Stability testing of natural products.docx

Stability is defined as the capacity of drug to remain within established specification limits to maintain its identity, strength, quality and purity throughout the retest or expiration dating period.
It is the ability of formulations to retain its physical, chemical, microbiological and toxicological parameters same that time of manufacturer.

1. MPG 104 T. INDUSTRIAL PHARMACOGNOSTICAL
TECHNOLOGY
M.pharm 1
st
semester
UNIT-4 TESTING OF NATURAL PRODUCTS:
STABILITY TESTING OF NATURAL PRODUCTS
INTRODUCTION:
Stability is defined as the capacity of drug to remain within established
specification limits to maintain its identity, strength, quality and purity
throughout the retest or expiration dating period.
It is the ability of formulations to retain its physical, chemical, microbiological and
toxicological parameters same that time of manufacturer.
PURPOSE OF STABILITY TESTING :
The purpose of stability testing is to provide evidence on how the quality of drug
substance varies with time under various environmental changes.
The four climatic zones-
The design of the stability testing programme should take into account the
climatic condition in the area in which the drug products will be used.
I. Zone I – temperate
II. Zone II – Sub tropical with high possibility of humidity
III. Zone III – hot/dry
IV. Zone IV – hot /humid
Advantages:
 Help in determination of shelf life and storage conditions.

2.  Help in selection of suitable formulation, excipients and container closure
systems for a product.
 Well being of the patient and manufacture by ensuring the good product
quality.
 Enhance patient confidence in herbal drugs and improve compliance.

Objectives:
 To provide evidence on how the quality of active substance varies with time
and environment factors.
 To establish re-test period for active substance.
 To establish shelf life of finished products.
 To evaluate the efficacy of drug.
 To develop suitable packing information for drug product.
 To submit stability information for regulatory agencies.

Factors affecting stability testing:
There are various factors that affects the stability testing of natural products.
 Temperature- chemical changes increase with increase in temperature.
 Light – many chemical changes due to exposure to light .
Eg. Auto oxidation of volatile oils.
 Moisture content- absorption of moisture on solid surface increases
decomposition.
 Oxidation – additional of oxygen, radical formation and degradation of
compound.
 Hydrolysis- reduction with water leads to degradation of compound.
 Geometric isomerism- trans and cis form, one form may be more
therapeutically active.
 Drug interaction- reaction of two or more drugs or between a drug.

3. Role of markers:
Markers are chemically known compound, which may or may not have
therapeutic effect, used to calculate the quality of herbal medicinal ingredients in
herbal medicinal products (HMPs).
Typical sources for finding markers are –
1. Monograph and drafts
2. Experience, transfer from plants/ constituents
3. Literature research
4. Scientific research
Analytical method:
 A systemic approach should be adopted to the presentation and evaluation
of stability information, which should include, as necessary, physical,
chemical, biological and microbiology test characteristics.
 All product characteristics likely to be affected by storage.
E.g. assay value or potency, content of products of decomposition,
physicochemical properties should be determined. For solid or semi-solid
oral dosage forms, dissolution tests should be carried out.
 The analysis of preparation is mostly done by chromatography technique
like hplc, GC, TLC and spectroscopic technique like uv-visible spectroscopy
or by the combination of chromatographic and spectroscopic technique
such as LC-MS, GC-MS
 Analytical methods should be validated or verified, and accuracy as well as
the precision should be recorded.
Types of stability study:
Different types of stability study according to the condition to be maintained
throughout the shelf life of drug.
1. Physical stability study.
2. Chemical stability study.

4. 3. Microbiological stability study.
4. Therapeutic stability study.
5. Toxicology stability study.
1.Physical stability study-
 The original physical properties, namely, appearance, uniformity,
dissolution and suspendability are maintained.
2.Chemical stability-
 Each and every active ingredient retains its chemical integrity as well as
potency specified on label, within the specified limits. It involves drug assay
and determination of drug degradation.
Material used for stability studies-
I. Active pharmaceutical ingredient ( API)
II. Finished pharmaceutical products.
Stages in stability testing-
I. Pre formulation.
II. Final product.
III. Post marketing.

3.Microbiology stability study-
 Sterility or resistance to microbial growth is maintained as per the specified
requirements.
4.Therapeutic stability study-
 The therapeutic effect remains unaltered.
5.Toxicology stability study-
 No valid increase in toxicity should occurs.

5. STABILITY TESTING:
 It is recommended that in case of a herbal medicinal product containing a
natural product or a herbal drug preparation with constituents of known
therapeutic.
Shelf life- The period of time during which a product, if stored correctly, is
expected to comply with the specification as determined by the stability studies
on a no. Of batches of the product. The shelf life is used to establish the expiry
date of each batch.
The determination of shelf life of herbal medicinal drug is same as chemically
defined APIs, but special nature of herbal product should be taken into
consideration.
Expiry date- The date after which something is no longer in effect. It is
established for each batch by adding the shelf-life period to the date of
manufacture.


Methods of stability testing:
1. Stress testing.
2. Accelerated stability testing.
3. Intermediate testing.
4. Forced degradation testing.
5. Long term stability testing products or real time testing.

6. Stress testing-
 Stress testing help to identify the degradation product, which can help to
establish the degradation pathway. Stress tests are usually considered
unnecessary for herbal drug and its preparation.
 Nature of stress testing depends on the individual drug substance and the
type of drug product involved.
 Generally conducted on a single batch of the drug substance.
 The testing should also evaluate the susceptibility of the drug to hydrolysis
across a wide range of pH values when in solution or suspension.
 Perform phytochemical analysis for constituents in given sample of herbal
medicinal products (HMPs) by HPLC, HPTLC.

Accelerated stability testing-
 The substance or medicinal product is challenged by a controlled
exaggerated stress (storage conditions) over a short time , so that the rate
of degradation reaction is enhanced.
 In short period of time, stability data can be obtained (3-6 months)
 Samples are stored at 40°C ± 2°±; 75% ± 5 % relative humidity (RH) for a
period of 6 months.
Procedure-
1) Note down description, identification, pH, viscosity, assay, heavy metals
and microbial purity.
2) Above parameter should match those specified in monographs or label.
3) Transfer HMPs
 Pet bottle with Al-cap, containing rubber cork.
 Glass bottle with Al-cap, containing rubber cork.
4) 1
st
set- 25°C ± 2°C/ 60% RH ± 5% ----- under controlled condition.
5) 2
nd
set - 40°C ± 2°C / 75% RH± 5% -----accelerated condition.
6) Carry out sampling at various intervals (0,1,2,3,4,5……months)

7. 7) Perform phytochemical analysis for constituents in given sample of HMPs
by HPLC, HPTLC.
Intermediate testing-
 Conducted when accelerated studies fail. At 25°C for longer duration of
time.
Forced degradation testing-
 Performed to provide intrinsic stability assessment of drug.
Long term stability testing or real time testing-
 Evaluation of the physical , chemical and microbiological characteristics of a
drug product over the expected duration of shelf-life under recommended
storage conditions and in the proposed container/closure system. The
results are used to establish the shelf-life, to confirm the projected shelf
life, and to recommend storage conditions.
 Derived storage condition for Real time testing is below.

Climatic
zone
Storage temperature (◦C) Relative humidity(%RH)
I 21 45
II 25 50
III 30 35
IV 30 70

Challenges in stability testing of herbal medicinal products (HMPs):
 Stability testing of herbal products is a challenge because-
 The whole herbal product is regarded as the active matter, regardless of
active constituents with defined therapeutic activity.
 HMPs are complex in nature due to their high no. of constituents.
 Constituents sometimes have very low concentrations in the finished
product.

8.  Different requirements for the different types of extracts.
Steps to overcome challenges:
i. Use of markers for HMPs.
 Marker concentration should be within ±5%
 When marker concentration is less in finished product then variation of
±10%
ii. Use of fingerprint chromatograms.
iii. Use of GC-MS, HPLC, HPTLC and spectroscopic technique.




Stability testing storage conditions for drugs as per ICH & WHO:

Storage of stability samples:-
Intended
storage
conditions
Stability test
method
Temperature and
humidity as per ICH
Temperature and humidity
as per WHO
Room
temperature
Long term 25±2°C/60±5%RH 25-30±2°C/60-75±5%RH
Intermediate 30±2°C/65±5%RH 30±2°C/65±5%RH
Accelerated 40±2°C/75±5%RH 40±2°C/75±5%RH
Refrigerator Long term 5°C ambient 5±3°C
Accelerated 25±2°C/605%RH 25±2°C/60±5%RH or
30±2°C/65±5%RH
Freezer Long term -20°C/ambient (12) -20°C±5°C

9. Test schedule for stability of new products
Method and climatic
zone
Environmental Frequency of testing
Long term for climatic
zones I and IV
25°C/60%RH 3,6,9,12,18,24,36 months
Long term for climatic
zones III
30°C/35%RH 3,6,9,12,18,24,36 months
Long term for climatic
zones Iva/ intermediate
for zones I and II
30°C/65%RH 3,6,9,12,18,24,36 months
Long term for climatic
zone IVb/ intermediate
for zones I and II
30°C/75%RH 3,6,9,12,18,24,36 months
Accelerated condition
for all zones
40°C/75%RH 3,6 months
F


PROTOCOL FOR STABILITY TESTING :
Protocol is a document describing the basic components of a well-controlled
stability study.
A well-controlled stability protocol should contain the following information:-
 Selection of batches and samples
 Test selection
 Analytical procedures
 Acceptance criteria
 Storage conditions
 storage period
 Testing frequency
 Sampling plan

10.  Container closure system
 Evaluation
 Statement labelling

1. Selection of batches and samples
 In general , this selection should constitute a random sample from
pilot or production batches that may involve a single batch or 2-3
batche.
2. Test selection
 The test that monitor the quality , potency , purity ,and identity that are
expected to vary upon storage are chosen as stability tests.
3. Analytical procedures
 Procedures given in the official compendia should be followed and if alternate
methods are to be used they need to be duly validated.
4. Acceptance criteria
 This should be fixed beforehand in form of statistical limits for the results
manifested in computable terms and pass or fail for qualitative tests.
5. Storage conditions
 After the stability of the product has been evaluated, following conditions are
recommended for storage.
 Store under normal storage conditions.
 Store between 2 and 8 ◦C ( under refrigeration, no freezing.)
 Store below 8 ◦C ( under refrigeration)
 Store between -5 and -20 ◦C ( in a freezer)
 Store below -18 ◦C ( in a deep freezer)
 These are based upon the marketing climatic zone of the drug.
 Storage condition such that thermal stability as well sensitivity to moisture can
be tested.
6. Storage period

11.  It generally extend from minimum of 3-6 months in accelerated and
stress testing and upto 12,18 or 60 months in ongoing ot follow-up
stability testing.

7. Testing frequency
 It should be sufficient to establish the stability profile of the drug test
schedules for different types of stability testing.
8. Sampling plan
 It involves devising for the no. of samples to be placed in the stability
chambers and taking out of the charged batch so as to cover the
entire study.
9. Container closure system
 The testing in actual containers as well as closures scheduled for
marketing, are to be tested separately with proper orientation of
storage of containers.
10.Evaluation
 The data on attribute is analysed to determine the time duration at
which 95% one-side confides limit for the mean curve intersects the
acceptance criterion.




11.Statement labelling
 A storage statement , retest period, and re test date based on the
stability evaluation of the drug substance should be established for
the labelling

12.  Labelling specification should match as those mentioned by
regulatory authority.