7/26/2010
VERIFICATION OF
COMPENDIAL
PROCEDURES
ANNE-MARIE ALDERSON, MBA
COOK MYOSITE INC.
JULY 29, 2010
Introduction
What are “compendial methods”?
What analytical test methods are attendees
using?
What obstacles are being encountered with
method verification?
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Relevant References
USP-NF <1225> Validation of Compendial
Procedures
USP-NF <1226> Verification of Compendial
Procedures
ICH Q2B
FDA Draft version Guidance for Industry,
Analytical Procedures and Methods Validation
Additional Resources
Validation of Analytical Methods for
Biopharmaceuticals: A Guide to Risk-Based Validation
and Implementation Strategies
Author: Stephan O. Krause
Available at https://store.pda.org/bookstore/
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Terminology
Validation
† The
process by which it is established, by
laboratory studies, that the performance
characteristics of the procedure meet the
requirements for the intended analytical
application
Verification
† Assessing
selected analytical performance
characteristics to generate appropriate, relevant
data rather than repeating the validation process
Validation vs. Verification
Non-Compendial methods must be validated
† An
alternative method proposed for some
application instead of the regulatory analytical
procedure
Compendial methods can be verified
Regulatory analytical procedure in USP-NF
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Should You Validate or Verify?
USP <1225> states:
†
“According to these regulations [21 CFR
211.194(a)(2)], users of analytical methods described
in USP-NF are not required to validate the accuracy
and reliability of these methods, but merely verify their
suitability under actual conditions of use.”
USP <1226> states:
†
“Users of compendial analytical procedures are not
required to validate these procedures when first used
in their laboratories, but documented evidence of
suitability should be established under actual
conditions of use.”
Why is it necessary to verify an
approved compendial method?
Necessary to show suitability of test method under
actual conditions of use
Variable parameters may not be addressed in
USP-NF:
Different drug manufacturers may have different
impurity profiles
† Excipients in a drug product can vary among
manufacturers
† Different antioxidants, buffers, container extractives,
excipients, etc. may interfere with the compendial
procedure
†
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Regulatory Expectations
USP <1226>
† “Verification
consists of assessing selected
analytical performance characteristics, such as
those described in <1225>, to generate
appropriate, relevant data rather than repeating
the validation process.”
† “Users of compendial analytical procedures are
not required to validate these procedures when
first used in their laboratories, but documented
evidence of suitability should be established
under actual conditions of use.”
Regulatory Expectations
<1226>
† Only
those characteristics that are considered to
be appropriate for the verification of the particular
method need to be evaluated
† Degree and extent of verification may depend on
the level of training/experience of user, on the
type of procedure and its associated equipment
or instrumentation, on specific procedural steps,
and on which article(s) are being tested
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Regulatory Expectations
21 CFR 211.194(a)(2) – excerpts
† Laboratory
records must include complete data
for all tests necessary to assure compliance with
specifications and standards, including
examinations and assays
„ “Suitability
of all testing methods used shall be verified
under actual conditions of use”
Regulatory Expectations
<1225> and <1226> do NOT cover
microbiological procedures because they are
covered in…
<51> Antimicrobial Effectiveness
† <61> Microbiological Examination of Non-Sterile
Products: Microbial Enumeration Tests
† <62> Microbiological Examination of Non-Sterile
Products: Tests for Specified Microorganisms
† <71> Sterility Tests
† <1227> Validation of Microbial Recovery from
Pharmacopeial Articles
†
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Regulatory Expectations
<1226> “Verification is not required for basic
compendial test procedures that are routinely
performed, unless there is an indication that the
compendial procedure is not appropriate for the
article under test”
Loss on drying
† Residue on ignition
† Various wet chemical procedures
†
„
†
Acid value
Simple instrumentation methods
„
pH measurements
Typical Analytical Characteristics Used
in Method Validation/Verification
<1225>
Accuracy
Precision
Specificity
Detection Limit
Quantitation Limit
Linearity
Range
Robustness
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Accuracy
The closeness of test results obtained by that
procedure to the true value, established across
its range
Calculated as the percentage of recovery by
the assay of the known added amount of
analyte in the sample, or as the difference
between the mean and the accepted true
value, together with confidence intervals
Accuracy
ICH recommends…
†
Determining Accuracy in a Drug Substance
†
†
Using a minimum of 9 determinations over a minimum of 3
concentration levels covering the specified range
Application of analytical procedure to analyte of known purity/Reference
Standard
Comparison of results of the proposed analytical procedure with those of
a second well-characterized procedure, the accuracy of which is stated
or defined
Determining Accuracy in Drug in a Formulated Product
†
†
Application of analytical procedure to synthetic mixes of the drug
product components to which known amounts of analyte have been
added within the range of the procedure
When impossible to obtain samples of all product components, add
known quantities of the analyte to the drug product or compare to the
results obtained from a second, well-characterized procedure of which
the accuracy is stated or defined
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Accuracy Assessment
Can be assessed in a variety of ways
Recovery of the analyte (percentage recovery) across
the range of the assay
† Linearity of the relationship between estimated and
actual concentration
†
Statistically Preferred:
Confidence interval for slope be contained in an
interval around 1.0
† Slope is close to 1.0
†
Acceptance criterion depends on the assay, its
variability and the product
Precision
The degree of agreement among individual test results when the
procedure is applied repeatedly to multiple samplings of a
homogeneous sample
Expressed as the standard deviation or relative standard deviation
(coefficient of variance) of a series of measurements
Measure of either the degree of reproducibility or of repeatability of
the analytical procedure under normal operating conditions
† Reproducibility = use of the analytical procedure in different
laboratories (different analyst, different equipment)
† Intermediate precision = within-laboratory variation (different
days, different analyst)
† Repeatability = use of the analytical procedure within a lab over
a short period of time (same analyst, same equipment)
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Precision
Determined by assaying a sufficient number of
aliquots of a homogeneous sample to be able
to calculate statistically valid estimates of
standard deviation or relative standard
deviation
ICH recommends repeatability be assessed
using a minimum of 9 determinations covering
the specified range for the procedure
†3
concentrations, 3 replicates of each OR
† 6 determinations at 100% of test concentration
Accuracy vs. Precision
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Specificity
The ability to assess unequivocally the analyte in the
presence of components that may be expected to be
present, such as impurities, degradation products, and
matrix components
Identification Test
†
Purity Test
†
Ensures the identity of the analyte
Ensures that all the analytical procedures performed allow
an accurate statement of the content of impurities of an
analyte
Assays
†
Provide an exact result, which allows an accurate
statement on the content or potency of the analyte in a
sample
Specificity Determination
In ID tests:
†
The ability to select between compounds of closely
related structure that are likely to be present
„
Confirm by obtaining positive results from samples
containing the analyte, coupled with negative results from
samples that do not contain the analyte and by confirming
that a positive response is not obtained from materials
structurally similar or closely related to the analyte
Purity tests:
†
Spike the drug substance/product with appropriate
levels of impurities and demonstrate that these
impurities are determined with appropriate accuracy
and precision
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Specificity Determination
Assay tests:
†
†
Demonstrate that the procedure is unaffected by the
presence of impurities or excipients
Spike the drug substance/product with appropriate levels
of impurities or excipients and demonstrate that the assay
result is unaffected by the presence of the extraneous
materials
If impurity or degradation product standards
unavailable
†
Compare the results of samples containing impurities or
degradation products to a well-characterized procedure
(USP or other validated procedure)
„
Include samples stored under relevant stress conditions (light,
heat, humidity, etc)
Detection Limit / Limit of
Detection
The lowest amount of analyte in a sample that
can be detected, but not necessarily
quantitated, under the stated experimental
conditions
Substantiate that the amount of analyte is
above or below a certain level
Usually expressed as the concentration of
analyte in the sample
† Percentage,
parts per billion, etc.
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Detection Limit Determination
Analyze samples with known concentrations of
analyte; determine the minimum level at which
the analyte can be reliably detected
Normally not necessary to determine actual
detection limit
If it is required that an impurity can be detected
at 0.01%, should demonstrate that the
procedure will reliably detect the impurity at
that level
Detection Limit and background
noise
Compare measured signals from samples with
known low concentrations of analyte with
those of blank samples (See ICH)
Acceptable signal-to-noise ratios are typically
2:1 or 3:1
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Quantitation Limit
Lowest amount of analyte in a sample that can be
determined with acceptable precision and
accuracy under the stated environmental
conditions
Characteristic of quantitative assays for low levels
of compounds in sample matrices, such as
impurities in bulk drug substances and
degradation products in finished pharmaceuticals
Generally determined by the analysis of samples
with known concentrations of analyte and by
establishing the minimum level at which the
analyte can be determined with acceptable
accuracy and precision
Quantitation Limit
Compare measured signals from samples with
known low concentrations of analyte with
those of blank samples
Acceptable signal-to-noise ratios is typically
10:1
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Linearity
The ability to elicit test results that are directly, or
by a well-defined mathematical transformation,
proportional to the concentration of analyte in
samples within a given range
Linearity of relationship between concentration
and assay measurement
In some cases, concentration/measurement may
be transformed to achieve linearity
Log, square root, reciprocal
† Not always possible to achieve linearity – may have to
use a nonlinear model
†
Linearity Determination
Establish by visual examination of a plot of signals
as a function of analyte concentration of content
Use statistical methods
ICH recommends a minimum of 5 concentrations
normally be used
Calculate regression line
Regression data may provide a useful estimate of
the degree of linearity
Correlation coefficient, y-intercept, slope and residual
sum of squares should be submitted
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Range
The interval between (and including) the upper
and lower levels of analyte that have been
demonstrated to be determined with a suitable
level of precision, accuracy, and linearity using
the procedure as written
Verify that the analytical procedure provides
acceptable precision, accuracy, and linearity
when applied to samples containing analytes
at the extremes of the range as well as within
the range
ICH Range Recommendations
Assay of a drug substance or finished product
†
Determination of an impurity
†
50% to 120% of the acceptance criterion
Content uniformity
†
80% to 120% of test concentration
Minimum of 70% to 130% of test concentration,
unless a wider/more appropriate range based on
dosage form is justified
Dissolution testing
†
±20% over the specified range
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Robustness
A measure of a procedure’s capacity to remain unaffected
by small but deliberate variations in procedural parameters
listed in the procedure documentation and provides an
indication of its suitability during normal usage
May be determined during development of the analytical
procedure
Establish a series of system suitability parameters to
ensure the validity of the procedure is maintained
whenever used
† Stability of analytical solutions, different equipment,
different analysts
† In liquid chromatography: mobile phase pH and
composition, various lots or suppliers of columns,
temperature, flow rate
† In gas chromatography: various lots or suppliers of
columns, temperature, flow rate
Different Tests Require Different
Verification Plans
Category 1
†
Category II
†
Analytical procedures for determination of impurities in bulk
drug substances or degradation compounds in finished
pharmaceutical products, such as quantitative assays and
limit tests
Category III
†
Analytical Procedures for quantitation of major components
of bulk drug substances or active ingredients (including
preservatives) in finished pharmaceutical products
Analytical procedures for determination of performance
characteristics
Category IV
†
Identification tests
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Required Elements by Category
Category II
Analytical
Performance
Characteristics
Category I
Quantitative Limit Tests
Category III Category IV
Accuracy
Yes
Yes
*
*
No
Precision
Yes
Yes
No
Yes
No
Specificity
Yes
Yes
Yes
*
Yes
Detection Limit
No
No
Yes
*
No
Quantitation Limit
No
Yes
No
*
No
Linearity
Yes
Yes
No
*
No
Range
Yes
Yes
*
*
No
* May be required – depends on nature of the specific test
What happens if verification is not
performed?
Risk receiving a 483
Abundant 483’s in validation/verification
Searchable FDA Database:
† http://www.fda.gov/iceci/enforcementactions/warni
ngletters/default.htm#browse
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Example 483’s
April 27, 2009
Nostrum Pharmaceuticals, LLC
6. The accuracy, sensitivity, specificity, and reproducibility of test
methods employed by your firm have not been established [21
CFR § 2l1.165(e)]. Refer to FDA 483, Observations 7 and 11. For
example, the related substances method used by your firm has not
been adequately verified in that no testing has been performed
to determine if the method can detect two of the known
impurities of [(b)(4)] at the USP-specified limits … Furthermore,
the actual procedure used by your firm to test Sucralfate Tablets
USP 1 g for dissolution varied from the method validated by the
developing contract laboratory. The method of calculation from
the standard curve in the method validated by the developing
contract laboratory was not the same in the procedure you use
and your firm's policy on rounding of calculated test results,
outlined in your SOP [(b)(4)], had the practical effect of creating
wider limits of acceptability for the bracketing standards than
are specified in the validated method.
Example 483’s
APR 21 2008
Changzhou SPL Company, Ltd (aka "Kaipu")
3. The test methods performed for heparin sodium USP have not been
verified to ensure suitability under actual conditions of use.
Our inspection found (Observation #4 of the FDA-483) that you have not
ensured that certain USP compendial test methods were verified
under actual conditions of use. Specifically, you have failed to conduct
adequate verification of USP compendial test methods as applied to
the production of your firm's API. The data you provided in your March
17, 2008, response did not include information about the suitability,
accuracy, and detection limits of certain test methods for API, such as
the protein test method, used by your firm. There was no indication from
these data that your firm's test methods could reliably detect and
quantify the presence of proteins in the finished API. In addition, your
firm had not conducted suitability testing of the method to determine
the limit of detection for the method. The suitability for use of the protein
method for in-process testing was also not established ... Please provide
data that demonstrate that the compendial test method has been verified
and determined to be suitable under actual conditions of use.
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Effective Test Method Verification
Documentation
Perform according to a written, approved
Protocol
† Title/Document
Number
† Introduction/Background
† Purpose
† Definitions
† Roles/Responsibilities
† Reference Documents
† Equipment, Reagents, Materials
† Procedure Steps
† Additional special instructions where needed
Effective Test Method Verification
Documentation
Protocol should include applicable analytical
characteristics (accuracy, precision, LOD, etc.)
and acceptance criteria for each characteristic
Explanation of deviations, protocol exceptions,
and/or out-of-specification results
Approval signatures
† Relevant
department managers
† Quality
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Handling a Failed Test Method
Verification
Troubleshoot/Seek assistance from USP staff
Procedure may not be suitable for use with the
particular article being tested
May be necessary to develop and validate an
alternate procedure
† Alternate
procedure is submitted to USP with
relevant data to support a proposal for inclusion
or replacement of the current compendial
procedure
When/If to reverify?
Depends on the nature of changes made, but
may be necessary in the case of
† Changes
in the synthesis of the product
† Changes in the composition of the product
† Changes in the analytical procedure
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Life Cycle Approach to
Validation
Change/
Develop
Review
Validation/
Verification
Analysis
Group Exercise
Divide into small groups
Select an analytical test method and develop
an outline for a verification protocol
† Determine
which category it is
† Briefly describe how you would test each attribute
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Final Thoughts / Questions?
aalderson@cookmyosite.com
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