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analysis
By: Yehualashet T. (BSc, MSc. Of pharmaceutical Analysis & Quality Assurance)
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Cont…
Quantitative calculations are common everyday practice for
pharmacists engaged in laboratory work.
This lesson therefore aims at familiarizing students with
such calculations mainly done in pharmaceutical analysis.
It exposes the reader to various approaches for problem
solving and aids in consolidating theoretical knowledge
by applying it to the solution of real problems.
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Expression of concentration
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Cont…
Example: What is the % v/v of a solution that has 5.0 mL
of hydrochloric acid (HCl) diluted to 100 mL with
deionized water?
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Percentage weight in volume (%w/v)
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Molarity
The molar concentration (Cx) of the solution of the chemical
species X is the number of moles of that species that is
contained in one liter of the solution (not one liter of the
solvent).
The unit of molar concentration is molarity, M, which has the
dimensions of mol L-1.
Cx= no mole solute = no m mole
solute no L solution
no ml solution
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Examples
1. Suppose you had 0.2 moles of solute dissolved into 100 ml of solution.
What's the molarity of the solution?
⚫ Normality (N) is defined as the number of equivalents of solute dissolved in one liter
of solution.
N= no of equivalent weight no of equivalent weight =
weight
⚫ The number of replacing units, n, is based on a reaction unit, which is that part of a
⚫ Normality makes use of the chemical equivalent, which is the amount of one
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chemical species reacting stiochiometrically with another chemical species.
Dilutions
It is the process of decreasing the concentration of a stock solution by
pharmaceutical analysis
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Dilutions……..
In order for an extract from a formulation or a solution of a pure drug
substance to be measured, it must be diluted so that it falls within
the working range of the instrument used to make the measurement.
Thus an understanding of dilution factors is fundamental to
calculations based on analytical data.
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Calculation example
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Self test
2. A 5 ml sample of an injection of a steroid phosphate was diluted to 100 ml.
Then 10 ml of the diluted injection was diluted to 100 ml and this dilution
was further diluted 10 to100 ml. From measurement by UV the diluted
sample was found to contain 0.249 mg/100 ml of the steroid. What was the
original concentration of the injection in %w/v and in mg/ml? Answers:
0.498% w/v, 4.98 mg/ml
3. 50.0 mg paracetamol is dissolved in 100.0 ml methanol in a volumetric
flask. From this solution, 1.00 ml is picked up by a transfer pipette and
diluted to 100.0 ml with methanol in a new volumetric flask. What is the
concentration of paracetamol in the final solution?
Answer:
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Physical and chemical properties of
drug molecules
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Physical and chemical properties of drug molecules
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Cont.…
A deeper understanding of the analytical method requires
knowledge about both the analytical technique and the
chemical properties of the analytes.
Chemical and physical behavior of drug molecules, influence
the development of analytical methods.
Here, the chemical and physical behavior of drug molecules
that influence the development of analytical methods.
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Cont.…
most drugs used in medicine behave in solution as either weak acids
or weak bases.
The pKa and pKb values give information about the strength of
acids and bases; they tell you the pH at which 50% of the drug is
ionised.
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How Do We Measure pH?
For accurate
more
measurements, one uses a pH
meter, which measures the
voltage in the solution.
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pH
The hydrogen ion concentration or pH is a measure of
the
acidity or alkalinity of a solution.
In an aqueous solution, the product of hydrogen
ion concentration and hydroxyl ion concentration is constant.
The pH of solution is defined as the negative logarithm of the
hydrogen ion concentration, in an aqueous solution
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Cont.…
A pH meter is a scientific instrument that measures the
hydrogen-ion activity in water-based solutions, indicating its
acidity or alkalinity expressed as pH
pH is the unit of measure that describes the degree of
acidity or alkalinity. It is measured on a scale of 0 to 14
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pH value of aqueous solutions
Dissociation of water
The pH of a solution is defined as –log [H+]
pH = - log[H+] = log1/[H+]
pKw
= pH + pOH = 14
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Cont.…
If an acid is introduced into an aqueous solution , the [H+] increases
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Strong acids and bases
The common strong acids are HCl, HClO4, HNO3, H2SO4 and
the common strong bases are NaOH, KOH
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Cont.….
Calculate the pH of the following solutions:
i. 0.05 M HCl
ii. 0.05 M NaOH
iii. 0.05 M H2SO4.
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pH for weakly acidic and basic solutions
weak acid
Weak acids and bases are those that are not completely dissociated in
aqueous solution and are in equilibrium with the undissociated acid.
the extent of dissociation is characterized by Ka
Consider weak acid HA in a solution at total solute concentration of C
The dissociation reaction is HA Ka H+ + A
- Ka = [H+][A-] / [HA]
Not completely dissociates, [H+] is less than C
Ka
HA H+ + A- ,[H+] = [A-], [HA] = c – [H+], [HA] is
undissociated one
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Cont.….
Ka = [H+]2/(c - [H+]), if weakly dissociated, c -
[H+]~ c
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Weak acids…..
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Weak base……….
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Self test
Calculate the pH of the following solutions:
i) 0.1 M HCOOH or formic acid (Ka = 1.77 x 10-4)
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Factors Affecting Acid Strength
The more polar the H-X bond and/or the weaker the H-X bond, the
more acidic the compound.
Acidity increases from left to right across a row and from top to bottom
down a group.
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Buffers
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Buffer cont…
The simplest type of buffer is composed of a weak acid or base in
combination with its conjugated a strong base or acid.
A common buffer system is the sodium acetate/acetic
acid buffer
system.
The most direct way of preparing this buffer is by the addition of
sodium hydroxide to a solution of acetic acid until the required pH is
reached.
pH of buffer is solution determined from Henderson–Hasselbalch equation
Buffer of weak acid and its salt:
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Cont….
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Cont….
Suppose we are making a buffer solution of ammonium
(NH4+) and ammonia (NH3). In this example, ammonium is
our weak acid and ammonia is the conjugate base. The
reaction is as follows.
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Cont…
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Problem:
Calculate the pH of a solution containing 0.200 M NH3
and 0.300 M NH4Cl given that the acid dissociation
constant for NH4+ is 5.7x10-10.
(0.200)
pH = 9.244 + log
(0.300)
pH = 9.07
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Limitation of HH
When using the Henderson-Hasselbalch to make a buffer solution the
ratio should be close to one.
This makes the most stable solution.
When the ratio moves far away from one, the capacity to buffer the
solution decreases as a small disturbance changes the pH dramatically.
The Henderson-Hasselbalch equation does not provide accurate pH
values for extremely dilute buffer solutions because it does not account
for the self-dissociation that occurs in water.
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Limitation….
If an acid can easily dissociate more than one hydronium ion
(H+) the equation does not account for that and may not work
well.
The Henderson-Hasselbalch equation cannot be used for
strong acids and strong bases
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Ionisation of drug molecules
When a weakly acidic or basic drug is administered to the body, the drug will
ionize to a greater or lesser extent depending on its pKa and the pH of the body
fluid in which it is dissolved.
The pH of the body varies widely, but the most important biological solution is
the blood, normally has a pH of 7.4.
The ionisation of drug molecules is important with regard to
their absorption into the circulation and
their distribution to different tissues within the body
Drugs are better absorbed in unionized form.
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Example:
1. Calculate the percentage of ionisation of the drugs shown in Figure
below, at pH 7.0.
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Example………..
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Example 2:
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Drug stability
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Drug stability
Drug stability means the ability of the pharmaceutical dosage form to
maintain the physical, chemical, therapeutic and microbial properties
during the time of storage and usage by the patient
The purpose of drug stability testing is:
– to provide evidence on how the quality of a drug
substance or drug product changes with time under the influence
of environmental factors such as temperature, humidity and light.
The aim of the testing is to
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Factors affecting Stability
1- Environmental factors : -
Temperature Light , Oxygen, Moisture &
2
Carbon
Drugsdioxide
or excipients in the dosage form: Particle size
of drug & pH of the vehicle
3 Microbial contamination
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Stability is used to determine
Quality of a drug substance or drug product
Shelf life for the drug product
Recommended storage conditions
Why stability testing is necessary :
Chemical degradation may lead lowering of concentration
of drug in dosage form
toxic product may form due to degradation of
active ingredients
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Cont….
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Types of stability
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A. Physical stability
The formulation is totally unchanged throughout its shelf life
and has not suffered any changes by way of appearance,
organoleptic properties, hardness, brittleness, particle size etc.
It is significant as it affects:
pharmaceutical elegance
drug content uniformity
drug release rate.
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Types of physical degradation are
2. Loss of H2O
3. Absorption of H2O
4. Crystal growth
5. Polymorphic changes
6. Color changes
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1. Loss of water
This can be seen in the following dosage forms:
a. Emulsions: Loss of water lead to separation of the two phases and change to
other type
substances added to aqueous phase to absorb water from atmosphere and prevent
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2. Crystal formation
Crystal formation in pharmaceutical preparations: Causes:
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3. Colour Changes:
Colour changes are of two types.
1. Loss of color due to PH change , Presence of reducing agent
2. Development of color · Exposure to light
Preventive measures for physical instability
Loss of volatile components, Loss
of water & Absorption of
water:
Products should be placed in well-closed
For solutions: Stabilizers are added
container.
Crystal
For suspensions · Minimum
growth: flocculation should be
temp. Incorporation of surface active managed,
agent ·
Color By increasing viscosity
changes: PHof should
suspendingnot be Exposure to light should be
material
changed
avoided and, An attempt has been made to prevent the fading by incorporating UV light
absorbing material.
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B. Chemical stability:
It implies lack of any decomposition in the chemical moiety
that is incorporated in the formulation as the drug,
preservatives or any other excipients
If any decomposition occur, it may influence the physical
and chemical stability of the drug
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Types of chemical degradation are as under
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i. Hydrolysis
“It is defined as the reaction of a compound with water.”
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Some Functional Groups Subject to Hydrolysis
Drug type Examples
Esters Aspirin, alkaloids
Dexmethasne sodium phosphate
Nitroglycerin
Lactones Pilocarpine
Spironolactone
Amides Chloramphenicol
Lactams Penicillins
Cephalosporins
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Hydrolysis Preventive Measures:
Adjustment of pH: Rate of decomposition is critically
dependent Upon pH.
Choice of solvent
Addition of surfactants
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ii. Oxidation
Oxidation of a compounds is explained is defined as loss
of electrons or gain of oxygen.
Thioethers Chlorpromazine
Carboxylic acids Fatty acids
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Factors lead to oxidation
Presence of oxygen, Light (It can cause photo-chemical reaction, i.e. chemical reaction
PH : each drug has its ideal pH for stability. Any change in pH affect drug stability and
Pharmaceutical dosage form; Oxidation reaction occur in solutions faster than in solid
dosage forms.
Type of solvent used : Oxidation reaction occur faster in aqueous solution than others.
Presence of unsaturated bonds : as double and triple bonds (oils) which undergo easier
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Factors induce Polymerization
1. Temperature
2. Light
3. Solvent
4. pH
5. Impurities
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Iv. Isomerization
It means conversion of drug to its isomer
reading assignment
1. Optical isomerization
2. Geometric isomerization
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C. Microbiological stability
Microbiological stability implies that: formulation has not
suffered from any microbiological attack and is meeting the standards
with respect to lack of contamination/sterility.
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CONT…
Sources Microbial Contamination: Water, Air, Raw
materials,
of containers and closures, Personnel & Instruments
and apparatus
To prevent contamination to the formulation during storage
1. suitably designing the containers
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Stability Testing
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Stability: is the capacity of product to remain
drug within
specification established to ensure its identity, strength, quality,
and purity
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Stability Testing
Provide evidence as to how the quality of the drug
product
varies with time.
Establish shelf life for the drug product
Determine container closure system suitability
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Order of reaction
Refers to the way in which the concentration of the reactant (s)
influence the rate of a chemical rxn.
1. Zero-order degradation
Zero order rxn: the rate of rxn is independent of
the concentration of the reactants and constant with respect to
time
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Zero-order degradation
In zero-order kinetics the rate of degradation is independent of
the concentration of the reactants.
Thus, if the rate constant for the zero-order degradation of a substance is
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Mathematically
where k0 is the zero-order rate constant [concentration(C)/time(t)].
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First-order degradation
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EXAMPLE 3
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Stability protocol
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Testing Frequency
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Accelerated stability testing
Study to predict the shelf life of the product, by accelerating the rate of
decomposition, preferably by increasing the temperature of reaction
conditions.
With the advancement in branch of kinetics, shelf life of a dosage form
can be predicted within months based on accelerated stability reports
Preparations are subjected to high stresses during stability testing.
Common high stresses include Temperature , Humidity and Light
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Thank you
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