Dr.

Talib Hussain
STABILITY ANALYSIS PhD Pharmaceutics
Lecturer, IPS, UVAS, Lahore.
METHOD OF STABILITY ANALYSIS
The storage conditions (temperature and humidity) and lengths of storage
for stability studies should be sufficient to cover the
Storage, Shipment, and Subsequent use.
For submission of drug application, the data for long - term storage
should cover a minimum of 12 months on three primary batches.
The testing should, however, continue for the duration of the proposed
shelf life and retest period.
The accelerated and, when necessary, intermediate (storage condition
designed to moderately increase rate of chemical degradation or physical
change) storage conditions should be carried out for 6 months.
 STORAGE CONDITIONS FOR STABILITY
EVALUATION OF APIS
Stability Study Type Stability Storage Conditions Minimum Time Period Covered
by Data at Submission (months)

Marketed API Intended for Room Temperature (General Case) [7] Storage Conditions
Long term 25 ° C ± 2 ° C, 60% RH ± 5% RH or 30 ° 12
C ± 2 ° C, 65% RH ± 5% RH

Intermediate 30 ° C ± 2 ° C, 65% RH ± 5% RH 6
Accelerated 40 ° C ± 2 ° C, 75% RH ± 5% RH 6
Marketed API Intended for Storage in Refrigerator
Long term 5°C±3°C 12
Accelerated 25 ° C ± 2 ° C, 60% RH ± 5% RH 6
Marketed API Intended for Storage in Freezer
Long term -20 ° C ± 5 ° C 12
 SAMPLING PROTOCOLS
ICH guidelines recommends testing every 3 months over the first
year, 6 months over the second year, and annually thereafter
throughout the retest period under long - term storage condition.
For a 6 - month accelerated storage stability condition, sampling
at 0, 3, and 6 months is recommended.
When significant change to established test specification occurs
under accelerated storage condition, sampling at time 0, 6, 9, and
12 is recommended for a 12 - month intermediate storage
condition.
DEFINITION OF CLIMATIC ZONES AND RECOMMENDED
LONG- TERM STABILITY CONDITIONS
Climatic Definition Criteria Long-Term Test
Zone [Mean Annual Temperatures Condition
Measured in Open Air (° C) [Temperature ( ° C)
and Mean Annual Partial and RH]
Vapor Pressure (hPa)]
I Temperate climate ≤15 °C, ≤11 hPa 21 ° C, 45% RH
II Subtropical and >15 – 22°C, >11–18 hPa 25 ° C, 60% RH
Mediterranean climate
III Hot and dry climate >22 ° C, ≤15 hPa 30 ° C, 35% RH
IVA Hot and humid >22 ° C, >15 – 27 hPa 30 ° C, 65% RH
climate
IVB Hot and very humid >22 ° C, >27 hPa 30 ° C, 75% RH
climate
PARAMETERS OF STABILITY STUDIES
❑Stability of Toxicological Formulations (Toxic properties i.e. ADRs)
❑Liquid State Stability (Solution state of drug)
❑Solid State Stability (Drug excipient compatibility & Storage conditions)
❖SUB PARAMETERS OF STABILITY ANALYSIS
➢Thermal stress testing (From method of stability test)
➢pH stress testing (extreme pH 1-8, Cosolvent, Ionic strength)
➢Oxidative stress testing (Oxygen with antioxidants)
➢Photostability (Light stress testing)
 STABILITY OF TOXICOLOGICAL
FORMULATIONS
➢Toxic limit of the dose (effects on cells and tissues
causing damage or ADRs)
➢The stability of toxic dose (LD50) (Dose at which
risk to benefit ratio is +ve)
➢The appearance of toxic response before storage
or formulation and then evaluating its response
after converting to solution as well as blending with
solid excipients and stored for specified period.
 LIQUID STATE STABILITY
Solution stability is required to designate the drug candidate
suitable for liquid dosage forms i.e. Syrups, Suspensions,
Emulsions, lotions etc.
Solid dosage forms also dissolve in GIT and converted to
liquid state before absorption and after absorption they stay in
blood and tissues.
➢Aqueous buffers are used to prepare solution over wide range
of pH (1-8) with constant level of drug, cosolvent and ionic
strength.
 LIQUID STATE STABILITY CONT.…
➢Constant ionic strength is required for parenteral
administration. Ionic strength of NaCl 0.9% solution is 0.15.
1
➢Formula µ = σ 𝑚1 𝑍1 2
2
•Where m = concentration of ions & Z = valence
➢Cosolvents are selected to achieve drug concentration
necessary to achieve analytical sensitivity i.e. alcohols and
organic solvents
➢Stability solutions prepared are sealed in glass ampoules by
flame sealing and stored at temperatures less than boiling point
of volatile cosolvents, and are analyzed according to the time of
stability method e.g. Long-term, intermediate or accelerated
OXIDATIVE DEGRADATION ANALYSIS
➢Presence of oxygen and light can be responsible for degradation of drug as
head space of ampoule contains oxygen.
➢Seal the drug solution in glass ampoules with oxygen as head space
➢ Seal the drug solution in glass ampoules with vacuum as head space
➢Seal the drug solution in glass ampoules with nitrogen (inert) as head space
➢Seal the drug solution in glass ampoules with oxygen as head space and
organic antioxidants such as BHT or BHA (Butylated Hydroxy
Anisole/Toluene)
➢Seal the drug solution in glass ampoules with oxygen as head space and
inorganic antioxidants such as Sodium metabisulphite
➢Store the samples according to stability method selected & evaluate for drug
 OXIDATIVE DEGRADATION ANALYSIS
RESULTS
1) If drug is stable in the presence of oxygen then there is no need to
change head space or addition of any antioxidants
2) If drug is not stable in presence of oxygen then check for containers
having nitrogen and vacuum as head space for stability. If
degradation occurs then its due to some solution incompatibility
other than oxygen.
3) If drug is not stable in presence of oxygen then check if it is stable
with antioxidants. Select the antioxidant from organic or inorganic
for formulation
4) If there is degradation due to incompatibility then select nitrogen or
vacuum as head space
PHOTODEGRADATION ANALYSIS
➢Presence of oxygen and light can be responsible for degradation
of drug as head space of ampoule contains oxygen.
➢Seal the drug solution in glass ampoules with oxygen, nitrogen,
vacuum as head space , oxygen with antioxidants in
1) Transparent glass container,
2) 2) Amber colored glass,
3) 3) Yellow glass container,
4) secondary packaging (aluminium foil)
➢Store the samples according to stability method selected & expose
samples to fixed light source preferably UV and IR radiations then
evaluate for drug concentration.
PHOTODEGRADATION ANALYSIS RESULTS

➢If drug is stable in transparent containers then there is no need to use

colored glass or secondary packaging
➢If drug is not stable in transparent then check the stability in amber and
yellow colored containers
➢If stable in amber colored container the pack in amber color and if
stable in yellow colored container then pack in transparent glass with
aluminium foil secondary packaging
 SOLID STATE STABILITY

For solid state stability analysis there are two

parameters to be evaluated

1. Storage conditions of NDM

2. Drug excipient compatibility analysis

 STORAGE CONDITIONS
➢Take 5 mg of drug sample (NDM) alone or dose of NDM and also blend it with
commonly used excipients to prepare solid dosage forms
➢Pack these in suitable glass containers (transparent, amber, transparent with Al++)
➢Pack the containers with oxygen, nitrogen, oxygens with antioxidant as head space
➢Expose these to UV-Visible light preferably 254nm and 366 nm with visible light.
➢Store these at stability temperature and humidity conditions for long-term,
intermediate, accelerated stability analysis.
➢Evaluate for temperature degradation also i.e. 50 ℃, 70 ℃, 90 ℃ with ambient
humidity conditions
➢Observe any polymorphic, color, or active ingredient quantity change on conditions
 DRUG EXCIPIENT COMPATIBILITY
Drug excipient compatibility is evaluated generally by two
techniques
A. Fourier transform infrared spectroscopy (FTIR)
B. Differential scanning calorimetry (DSC)
1. FTIR:
➢FTIR analysis measures a sample's absorbance of infrared light at
various wavelengths to determine the material's molecular
composition and structure.
➢The wave number on the infrared spectrum is plotted between
4,000 to 400 cm-1
 FTIR
➢Absorbance bands are assembled within two types:
➢ 1. Group frequencies and 2. Fingerprint frequencies.
➢Group frequencies are characteristic of small groups of atoms or
functional groups such as CH₂, OH, and C=O (> 1500cm-1), they’re
usually unique to a specific functional group.
➢Fingerprint frequencies, these are highly characteristic of the
molecule as a whole (< 1500cm-1), can verify the presence of molecule.
➢Interpreting FTIR spectra starts at the high frequency end to identify
the functional groups present. The fingerprint regions are then studied
to positively identify the compound. There are vast libraries of infrared
spectra available, allowing to compare unknown materials to ensure
quick and accurate identification
 DIFFERENTIAL SCANNING CALORIMETRY (DSC)
Thermal analysis used to find purity of the material (NDM) based on
melting point of molecules.
➢5 mg of NDM, excipients each, product formulation is packed in
aluminium pan separately and placed in heating mantle of the DSC
instrument.
➢Flush nitrogen to mantle to avoid any incompatibility due to oxygen or
moisture
➢Apply heat according to standard heating rate i.e.
➢2 ℃ / minute, 5 ℃ / minute, 10 ℃ / minute over a given range 20-250 ℃
➢The heat range vary according to the melting point of NDM and excipients
 DSC CONT.…
➢The results are obtained in the form of thermograms either
endotherms (generally) or exotherms.
➢Sharp peaks are usually obtained at the melting points of each
material (NDM, excipients)
➢Comparison of peaks are made for melting point peak presence in
formulation at same temperature as was with pure ingredient
➢The presence of same peaks in thermograms indicate compatibility
➢The shifting of peaks from original position describe interactions
➢The presence of new peak and absence of existing peak describe
incompatibility or chemical complexation.
A, B ARE EXCIPIENTS
C IS DRUG MOLECULE
D IS FORMULATION