The document discusses various topics related to impurity profiling and stability testing of pharmaceutical drugs including definitions of impurities, classification of impurities, impurity profiling methods, factors affecting drug stability, types of stability testing, regulations for stability testing, and methods for accelerated stability testing and shelf life calculations. Impurities are classified as identified, unidentified, or according to USP and ICH guidelines. Various spectroscopic, separation, isolation, and characterization methods are described for identifying impurities.

1. Presented by:
CHANDU . CHATLA
Rig. no: Y18MPH301 M.PHARM
Under the guidelines of
Mrs.CH. SRUJANI
ASSISTANT PROFESOR
ACHARYA NAGARJUNA UNIVERSITY

2.  DEFINITIONS
 CLASSIFICATION OF IMPURITIES
 IMPURITY PROFILING
 STABILITY
 TYPES OF STABILITY
 REGULATIONS REQUIRED
 STABILITY TESTING METHODS
 ACCELERAED STABILITY AND SHELF LIFE
CALCULATIONS

3. Impurity: Any component of the new drug product that
is not the drug substance or an excipient in the drug
product.
• Identified impurity: An impurity for which
structural characterisation has been achieved.
• Unidentified impurity: An impurity for which
structural characterisation has not been achieved.

4. United States Pharmacopoeia (USP)
• According to USP impurities are classified into three sections
1. Impurities in Official Articles
2. Ordinary Impurities
3. Organic Volatile Impurities
• The ICH Terminology
According to ICH guidelines, impurities in drug substance
produced by chemical synthesis can be broadly classified
into following three categories
1. Organic Impurities (Process and drug-related)
2. Inorganic Impurities (Reagent, ligands, catalysts)
3. Residual Solvents (Volatile solvents)

5. • Impurity profile describes the identified and
unidentified impurities present in a new drug
substance.
• Impurity profiling is the common name of a group of
analytical activities, the aim of which is the detection,
identification/structure elucidation and quantitative
determination of organic and inorganic impurities as
well as residual solvents in bulk drugs and
pharmaceutical formulations.

6. • New drug development requires meaningful and reliable
analytical data to be produced at various stages of the
development .
a) Sample set selection for analytical method development
b) Screening of Chromatographic conditions and Phases,
typically using the linear solvent- strength model of
gradient elution's.
c) Optimization of the method to fine-tune parameters
related to ruggedness and robustness

7. The impurities can be identified
predominantly by following methods;
• Reference standard method
• Spectroscopic method
• Separation method
• Isolation method
• Characterization method

8. • The key objective of this is to provide clarity to the
overall life cycle, qualification and governance of
reference standards used in development and control
of new drugs.
• Reference standards serve as the basis of evaluation of
both process and product performance and are the
benchmarks for assessment of drug safety for patient
consumption.
• These standards are needed, not only for the active
ingredients in dosage forms but also for impurities,
degradation products, starting materials, process
intermediates, and excipients.

9. SPECTROSCOPIC METHODS :
• The UV, IR, MS, NMR and Raman spectroscopic
methods
• are routinely being used for characterizing
impurities.
SEPARATION METHODS :
• The Capillary electrophoresis (CE), Chiral
Separations, Gas Chromatography (GC)
• Supercritical Fluid Chromatography (SFC), TLC,
HPTLC, HPLC are regularly
being used for separation of impurities and
degradation products.

10. • It is often necessary to isolate impurities. But if the
instrumental methods are used, isolation of
impurities is avoided as it directly characterizes the
impurities.
• Generally, chromatographic and non-
chromatographic techniques are used for isolation of
impurities prior its characterization.

11. A list of methods that can be used for
isolation of impurities is given below :
• Solid-phase extraction methods
• Liquid-liquid extraction methods
• Accelerated solvent extraction methods
• Supercritical fluid extraction
• Column chromatography
• Flash chromatography
• TLC
• GC
• HPLC
• HPTLC
• Capillary electrophoresis (CE)
• Supercritical fluid chromatography (SFC)

12. CHARACTERIZATION METHODS :
• Highly sophisticated instrumentation, such as MS
attached to a GC or HPLC, are inevitable tools in
the identification of minor components (drugs,
impurities, degradation products, metabolites) in
various matrices. For Characterization of
impurities, different techniques are used; which
are as follows:
NMR and MS

13. • The ability of NMR to provide information regarding the
specific bonding structure and stereochemistry of molecules
of pharmaceutical interest has made it a powerful analytical
instrument for structural elucidation.
• The ability of NMR- based diffusion coefficient determination
to distinguish between monomeric and
dimeric substances was validated using a standard mixture of
authentic materials containing both monomers and dimers .
• Unfortunately, NMR has traditionally been used as a less
sensitive method compared to other analytical techniques.
Conventional sample requirements for NMR are on the order
of 10 mg, as compared with MS, which requires less than 1
mg.

14.  It has an increasingly significant impact on the pharmaceutical
development process over the past several decades.
 Advances in the design and efficiency of the interfaces, that
directly connect separation techniques with Mass
Spectrometers have afforded new opportunities for
monitoring, characterizing, and quantification of drug related
substances in active pharmaceutical ingredients and
pharmaceutical formulations.
 If single method fails to provide the necessary selectivity,
orthogonal coupling of chromatographic techniques such as
HPLC-TLC and HPLC-CE, or coupling of chromatographic
separations with information rich spectroscopic methods such
as HPLC-MS or HPLC-NMR may need to be contemplated, but
hopefully only as a development tool rather than a tool for
routine QC use.

15. HYPHENATED METHODS:
• LC-MS-MS
• HPLC-DAD-MS
• HPLC-DAD-NMR-MS
• GC-MS
• LC-MS
• A common goal for investigation of both process
and product degradation-related impurities is to
determine which of the many potential impurities
are, in fact, produced in the manufacturing process
and which occur under a given set of storage
conditions.

16. Definition:
• The capacity of a drug or product to remain within
established specifications of identity , quality, purity
in a specific period of time.
or
• The capacity or the capability of a particular
formulation in a specific container to remain with in
particular chemical , microbiological , therapeutically
, and toxicological specifications.

17. FACTORS EFFECTING DRUG STABILITY:
 The primary factors effecting stability :
• PH , Temperature , Moisture , humidity , light ,
Storage closure and containers , Oxygen
 The major factors effecting drug stability are :
• Particle size (suspension and emulsion) , PH ,
additives and molecular binding and diffusion of
drugs and excipients .
OBJECTIVES
1. To determine maximum expiration date/ shelf life.
2. To provide better storage condition.
3. To determine the packaging components.
4. To gather information during pre formulation
stage to produce a stable product.

19. TYPES OF STABILITY THAT MUST BE
CONSIDERED FOR ANY DRUG
 CHEMICAL:
• Each active ingredient retains its chemical integrity and labelled potency
within the specified limit.
 PHYSICAL :
• The physical stability properties includes appearance, palatability
,uniformity ,dissolution and suspend ability are retained.
 MICROBIOLOGICAL :
• Sterility or resistance to microbial growth is retained according to
specified requirement.
 THERAPEUTIC:
• Therapeutic activity remains unchanged .
 TOXICOLOGIC :
• No significant increase in toxicity occurs.

20.  Stability study requirement and expiration dates are
covered in the current GMP , USP and FDA
 GMP (Good Manufacturing Practice) states that
there will be written testing program design to
access testability characteristics of drug products .
And result of such stability testing will be used to
determine appropriate storage condition and
expiration dates

21. ICH GUIDELINES FOR STYABILITY TESTING
• The ICH has so far released six guidelines for stability studies
as indicated in table :
ICH GUIDELINES TITLE
Q1A Stability testing of new drug substances and products (second
revision)
Q1B Stability testing : photo stability testing of new drug substance
and products.
Q1C Stability testing for new dosage forms
Q1E Bracketing and matrixing designs for stability testing of drug
substances and products
Q1D Evaluation of stability data
Q1F Stability data package for registration application in climatic
zones III and IV

22. AS per ICH and WHO guidelines ,world has
been divided into four zones :
ZONE 1 - TEMPERATE
ZONE2 - SUBTROPICAL WITH POSSIBLE HIGH HUMIDITY
ZONE 3 - HOT, DRY
ZONE 4 - HOT,HUMID

23. • Stability testing is a routine procedure performed on drug
substances and products and is employed at various stages of
the product development.
• Depending upon the aim and steps followed, stability testing
procedures have been categorized into the following four
types.
1. Real-Time stability testing.
2. Accelerated stability testing.
3. Retained sample stability testing.
4. Cyclic temperature stress testing.

24.  In real-time stability testing, a product is stored at
recommended storage conditions and monitored until it fails
the specification.
 In accelerated stability tests, a product is stored at elevated
stress conditions (such as temperature, humidity, and pH).
 Degradation at the recommended storage conditions can be
predicted using known relationships between the acceleration
factor and the degradation rate.

25.  In accelerated stability testing, a product is stressed at
several high (warmer than ambient) temperatures and
the amount of heat input required to cause product
failure is determined.
 This is done to subject the product to a condition that
accelerates degradation.
 This information is then projected to predict shelf life or
used to compare the relative stability of alternative
formulations.
 This usually provides an early indication of the product
shelf life and thus shortening the development schedule.
 In addition to temperature, stress conditions applied
during accelerated stability testing are moisture, light,
agitation, gravity, pH and package

26. procedure:
• Store the pharmaceutical at different higher
temperature.
• Assay the pharmaceutical product at different time
intervals.
• Determine the order of degradation by graphical
method.
• Form the graph, find out the rate constant for drug
degradation at different temperature.

27. • Construct an Arrhenius plot log K vs. 1/T and
extrapolate the straight line to room temperature.
from this , find the rate constant and shelf life at
25◦C . The Arrhenius equation and graph given
below:

28.  This is a usual practice for every marketed product for which
stability data are required. In this study, stability samples, for
retained storage for at least one batch a year are selected.
 If the number of batches marketed exceeds 50, stability samples
from two batches are recommended to be taken. At the time of first
introduction of the product in the market, the stability samples of
every batch may be taken, which may be decreased to only 2% to
5% of marketed batches at a later stage.
 In this study, the stability samples are tested at predetermined
intervals i.e. if a product has shelf life of 5 years, it is conventional
to test samples at 3, 6, 9, 12, 18, 24, 36, 48, and 60 months.
 This conventional method of obtaining stability data on retained
storage samples is known as constant interval method

29. Retained sample stability testing :
 Stability testing by evaluation of market
samples is a modified method which involves
taking samples already in the market place and
evaluating stability attributes.
 This type of testing is inherently more realistic
since it challenges the product not just in the
idealized retained sample storage conditions,
but also in the actual marketplace .

30. • This is not a routine testing method for marketed
products.
• In this method, cyclic temperature stress tests are
designed on knowledge of the product so as to
mimic likely conditions in market place storage.
• The period of cycle mostly considered is 24 hours
since the diurnal rhythm on earth is 24 hour, which
the marketed pharmaceuticals are most likely to
experience during storage.

31. • The minimum and maximum temperatures for the
cyclic stress testing is recommended to be selected
on a product by- product basis and considering
factors like recommended storage temperatures for
the product and specific chemical and physical
degradation properties of the products. It is also
recommended that the test should normally have 20
cycles

33. SHELF LIFE
 Shelf life is the period of time, from the date of
manufacture, that a drug product is expected to
remain within its approved product specification
while stored under defined conditions.
 Shelf life is typically expressed in units of months,
i.e. 24 months, 36 months, to a maximum of 60
months.

34. • The Preparation is stored at different elevated
temperatures, to accelerate the degradation
• Samples are withdrawn at different time intervals
• The Order of the reaction is determined by plotting
the appropriate function of concentration against
time and linear relationship is determined
• Straight line in a graph permits the estimation of k
value from the slope
• Similarly graphs are drawn for different elevated
temperatures.
• K value for each temperature are calculated.
• By using Arrhenius relationship, Log k values are
plotted against reciprocal of absolute temperature,
energy of activation can be calculated.

35. • Extrapolate the straight line to room temperature
(k25) or refrigerated temperature and read the log k
value on y-axis.
• Substitute the k value in the appropriate equation to
get the shelf life of the product.

36. • Arrhenius plot for predicting the rate constant at
ambient temperature(25ºC).

37. • Patrick J.Sinko , Martin’s Physical Pharmacy and
Pharmaceutical Sciences.
• Text book of physical pharmacy by cvs. subrahmanyam.
• Journal of applied pharmaceutical sciences “Stability
Testing of Pharmaceutical Products” Sanjay Bajaj, Dinesh
Singla and Neha Sakhuja.
• Journal of pharmaceutical sciences and reserch “impurity
profiling theory and practices” P. Venkatesan and
K.Valliappan.
• INTERNATIONAL JOURNAL OF PHARMACY & LIFE
SCIENCES Advance approaches for the impurity profiling
of pharmaceutical drugs: A review S. J. Ingale*, Chandra
Mohan Sahu, R.T. Paliwal, Shivani Vaidya and A.K.
Singhai