Transcript STABILITY STUDIES

Supplementary Training Workshop on
Good Manufacturing Practices (GMP)
VALIDATION of
ANALYTICAL TEST
METHODS
János Pogány, pharmacist, PhD,
consultant to WHO
Pretoria, South Africa, 28 June 2005
E-mail: [email protected]
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GMP - 4.11 Analytical methods,
computers and cleaning procedures
 „It is of critical importance that particular attention is
paid to the validation of analytical test methods,
automated systems and cleaning procedures.”
 Validation of analytical procedures used in the
examination of pharmaceutical materials (WHO Expert
Committee on Specifications for Pharmaceutical Preparations.
32nd Report. Geneva, WHO, 1992 (WHO Technical Report
Series, No. 823).
 Text on Validation of Analytical Procedures Q2A
(1994) Validation of Analytical Procedures:
Methodology Q2B (1996) ICH Harmonized Tripartite
Guidelines
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Characteristics of methods
 Accuracy (also termed trueness)
 Precision

repeatability

intermediate precision (intra-laboratory variation)

reproducibility (inter-laboratory variation)
 Robustness, ruggedness
 Stability
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Accuracy and precision
Inaccurate &
imprecise
Inaccurate and imprecise
Inaccurate but
precise
Precise
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Accurate but
Accurate
imprecise
Accurate and precise
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Accuracy (of any process)
The accuracy of an analytical  application of an analytical
procedure to an analyte of known
procedure expresses the
purity (e.g. reference material);
closeness of agreement
 comparison of the results of the
between the value, which is
proposed analytical procedure
accepted either as a
with those of a second wellconventional true value or an
characterized procedure, the
accepted reference value and
accuracy of which is stated
the value found (individual
and/or defined
observation or mean of
 accuracy may be inferred once
measurements).
precision, linearity and
specificity have been established.
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Precision (of any process)
Measured mean
Real mean
The precision (VARIABILITY)
of an analytical procedure is
usually expressed as the standard
deviation (S), variance (S2), or
coefficient of variation (= relative
standard deviation, R.S.D.) of a
series of measurements.
The confidence interval should be
reported for each type of
precision investigated.
PRECISION
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Repeatability (of any process)
Measured mean
Repeatability expresses the
precision (spread of the data,
variability) under the same
operating conditions over a
short interval of time.
Repeatability is also termed
intra-assay precision.
REPEATABILITY
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Intermediate Precision and
Reproducibility
Measured means
Intermediate precision
or Reproducibility
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Intermediate precision expresses
within-laboratories variations. #1,
#2 and #3: different days, different
analysts, different (manufacturing)
equipment, etc.
Reproducibility expresses the
precision between laboratories #1,
#2 and #3 (collaborative studies,
usually applied to standardization
of methodology). (Transfer of
technology)
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Sensitivity and robustness
Input-output relationship
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Stability
Stability Measured means
Stability (of the analytical
solution) expresses variation
of the measured mean as a
function of time.
(Manufacturing machines)
#1 … First mesurements
#2 … Second mesurements
of the same sample within a
relatively short period of
time.
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Characteristics of analytical
procedures
 Linearity
 Range
 Specificity (selectivity)
 Sensitivity (versus robustness)
 Limit of detection
 Limit of quantitation
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Linearity
Measured
mean
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Real
mean
Precision
Linearity expresses
differences in precision at
different points of a given
range.
„The linearity of an
analytical procedure is its
ability (within a given
range) to obtain test
results, which are directly
proportional to the
concentration (amount) of
analyte in the sample.”
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Range (minimum requirements)
 Assay of an API or a FPP: ± 20% of the test concentration.
 Content uniformity: ± 30% of the test concentration (unless a
wider more appropriate range, based on the nature of the dosage form
(e.g., metered dose inhalers), is justified).
 Dissolution testing: ± 20 % over the specified range.
 Impurity: from the reporting level of an impurity to 120% of
the specification. (Unusually potent or toxic impurities, LOD and LOQ
should be commensurate with ICH requirement.)
 If assay and purity are performed together as one test and only
a 100% standard is used, linearity should cover the range from
the reporting level of the impurities to 120% of the assay
specification
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Specificity (selectivity)
 Specificity is the ability to assess unequivocally
the analyte in the presence of components, which
may be expected to be present. Typically these
might include impurities, degradants, excipients,
etc.
 Stability indicating analytical methods should
always be specific.
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LOD, LOQ and SNR
 Limit of Quantitation (LOQ)
 Limit of Detection (LOD)
 Signal to Noise Ratio (SNR)
Peak B
LOQ
Peak A
LOD
Baseline
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Classes of analytical tests

„The objective of validation of an analytical procedure is to demonstrate that
it is suitable for its intented purpose.”
 Class A: To establish identity
 Class B: To detect (Bd) and quantitate (Bq) impurities
 Class C: To determine quantitatively the concentration,
or assay
 Class D: To assess characteristics
 Other classes not covered in the guides
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Criteria for analytical classes
Criteria
A
Bq
Bd
C
D
Accuracy
X
X
X
Precision
X
X
X
X
X
X
X
X
X
Robustness
X
Linearity and range
Specificity
X
X
X?
X
Limit of detection
X
X
Limit of quantitation
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X
X
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General requirements
 Qualified and calibrated instruments
 Documented methods
 Reliable reference standards
 Qualified analysts
 Sample integrity
 Change control (e.g., synthesis, FPP composition)
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HPLC Method Development and
Validation for Pharmaceutical
Analysis by Ghulam A. Shabir
Pharmaceutical Technology
Europe, 1 March 2004
Prequalification requirements
 Analytical method validation is required by WHO
for the prequalification of product dossiers. Noncompendial ARV APIs and FPPs were/are tested
with methods developed by the manufacturer.
 Analytical methods should be used within GMP
and GLP environments, and must be developed
using the protocols and acceptance criteria set out
in the ICH guidelines Q2A and Q2B.
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HPLC system
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Linearity and range
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ICH requirements
 Concentration range 0.025–0.15 mg/mL (25–150% of the
theoretical concentration in the test preparation, n=3)
 Regression equation was found by plotting the peak area
(y) versus the analyte concentration (x) expressed in
mg/mL: y = 3007.2x + 4250.1 (r2 = 1.000).
 The regression coefficient demonstrates the excellent
relationship between peak area and concentration of
analyte.
 The analyte response is linear across 80-120% of the
target progesterone concentration.
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Accuracy
The data show that the recovery of analyte in spiked samples met the
evaluation criterion for accuracy (100 ± 2.0% across 80–120% of target
concentrations).
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Specificity
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Specificity
 An example of specificity criterion for an assay
method is that the analyte peak will have baseline
chromatographic resolution of at least 2.0 minutes
from all other sample components.
 In this study, a weight of sample placebo
equivalent to the amount present in a sample
solution preparation was injected to demonstrate
the absence of interference with progesterone
elution. Former slide demonstrates specificity.
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Repeatability
The repeatability
precision obtained by
one analyst in one
laboratory was 1.25%
RSD for the analyte and,
therefore, meets the
evaluation criterion of
RSD ≤2%.
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Intermediate precision
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Limit of detection
 The limit of detection (LOD) is defined as the
lowest concentration of an analyte in a sample that
can be detected, not quantified. It is expressed as a
concentration at a specified signal:noise ratio
(SNR), usually between 3 and 2:1.
 In this study, the LOD was determined to be
10ng/mL with a signal:noise ratio of 2.9:1.
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Limit of quantitation
 The limit of quantitation (LOQ) is defined as the
lowest concentration of an analyte in a sample that
can be determined with acceptable precision and
accuracy under the stated operational conditions of
the method. The ICH has recommended a signal :
noise ratio (SNR) of 10:1.
 The LOQ was 20 ng/mL with a signal:noise ratio of
10.2. The RSD for six injections of the LOQ
solution was ≤2%.
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Analytical solution stability
Standard and sample solutions stored in a capped volumetric flask on
a lab bench under normal lighting conditions for 24 h were shown to
be stable with no significant change in progesterone concentration
during this period.
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Main Points Again
 Validation of analytical procedures is a critical
requirement in risk assessment and management:


establishment of product-specific acceptance criteria, and
stability of APIs and FPPs.
 Validation should demonstrate that the analytical
procedure is suitable for its intented purpose.
 HPLC systems and method validation deserves
special attention during the inspection of QC
laboratories.
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