Introduction to Pharmaceutical Analysis

AAU, CHS, School of Pharmacy

Minichil C.(B. Pharm, Msc Student)

 Objectives
 students are able to:-
 define pharmaceutical analysis, QC, QA and related terms.
 discuss on law & pharmaceutical analysis.
 discuss the contents of pharmaceutical monographs & their
role (USP, BP).
 classify the analytical errors &validation parameters of method
development.
 exercise the d/t units of expressing concentration and
percentage/content.
 PH, Partition coefficient, ……

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 Introduction:
 Pharmaceutical Analysis deals with qualitative and quantitative
analysis of drugs, their metabolites or impurities
Quality, Safety and Efficacy of pharmaceutical products.

 It is often described as the area of pharmaceutical chemistry

responsible for characterizing the composition of matter?,
both
Qualitatively
 Quantitatively.

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 Intro…
 Pharmaceutical analysis is not simply performing routine
analysis on routine samples , but
improves in improving established methods,
extending existing methods to new types of samples, and
 developing new methods for measuring chemical
phenomena.

 “. . .the science of inventing and applying the concepts,

principles, and . . .strategies for measuring the
characteristics of chemical systems and species”.

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 Intro…

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 Scope of pharmaceutical analysis:
 Pharmaceutical industry:

Raw material control

In-process control
Dosage form control

 Govt. drug control lab. (Regulatory Agencies):

Manufacture lab. Procedure control.

Products control.

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 Scope …
 Pharmaceutical research labs

 Advanced research
– Development of analytical method
– Dosage form evaluation
– Stability studies

 Basic research
– Separation
– Identification
– Quantization
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 Other Applications:
 Hospital pharmacy
Clinical pharmacy--- TDM

 Community pharmacy

Pramit Dey ‘’no matter what cpds you discover or

formulation you make nothing is valid until it is
evaluated, analysed and validated’’.

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 Intro…
 The Quality of a product may deviate from the std required.
 But the quality of the analysis itself is of the std required.

 Testing a pharmaceutical product involves chemical, physical

and sometimes microbiological analyses.

 Quality of pharmaceutical products is essential to assure the

maximum level of pt’s satisfaction.
 safety, potency, efficacy, stability, pt acceptability &
regulatory compliance.

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 Introduction to QC & QA
QC- Quality + Control

 Quality---inspection/measurement approach used in QC when

quality consists of conformity with a predetermined
specification.

‘Quality is the degree to which a specific product conforms

to a design or specification’.

ISO:“Fitness for purpose with customer

satisfaction’’
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 QC & QA….
 QC
checking or testing that specifications are met
checking that the product is fit for purpose.

 The aim of QC is to evaluate whether the sample of drug

complies with the appropriate specifications.

 QC is integral to all modern industrial processes.

Pharmaceutical industry.

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 QC & QA….
 Medicines are exposed to strict quality control.

Why?
 Medicines are special because virtually no other product is
consumed by the public on such utter trust-
trust that the medicine will not do them more harm than the
illness it is meant to cure.
the belief that it will cure their ill health and it will not make it
worse!

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 QC & QA….

 Manufacturing Impacts of Low-Quality

process
 Packaging
Medicines
 Transportation
Therapeutic failure or
 Storage ? drug resistance.
condition
MEDICINE Toxicity,
QUALITY Prolonged illness.
 death.
Loss of customer’s
confidence in the brand,
Loss of credibility
Enormous economic
13 losses .
 QC & QA….
 QC alone is considered as testing Quality into the
product.
only accept those materials that pass specification.

 QC is not adequate for the following reasons.

Only what is expected to be found is tested
Only small portion of the bulk of the material is tested.
There could be a small proportion of defective material
in a batch
It is doubtful whether, in the case of medicines, the
consumer would detect defects.
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 QC & QA….
 To assure the quality of medicinal products, Quality must be
built at each stage of the manufacturing process and not merely
tested in.
Any factor that could have an effect on the quality of the final
medicinal product must be controlled.

 These factors could be

anything from the design of the production facility used
the environment the material is isolated in,
the analytical test methods employed at each stage of
production.

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 QC & QA….
 QA --- batch to batch consistency should be maintained

 Hence, if written procedures control all of these factors and

trained personnel follow these procedures, then a product
consistently meeting its predetermined specification should be
produced.
 End product testing is then becomes just a final check of the quality of
the product.

 This testing is then used in conjunction with the written records

as the supporting documentation to allow the material to be
released for use.
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 QA
Is the sum total of the organized
arrangements with the objective of
ensuring that products will be of the
quality required for their intended use

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 GMP
Is that part of QA aimed at EU GMP
ensuring that products are
US GMP
consistently manufactured
to a quality appropriate to
their intended use

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 QC & QA….

Who Ensures Medicine

Quality?  Drug regulatory
authority

 Drug and  Physicians

Therapeutics and other
Committee Medicine prescribers
Quality

 Hospital  Pharmacy
procurement (and dispensers)
office  Patients
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 QC vs QA
 function limited - testing  defines systems and methods to
methods ensure quality of a product-
“Controls the product” in  Ensures that stds & procedures
production, are followed and delivered
 tests various aspects of a product products or services meet
as it is being developed performance requirements.
 Operational laboratory  All those planned or systematic
techniques and activities used to actions necessary to provide
fulfill the requirement of Quality adequate confidence that a
product will satisfy the
requirements for quality,
Laboratory based
 Includes process checklists &
project audits.
Company based
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 The Law and Pharmaceutical Analysis
 Assuring continuous availability of the required
pharmaceuticals of proven
• Quality,
• Safety and
• Efficacy
requires pharmaceutical policy which does have legal instrument

 pharmaceutical legislation and pharmaceutical regulation.

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 Law….FDA
 Drugs could be bought and sold like any other consumer good.

 Regulation become more stringent

 medical technology improved and high profile incidents highlighted the
need for at least some degree of gov.t monitoring.

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 Law….FDA
 FDA began an initiative to address CGMPs for the 21st Century.
 taking a new look at both the regulatory & industrial systems for
ensuring drug quality.

FDA is the federal agency responsible for ensuring that

 Foods are safe, wholesome and sanitary
 Human and veterinary drugs, biological products and medical devices
are safe and effective
 Cosmetics are safe
 Electronic products that emit radiation are safe.

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 Local drug regulation (Ethiopia)
Major Duties and Responsibilities
1. Sets stds of quality, safety and efficacy of drugs
2. Sets stds of competence and issues, controls, renews, revokes, suspends
certificate of competence
3. Undertake inspection of pharmaceutical premises, establishments and
port of entry and exit.
4. Prepares categories and revises national list of drugs.
5. Formulates police, and prepares draft legislation.
6. Control quality of raw and packing materials used for the production of
drugs and finished products.
7. Prepares stds of quality, safety and efficacy of traditional drugs
8. Serves as DIC
9. Controls Narcotic and psychotropic drugs
10. Creates favorable conditions for the promotion and expansion of drug
trade?
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 Local…
FMHACA
 Scope of Control  Institutions Controlled
 Human and veterinary  Manufacturers
drugs  Importers and/or wholesalers
 Radio-pharmaceuticals
 Exporters
 Traditional medicines
 Retail-outlets
 Pesticides?
 Drug QC Labs
 Medical supplies and
equipments  Scientific offices,
 Sanitary items  Commission agents
 Cosmetics  Health institutions
 Raw and packaging materials

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 Compendia /Pharmacopeia/
 Pharmacopeia- book
 It contains a list of medicinal substances (drugs) with
descriptions,
specifications, USP
BP
tests, and
IP
 acceptance criteria, EUP
JP
specified by some recognized authority.

 In most countries, the recognized authority that issues the

books of stds is the gov.ment
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 Compendia….
 The pharmacopeias are revised and supplements are issued b/n
revisions in order to keep them as to up to date as possible.

 Quality of pharmaceuticals is based on pharmacopeia

specifications /stds.

 Std is required because

 To protect the user against drugs of poor quality.
 To guarantee/ ensure/ that the analytical methods used are std, keep
abreast of recent developments both in analytical techniques and in
quality considerations of the drug in question.

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 Compendia….
 Monograph- a written description of the principal feature
of the substance.
 the ways that the features can be determined.

 Definition of the nature of material

 Statement of the minimum std of purity as determined by assay
 Description of its physical characteristics
 Tests for use in verifying the identity of the product
 Limit tests to exclude excessive contamination
 Quantification & identification procedures
 Other information
 Packaging and storage conditions
 Labeling, dosage form
 Cautionary notices on cytotoxic and other such materials

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 Compendia….
 General Notices- Provide the overall guiding principles
for using the monographs and general chapters.
Provide in summary form the basic guidelines for
interpretation and application of the stds, tests, assays ….

 General tests and assays- Contain information on basic

test procedures.
 Apparatus for tests and assays
 Microbial tests
 Biological Assays
 Chemical tests and assays
 Physical tests and determinations
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 Analytical Errors
–"No amount of experimentation can ever prove me right; a
single experiment can prove me wrong." —Albert Einstein

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 Analytical Errors
 A quantitative analysis is not great deal of use unless there is some
estimation of how prone to error the analytical procedure is.

 Simply accepting the analytical result could lead to rejection or

acceptance of a product on the basis of a faulty analysis.

 usual to make several repeat measurements of the same sample.

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 Errors…
Error- the difference b/n a measured, calculated or observed
value and the "true" or accepted value.
1. Systematic errors- determinate
 It affects the accuracy of an analysis

 It is characterized by a systematic deviation from the true value.

 all the individual measurements are either too large or too
small.

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 Errors…
 Sampling error- error introduced during the process of
collecting a sample for analysis.
 Method error- error due to limitations in the analytical
method used to analyze a sample.

 Measurement error- error due to limitations in the

equipment and instruments used to make measurements.
 Personal error- error due to biases introduced by the
analyst.

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 Errors…
2. Random Errors- indeterminate errors
 Errors that cause results to fluctuate in a random fashion about the
"true" or accepted value.
completely random in nature.

 The errors are indicated by small differences in successive

measurements made by the same analyst under almost identical
experimental conditions.

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 Validation of Analytical Methods
 Qualification action of proving that any
EQUIPMENT works correctly and actually leads to the
expected results.

 Identification of particular attributes of equipment, utilities

or processes related to the performance of a particular
function.

 DQ, IQ, OQ & PQ

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 Validation,,,
 Method validation- the process used to confirm that the
analytical procedure employed for a specific test is suitable for
its intended use.

 Results from method validation can be used to judge the

quality, reliability and consistency of analytical results.

integral part of any good analytical practice!

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 Validation,,,
 Analytical methods need to be validated or revalidated:-

Before their introduction into routine use;

Whenever the conditions change for which the method has
been validated
Whenever the method is changed and the change is outside
the original scope of the method.

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 Validation parameters
1. Accuracy- the extent to which test results generated by
the method and the true value agree.

 The closeness of agreement b/n the true or accepted reference

value, and the value found.

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 Validation parameters ….
2. Precision- closeness of agreement (degree of scatter) b/n a
series of measurements
 obtained from multiple sampling of the same homogeneous sample under
the prescribed conditions.

-intra-assay precision

 %RSD should be less than or equal to 2.

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 Validation parameters ….
3. Specificity -the ability to assess clearly the analyte in the
presence of components which may be expected to be present.
 impurities, degradants, matrix, etc.

4. Sensitivity
 The sensitivity of method indicates how responsive it is to a
small change in the concentration of analyte.
 change in signal per unit change in the amount of analyte.

5. Range- is the interval b/n the upper and lower

concentration (amounts) of analyte in the sample

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 Validation parameters ….
6. Linearity- its ability (within a given range) to obtain test
results which are directly proportional to the concentration
(amount) of analyte in the sample.
r of > 0.96 is regarded as indicating linearity.

7. Robustness- a measure of its capacity to remain unaffected

by small, but deliberate variations in method parameters.
indication of its reliability during normal usage.

 It refers to how resistant the precision and accuracy of an assay

is to small variations in the method

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 Validation parameters ….
8. LoD- The detection limit of an individual analytical
procedure is the lowest amount of analyte in a sample which can
be detected but not necessarily quantitated as an exact value.
 It is the smallest concentration or absolute amount of analyte
that has a signal significantly larger than the signal arising from a
reagent blank.
LoD = 3 σ/s

9. LoQ -the lowest amount of analyte in a sample which can be

quantitatively determined with suitable precision and accuracy.
LoQ = 10 σ/s

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 Expressions of concentration
 Concentration --- the amount of solute present in a known
amount of solution.
 The actual units for reporting concentration depend on how
the amounts of solute and solution are measured.

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 Expressions…
 Although, in chemistry, all concentrations are expressed in
molarity, pharmaceutical analysts have to contend with the
medical profession.
 tends to prescribe drugs in units of mass per volume
(weight per milliliter).

 The most common way to express the concentration of active

drug in a medicine is in terms of
 mass of API/100g
 volume of API/100ml.

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 Expressions…
% w/v- the number of grams of drug in 100 ml of final
product.
the active ingredient is a solid.
 solutions, suspensions, etc.
 Example, 5% dextrose infusion.

% v/v - the number of ml of drug in 100 ml of final product.

the active drug and the final product are both liquids.
 Example 70% ethanol contains.

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 Expressions…
% w/w- the number of grams of drug in 100 g of final
product.
Powders, creams, ointments…
 Example: 1% salicylic acid ointment.

% v/w - the number of ml of drug in 100 g of final product.

 Only encountered in ointments and creams where the active
ingredient is a liquid,
 Example. 1% glycerol ointment.

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  ppm= mass of solute X 106 (For very dilute solutions)
mass of solution

 ppb = mass of solute X 109

mass of solution

 Example: 0.106 g ethanol dissolved in 999.911 g water. The

concentration of ethanol in ppm.
 Ans:106 ppm

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 Expressions…
 Normality makes use of the chemical equivalent, which is
the amount of one chemical species reacting stoichiometrically
with another chemical species.
N =EW per unit volume,

 Although a solution of H2SO4 has a fixed M, its N depends on

how it reacts.

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 Expressions…
 The number of equivalents, n, is based on a rxn unit,
which is that part of a chemical species involved in a rxn.

Precipitation rxn, n=the charge of the cation or anion.

Acid–Base rxn, n=the number of H+ ions donated by an
acid or accepted by a base
Complexation rxn, n=the number of electron pairs that
can be accepted by the metal or donated by the ligand.
Oxidation–Reduction rxn, n= the number of electrons
released by the reducing agent or accepted by the oxidizing
agent.

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 Practice Questions
1. What is the Molarity of the solution if 10.0 g of sodium carbonate
(M.wt =106 gm/mol) is dissolved in 200 mL of solution?
 Ans. = 0.47 M
2. What will be the molarity of the solution if 3.5gm of sodium
hydroxide (M.wt = 40 gm/mol) are dissolved to make 150 ml of
solution?
 Ans. = 0.583 M
3. Calculate the molar concentration of ethanol in aqueous solution
that contains 2.30 gm of ethanol (46.07 gm/mol) in 3.50 L
solution.
 Ans. = 0.0143 M
4. Calculate the normality of 2.56 gm/L of Na2CO3 (M.wt =106
gm/mol)
 Ans. =0.0483 M
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 Preparing Solutions by Dilution
 Solutions with small concentrations are often prepared by
diluting a more concentrated stock solution.

 A known volume of the stock solution is transferred to a new

container and brought to a new volume.

Since the total amount of solute is the same before and after
dilution,

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 Practice Questions
1. Describe how to prepare 250 ml of 0.25 M HCl acid from 6 M
HCl stock solution.
Ans. = take 10. 42 ml of HCl, add to small amount water and
dilute to the mark (250 ml
2. 2 ml volume of eye drops containing the local anaesthetic
proxymetacaine HCl is diluted to 100 ml and then 5 ml of
the dilution is diluted to 200 ml. The diluted sample was
measured by UV spectrophotometry and was found to
contain 0.512 mg/100 ml of the drug. Calculate the %w/v of
the drug in the eye drops.
 Ans= 1.024% w/v.

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 M1
Physicochemical properties of Drugs
 The ability of a chemical cpd to elicit a therapeutic effect is related to the influence of
various physicochemical properties

 Physical properties of drug molecules, along with simple

chemical derivatisation and degradation reactions, play an
important part in the design of analytical methods.

 Drug molecules can be complex

 Multiple functional groups.

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Slide 54

M1 Minichil, 10/28/2021
 Physicochemical properties of Drugs
PH Calculation
 The pH of a solution is defined as –log[H+].

The PH of H2O is thus

–log[10-7] =7

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 Physico…
 Strong acids and bases are completely ionized in water.

 The concentration of [H+] and [OH-] is equal to the molarity

of the acid and the base, respectively.

Example: 0.1M HCl and 0.1M NaOH

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 Physico…
 Weak acids are not completely ionized in aqueous solution
and are in equilibrium with the undissociated acid, which is a
very weak acid.

 For instance in a 0.1 M solution of acetic acid (Ka= 1.75 x10-5)

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 Physico…

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 Physico…
 Example, in a 0.1 M solution of ammonia (Kb =1.8 x 10-5)
the equilibrium can be written as follows:

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 Physico…
Henderson-Hasselbach Equation

For Acids

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 Physico…
Buffer Solutions
 A buffer solution is a solution that resists changes in pH.
If acid is added then, within reason, the pH does not fall; if
base is added, the pH does not rise

 Buffers are usually composed of a mixture of weak acids or

weak bases and their salts.
 Human plasma is buffered at pH 7.4 by a carbonic acid/bicarbonate buffer system.

Buffer ratio = [Salt]/[Acid] or [Salt]/[Base].

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 Physico…
 Buffers are used in the
Preparation of mobile phases for chromatography and
Extraction of drugs from aqueous solution.

 Example sodium acetate/acetic acid buffer system.

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 Physico…
Ionization of Drugs
 When a weakly acidic/ basic drug is administered to the body,
the drug will ionize
 to a greater/ 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,

Blood has a pH of 7.4.
 Design of analytical
methods
 ADME
 • Extraction
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Q. Ibuprofen is an OTC drug with the carboxylic acid moiety
(COOH) the proton donor, which has a pKa of around 4.4.
What happens once it’s swallowed? Assume that the pH of
stomach is3 & that of large intestine is 8.

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 [S ] [S ]
3  4.4  log ,,,  10 1.4  0.03981
[ A] [ A]
0.03981
% _ Salt  x100  4%
1.03981
 Most of the drug is in acid form and therefore could cross the stomach lining to enter
the bloodstream; i.e., ibuprofen can be absorbed from the stomach.
 Next the large intestine, assuming pH = 8
[S ] [S ]
8  4.4  log ,,,  103.6  3981
[ A] [ A]
3981
% _ Salt  x100  99.97%
3982

 Most of the drug is in salt form & so shouldn’t readily cross the intestinal lining &
enter the bloodstream.

65
 Physico…
Partition Coefficient
 When a substance (or solute) is added to a pair of immiscible
solvents, it distributes itself b/n the two solvents according to
its affinity for each phase.

 Partition law- ‘a given substance, at a given temperature,

will partition itself between two immiscible solvents in a
constant ratio of concentrations’.
partition coefficient

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 Physico…
Q. The distribution of 100 mg of a drug X between 50 ml of
chloroform and 50 ml of water was allowed to equilibrate in a
separating funnel and. When the organic layer is analysed, it is
found to contain 66.7 mg of compound. From these data,
calculate P and the percentage of the drug extracted into the
organic layer.

% Drug extracted=
66.7/100 *100= 66.7%.
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 Physico…
Importance of P
 Extraction and chromatography of drugs.
 Predict the absorption, distribution and elimination of drugs
within the body.
 predict the onset or the duration of action of drugs, or to tell whether a
drug will be active at all.

 RDD… SAR, where P is used relate the biological activity of a

drug to its physical and chemical characteristics.

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 Physico…
Stability of Drugs
 Drugs have quite complicated chemical structures.
result in their decomposition & deterioration.

Oxidation & Hydrolysis.

 Shelf-life (t90) --the length of time the product may safely

be stored on the dispensary shelf before significant
decomposition occurs.
 It is time for decomposition of 10% of the active drug to
occur, leaving 90% of the activity.
 determined from stability study.
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 Physico…
Measurement of Optical Rotation
 R/S
 The light waves vibrate randomly in all directions
perpendicular to the direction of propagation of the wave.

 If normal light of this type is passed through a material that is

itself chiral (e.g. Nicol prism), then the waves of light interact
with the chiral material to produce light that is oscillating in
only one plane.
 plane-polarized light.

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 Physico…
 When plane-polarized light is passed through a solution
containing an optically active substance, the chiral cpd causes
the plane of vibration of the light to rotate.
 The number of degrees of rotation can be measured.

 Where: [α] = specific optical rotation, α = measured rotation in degrees, l= length of sample tube
in decimeters (1 dm = 10 cm), c = concentration of sample in %w/v.

Applicable in Enantiomeric Purity Measurement.

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 Physico…
Q. Optical rotation measurements were made using the sodium
D line at 25 0C in a 1 dm cell and the readings obtained were as
follows:
 Phenylephrine HCl 2.6% w/v in 0.1 M HCl; -0.98. Calculate
[a] for this drug .
Ans: -37.7.

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 Thank YOU a lot!!!

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