atomic emission, a sample is subjected to a high energy thermal environment in
order to produce excited state atoms, capable of emitting light. The energy source
can be an electrical arc, a flame, or more recently plasma. The emission spectrum of
an element exposed to such an energy source consists of a collection of the allowa-
ble emission wavelengths, commonly called emission lines, because of the discrete
nature of the emitted wavelengths- This emission spectrum can be used as a unique
characteristic for qualitative identification of the element. Atomic emission using
electrical arcs has been widely used in qualitative analysis.
Emission techniques can also be used to determine how much of an element is pre-
sent in a sample. For a “quantitative” analysis, the intensity of light emitted at the
wavelength of the element to be determined is measured. The emission intensity
at this wavelength will be greater as the number of atoms of the analyte element
increases- The technique of flame photometry is an application of atomic emission
for quantitative analysis.
Atomic Absorption
Formative Assessment
Unit IV
Molecular Spectrocopy 1: Uv-visible And Ir
http://en.wikipedia.org/wiki/Molecular_energy_state
http://www.scienceofspectroscopy.info/edit/index.php?title=UV_Absorption_Ta-
ble
http://teaching.shu.ac.uk/hwb/chemistry/tutorials/molspec/uvvisab4.htm
http://www.scienceofspectroscopy.info/edit/index.php?title=UV-Visible_Spec-
troscopy
http://ull.chemistry.uakron.edu/analytical/Spectrophotometry/
http://www.cem.msu.edu/~parrill/AIRS/name_list.html
African Virtual University 80
Electronic Transitions
The absorption of light energy by organic compounds in the visible and ultraviolet
region involves promotion of electrons in δ, π, and n orbitals from the ground state to
higher-energy states. These higher energy states are called anti bonding orbitals. The
anti bonding orbital associated with the a bond is called the δ* (sigma star) orbital
and that associated with the π bond is called the π* (pi star)
Many molecules contain atoms with valence electrons that are not directly involved
in bonding; these are called nonbonding or n electrons and are mainly located in
atomic orbitals of oxygen. sulphur, nitrogen, and the halogen.
The n electrons do not form bonds, therefore, there are no anti bonding orbitals as-
sociated with them. The presence of an electron in an anti bonding orbital indicates
that the molecule is in a high-energy state. The electron density between the atomic
nuclei is less than that at the same distance from the nucleus in an isolated atom. In
the excited state some, but not all, of the electrons in a molecule occupy antibonding
orbitals.
The electronic transitions (→) that are involved in the ultraviolet and visible regions
are of the following types: δ→ δ*, n —> δ*, n →• π *, and •n —> π*. The energy
required for the δ —> δ* transition is very high; consequently, compounds in which
all valence shell electrons are involved in single-bond formation, such as saturated
hydrocarbons, do not show absorption in the ordinary ultraviolet region.
The light comes from the source and a thin ray is allowed in by slit 1, the diffraction
grating selects the required wave length and a thin ray is again selected by slit 2 the
filter allows in only selected wavelength mirror 2 rotates the through the half mirror
where part of the beam is reflected to mirror 3 and part is transmitted to mirror4, to
form reference beam and sample beam.
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Formative Exercise.
This exercise reinforces the knowledge covered in the above section identification of
Chromophores using UV
i) Which of the following compounds will absorb in the UV region?
a). CO2 b). H2 c) CH3CO
ii) Arrange the following compounds in order of the increasing frequency or wave
number at which they will absorb
a). CH3CO b). CH2=CH-CH=CH2 c). CH2=CH-CH=CH-CH=CH2
iii) Identify possible electronic transitions that will lead to UV absorption in the
following compounds
a). CH3CO b).CH2=CH-CH=CH2 c).CH3CN
Infrared Spectroscopy
IR absorption of C=O
Ketones
If a compound contains a carbonyl group, the absorption caused by C==0 stretching
is generally among the strongest present.
Carbonyl groups of ketones generally absorb in the region 5.7-6.0 µm (1754-1667
cm-1); the position of absorption is sensitive to ring size and to the degree of conju-
gated unsaturation, among other factors.
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Aldehydes
The absorption owing to the carbonyl stretching vibration of Aldehydes appears in
the same general region as that of ketones. The other striking characteristic of the
aldehyde functional group absorption is the presence of two weak bands owing to
C—H stretching vibrations. The wavelength of this absorption is increased (the wave
number is lowered) from the normal C—H stretching position near 3.4 µm (-294.0
cm-1) to about 3.55 and 3.68 µm.
(-2820 and 2720 cm-1). The presence of two absorption bands in this region is due
to the symmetric and asymmetric stretching modes of the C—H bond and C=0
bonds.
Esters and lactones
The position of absorption of the carbonyl stretching vibration of esters and lactones
is dependent, as with ketones, on conjugated unsaturation and ring size. But it is in
the general area of the C=O abs
Carboxylic Acid
The absorption owing to the carbonyl stretching vibration of a saturated carboxylic
acid (5.83 µm., 1715 cm-1) is shifted to longer wavelength (lower wave number) if
conjugated with an unsaturated group (benzoic acid, 5.88 µm 1701 cm-1).
Amides
All amides show strong absorption owing to carbonyl stretching and Absorptions
resulting from N—H stretching vibrations of primary and secondary amides are in
the 2.8-3.2 μm, (~3570-3125 cm-1)
Amines
The most characteristic absorption of amines is that owing to N—H stretching vi-
brations in the region 2.8-3.0 µm, (~3570-3333 cm-1). In dilute solution in an inert
solvent, the spectra of primary amines have two sharp bands in this region, owing
to symmetric and asymmetric N—H stretching vibrations; the spectra of secondary
amines have only one band in this region, and tertiary amines do not absorb in this
region.
lene bond is conjugated with some unsaturated group. Absorption owing to olefinic
C=C—H stretching vibration is observed as a small peak at 3.19 µm (3135 cm-1)
near the larger alkane C—H stretching vibration absorption
Triple bonds
Absorptions resulting from carbon carbon triple bond stretching vibrations of acety-
lenic compounds occur in the region 4.4-4.8 µm (~2275-2085 cm-1). The absorption
is weak, especially if the acetylenic linkage is non terminal. The stretching vibration
results only in a linear expansion and contraction of the molecule, and hence the dipole
moment is not much affected. Absorption caused by the acetylenic C—H stretching
vibration occurs as a fairly strong, sharp band near 3.0 µm (~3333 cm-1).
The absorption caused by the stretching vibration of the triple bond of nitriles occurs
in about the same region as that of acetylenes, but the absorption is much more intense.
This absorption of benzonitrile appears at 4.44 µm (2252 cm-1).
Aromatic compounds
There are four absorption bands in the 6-7 am. (1667-1429 cm-1) region that are dia-
gnostic of aromatic structure. These occur near 6.25, 6.32, 6.67, and 6.90 µm (—1600,
1580, 1500, and 1450 cm-1) and are caused by C=C skeletal in-plane vibrations. The
second band is frequently observed only as a shoulder of the first, but is intensified if
the aromatic nucleus is conjugated with some unsaturated group; the fourth band is
frequently obscured by strong absorptions resulting from —CH2—bending vibrations
if aliphatic groups are present. The absence of absorption by a compound in these
regions is fair assurance that the compound is not aromatic.
A number of absorption bands of variable intensity appear in the 10-15 µm (1000-
670)
Region that is caused by C—H bending vibrations. These absorptions depend on
the-number of adjacent free hydrogen atoms that an aromatic nucleus contains. An
aromatic compound containing five adjacent hydrogen atoms absorbs strongly in both
the 13.3 and 14.3 μm regions (~750 and 700 cm’”1); if the compound contains four
adjacent hydrogen atoms, as, for example, an o-disubstituted benzene, it absorbs
strongly only near 13.3 μm. (~750 cm”1). The remaining absorptions owing to fewer
adjacent hydrogen atoms (higher degree of substitution on the aromatic nucleus)
are usually weak and not easily assigned. Of particular importance for benzene
compounds is the absorption near 14.3 μm (~700 cm”1); if the compound does not
absorb strongly in this region, it cannot be a mono substituted benzene compound.
The spectra of biphenyl Fig. above and other mono substituted benzene compounds
show these absorptions.
The region 5-6 μm (2000-1670 cm-1) of the spectra of benzenoid compounds contains
absorption bands of low intensity that are overtone or combination bands. The number
and relative position of these bands are remarkably dependent upon the particular
substitution type of the benzene ring.
African Virtual University 88
Formative Assessment
Interpretation of IR Spectra
Unit V
Molecular Spectroscopy 2: Nuclear Magnetic Resonance
http://www.scienceofspectroscopy.info/edit/index.php?title=NMR_Spectros-
copy
http://en.wikipedia.org/wiki/NMR_spectroscopy#Chemical_Shift
http://www.mhhe.com/physsci/chemistry/carey/student/olc/ch13nmr.html#basi
http://teaching.shu.ac.uk/hwb/chemistry/tutorials/molspec/uvvisab4.htm
http://www.scienceofspectroscopy.info/edit/index.php?title=UV-Visible_Spec-
troscopy
http://www.scienceofspectroscopy.info/edit/index.php?title=UV_Absorption_Ta-
ble- non open source
UV visible
http://ull.chemistry.uakron.edu/analytical/Spectrophotometry/
http://www.chem.ucla.edu/cgi-bin/webspectra.cgi?Problem=bp1&Type=C
http://www.chem.ucla.edu/~webspectra/search.html
African Virtual University 90
Formative Assessment
Proton NMR
The most useful nuclei for organic NMR the proton Mass Num 1 A.M 1 and Carbon
13 because they occur in many organic compounds.
The proton has a spin number of ½ the magnetic nucleus may therefore assume any
one of (2I + 1) ranging from -½ , to ½ in steps of 1 orientations with respect to the
direction of the applied magnetic field. Thus, a proton (I = ½ ) will be able to assume
only one of two possible orientations that correspond to energy levels of ± μ.H in an
applied magnetic field, where H is the strength of the external magnetic field
Therefore these energy levels are said to be quantised.
A proton in a static external magnetic field may assume only two orientations cor-
responding to energies of ±μH. The low-energy orientation corresponds to that state
in which the nuclear magnetic moment is aligned parallel to the external magnetic
field, and the high-energy orientation corresponds to that state in which the nuclear
magnetic moment is aligned antiparallel (opposed) to the applied magnetic field. It
is possible to induce transitions between these two orientations; the frequency v of
electromagnetic radiation necessary for such a transition is given by v =-2μHo,/h.
where Ho is the strength of the external magnetic field. Note unlike the absorption in
UV and IR this absorption the frequency ν is dependent on the applied field.
African Virtual University 92
Under higher resolution the peaks of ethyl alcohol attributed to methylene and methyl
protons appear as multiplets.
The methyl CH3 absorption is split into three of relative area 1:2:1 and the meth-
ylene CH2 is split into four peaks of relative area 1:3:3:1. This is explained by the
methyl CH3 group interacts with CH2 group splitting into 4 and the methelene CH2
group interacts with the methyl CH3 group splitting it into 3 This effect is called the
spin-spin interactions
The magnitude of multiple separation resulting from spin-spin interactions is inde-
pendent of the strength of the applied field.
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Hydrogen Bonding
Protons that are involved in hydrogen bonding (this usually means -OH or -NH) are
typically observed over a large range of chemical shift values. The more hydrogen
bonding there is, the more the proton is deshielded and the higher its chemical shift
will be. However, since the amount of hydrogen bonding is susceptible to factors
such as solvation, acidity, concentration and temperature, it can often be difficult to
predict.
The power and usefulness of 1H NMR spectroscopy as a tool for structural analysis is
much appreciated. Unfortunately, when significant portions of a molecule lack C-H
bonds, no information is forthcoming. Examples include polychlorinated compounds
such as chlordane, polycarbonyl compounds such as croconic acid, and compounds
incorporating triple bonds (structures below, orange colored carbons).
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When acquired in this manner, the carbon NMR spectrum of a compound displays a
single sharp signal for each structurally distinct carbon atom in a molecule (remem-
ber, the proton couplings have been removed). The spectrum of camphor, shown on
the left below, is typical. Furthermore, a comparison with the 1H NMR spectrum on
the right illustrates some of the advantageous characteristics of carbon NMR. The
dispersion of 13C chemical shifts is nearly twenty times greater than that for protons,
and this together with the lack of signal splitting makes it more likely that every
structurally distinct carbon atom will produce a separate signal. The only clearly
identifiable signals in the proton spectrum are those from the methyl groups. The
remaining protons have resonance signals between 1.0 and 2.8 ppm from TMS, and
they overlap badly thanks to spin-spin splitting.
Unlike proton NMR spectroscopy, the relative strength of carbon NMR signals is
not normally proportional to the number of atoms generating each one. Because of
this, the number of discrete signals and their chemical shifts are the most important
pieces of information delivered by a carbon spectrum. The general distribution of
carbon chemical shifts associated with different functional groups is summarized in
the following chart. Bear in mind that these ranges are approximate, and may not
encompass all compounds of a given class. Note also that the over 200 ppm range of
chemical shifts shown here is much greater than that observed for hydrogen shifts
African Virtual University 98
13
c Chemical Shift Ranges*
Low Field
Region
*
For samples in CDCl3 solution. The δ scale is relative to TMS at δ=0.
Formative Assessment
Figure 34 : C10H13NO2
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2- The carbon-13 NMR spectrum of one of the butyl acetate isomers (C4H9OCOCH3)
showed signals at δc22, 28, 80 and170. What is its structure? Why is the intensity
of the peak at δ 28 much more intense than that at δ 22 (by factor of approxima-
tely eight)? How would the multiplicity and signal intensity in the proton NMR
spectrum of this compound confirm your deductions?
African Virtual University 100
Unit VI
Mass Spectrometry
http://www.cem.msu.edu/~reusch/VirtualText/Spectrpy/spectro.htm#contnt
http://riodb01.ibase.aist.go.jp/sdbs/cgi-bin/cre_index.cgi?lang=eng
http://ull.chemistry.uakron.edu/analytical/Mass_Spec/index.html/
Mass Spectrometry
Mass spectrometer identifies compounds by ionizing the compound and breaking the
compound into pieces called fragment and analyzing these fragments by passing the
pieces through analyser. The analyser sorts the fragments according to their mass
charge ratios. The results of the analyser are displayed as a mass spectrum.
A small sample of compound is ionized, usually to cations by loss of an electron-The
Ion Source
The ions are sorted and separated according to their mass and charge.- Mass Ana-
lyzer
The separated ions are then detected and tallied, and the results are displayed on a
computer.
Because ions are very labile (as they would react with ambient species) their formation
and analysis is conducted in a vacuum.
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Ion Source
In the ion source molecules of the sample are bombarded by high energy electrons
coming from a heated filament accelerated across an electric field. Ions formed by the
electron bombardment are pushed away by a charged repeller plate, and accelerated
toward other electrodes, having slits through which the ions pass as a beam. Some of
these ions fragment into smaller cations and neutral fragments. Therefore the initial
molecule results in a large number of cations which is formed into an ion beam.
Isotopes
Since a mass spectrometer separates and detects ions of slightly different masses, it
easily distinguishes different isotopes of a given element. This is manifested most
dramatically for compounds containing bromine and chlorine, as illustrated by the
following examples. Since molecules of bromine have only two atoms, the spectrum
on the left will come as a surprise if a single atomic mass of 80 amu is assumed for
Br. The five peaks in this spectrum demonstrate clearly that natural bromine consists
of a nearly 50:50 mixture of isotopes having atomic masses of 79 and 81 amu respec-
tively. Thus, the bromine molecule may be composed of two 79Br atoms (mass 158
amu), two 81Br atoms (mass 162 amu) or the more probable combination of 79Br-81Br
(mass 160 amu). Fragmentation of Br2 to a bromine cation then gives rise to equal
sized ion peaks at 79 and 81 amu.
bromine
methylene chloride
vinyl chloride
Figure 35 : Mass spectra of Bromine, Vinyl Chloride, Methylene Chloride
African Virtual University 102
The center and right hand spectra show that chlorine is also composed of two isotopes, the
more abundant having a mass of 35 amu, and the minor isotope a mass 37 amu. The precise
isotopic composition of chlorine and bromine is: Chlorine: 75.77% 35Cl and 24.23% 37Cl
Bromine: 50.50% 79Br and 49.50% 81Br
The presence of chlorine or bromine in a molecule or ion is easily detected by noticing
the intensity ratios of ions differing by 2 amu. In the case of methylene chloride, the
molecular ion consists of three peaks at m/z=84, 86 & 88 amu, and their diminishing
intensities may be calculated from the natural abundances given above. Loss of a
chlorine atom gives two isotopic fragment ions at m/z=49 & 51amu, clearly incor-
porating a single chlorine atom. Fluorine and iodine, by contrast, are monoisotopic,
having masses of 19 amu and 127 amu respectively. It should be noted that the pre-
sence of halogen atoms in a molecule or fragment ion does not change the odd-even
mass rules given above.
Two other common elements having useful isotope signatures are carbon, 13C is 1.1%
natural abundance, and sulfur, 33S and 34S are 0.76% and 4.22% natural abundance
respectively. For example, the small m/z=99 amu peak in the spectrum of 4-methyl-
3-pentene-2-one (above) is due to the presence of a single 13C atom in the molecular
ion. Although less important in this respect, 15N and 18O also make small contributions
to higher mass satellites of molecular ions incorporating these elements.
Fragmentation Patterns
The nature of the fragments provides a clue to the molecular structure, but if the
molecular ion has a lifetime of less than a few microseconds it will not survive long
enough to be observed. Most organic compounds give mass spectra that include a
molecular ion, and those that do not if different ionisation conditions are used the
molecular ion may be observed.
Among simple organic compounds, the most stable molecular ions are those from
aromatic rings, other conjugated pi-electron systems and cycloalkanes. Alcohols,
ethers and highly branched alkanes generally show the greatest tendency toward
fragmentation. The stable fragments will appear in the final spectrum.
Hydrocarbons
The mass spectrum of dodecane on the right illustrates the behavior of an unbranched
alkane. Since there are no heteroatoms in this molecule, there are no non-bonding
valence shell electrons. Consequently, the radical cation character of the molecular ion
(m/z = 170) is delocalized over all the covalent bonds. Fragmentation of C-C bonds
occurs because they are usually weaker than C-H bonds, and this produces a mixture
of alkyl radicals and alkyl carbo-cations. The positive charge commonly resides on
the smaller fragment, so we see a homologous series of hexyl (m/z = 85), pentyl
(m/z = 71), butyl (m/z = 57), propyl (m/z = 43), ethyl (m/z = 29) and methyl (m/z =
African Virtual University 103
Formative Assessment
A
Number of distinct carbon atoms: ❑
Number of distinct hydrogen groups: ❑
Number of distinct carbon atoms: ❑
B
Number of distinct hydrogen groups: ❑
Teaching Tips
This module aims at presenting the most common instrumental analytical techniques
to the learners.
With three principal aims of providing
• Knowledge of principles of the analytical techniques
• Skills for interpreting analytical data generated by instrument s,
• Practice for the skills and knowledge delivered
For undergraduate level students there is the delicate task of managing the level of
complexity of information and skills delivered. For this module there are lots of re-
sources many of them intended for the practicing professional and advanced graduate
student and therefore unsuitable for our purposes. The material was therefore selected
and presented in a simple form to provide a working knowledge of the subject matter,
leaving room for the more enthusiastic student to pursue the matter further, without
confusing the average student.
The basic teaching aid is the material presented in the module which forms the back
borne of the course. The recommended texts and online materials are required to
add depth to the understanding of the module by providing detail and extra practice
for the learner.
The major task of the e learning teacher is to encourage the learner and pace him
through each learning unit. Each learning unit should be covered and mastered before
advancing to the next unit.
The material presented should be preferably covered in the order presented in the
module.
African Virtual University 111
XIX. References
Crow D.R. : Principles and applications of Electrochemistry Chapman and
Hall, 2nd Edition 1996
Galen Wood Ewing Instrumental methods of chemical analysis
Publisher: MacgrawHill. 1986
Braun. D Robert Introduction to chemical Analysis Publisher: McGrawHill 1st
Edition 1982.
Heslop R.B, Wild Gillian M.. S I Units in chemistry
Applied Science Publishers, 1971.
Hobarth Willard, Lynne Merritt, John Dean, and Frank Settle, Instrumental
Methods of Analysis Wadsworth Publishing Company; 7 Sub edition (Fe-
bruary 1988)