Statistical Viewpoint.

David LeBlond, Daniel Griffith, and Kelly Aubuchon

Linear Regression 102: Stability

Shelf Life Estimation Using
Analysis of Covariance
David LeBlond, Daniel Griffith, and Kelly Aubuchon
Statistical Viewpoint addresses principles of statistics useful to practitioners in compliance and validation. We intend to present these concepts in a meaningful way so as to enable their application in daily
work situations.
Reader comments, questions, and suggestions are
needed to help us fulfill our objective for this column. Please contact managing editor Susan Haigney
at shaigney@advanstar.com with comments, suggestions, or manuscripts for publication.

KEY POINTS
The following key points are discussed:
Analysis of covariance (ANCOVA) is an important
kind of multiple regression that involves two predictor variables: one continuous (e.g., time) and one
categorical (e.g., batch of material).
Like simple linear regression, simple ANCOVA
fits straight lines to response measurements (e.g.,
potency, related substance, or moisture content)
over time: one line for each level (i.e., batch) of the
categorical variable.
A key objective of ANCOVA is to determine whether
the straight lines for all batches are best described as
having a common-intercept-common-slope (CICS)
model, a separate-intercepts-common-slope (SICS)
model, or a separate-intercepts-separate-slopes (SISS)
model.
In ANCOVA, model choice is based on two statistical

For more Author

information,
go to
gxpandjvt.com/bios

F-tests: one comparing slopes and one comparing

intercepts among batches. In the case of pharmaceutical shelf life estimation, the US Food and Drug
Administration recommends a p-value < 0.25 for
significance in these tests.
ANCOVA model adequacy can be assessed by examining measures such as a root mean square error
(RMSE), lack of fit, PRESS, and predicted R-square.
Once the appropriate model (i.e., CICS, SICS, or
SISS) has been identified for a given data set, it can
be used to obtain expected values, confidence intervals, and prediction intervals of potency of a given
lot at a given time.
When a lower or an upper specification limit can
be identified for the response, the ANCOVA model
can be used to estimate the shelf life for the batches
tested.
The shelf life for a pharmaceutical batch is defined as
the maximum storage period within which the 95%
confidence interval for the batch mean response level
remains within the specification range. Depending
on the response, the confidence interval may be one
or two sided.
The shelf life for a pharmaceutical product is taken
to be the minimum shelf life for batches on stability.
ANCOVA analysis and shelf life estimation using the
Minitab Stability Studies Macro is illustrated in the
cases of pharmaceutical potency, related substance,
and moisture content responses.

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ABOUT THE AUTHORS

David LeBlond, Ph.D., is senior statistician in Exploratory Statistics, Global Pharmaceutical R&D,
Abbott Global Pharmaceutical, Abbott Park, IL. He may be contacted by e-mail at david.leblond@abbott.com. Daniel Griffith is a statistician in the Technical Support Department at Minitab Inc. Kelly
Aubuchon is a statistician in the Technical Support Department at Minitab Inc., State College, PA.

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INTRODUCTION
A previous installment of Statistical Viewpoint
described simple linear regression in which there
is a single continuous independent variable such as
time, temperature, concentration, or weight (1). Many
important relationships involve multiple independent variables, some of which may be categorical in
nature (e.g., batch of material, supplier, manufacturing
site, laboratory, preservative type, clinical subject).
Understanding such relationships requires the use of
multiple linear regression. In this installment, we deal
with the simplest kind of multiple linear regression
in which there are two independent variables: one
continuous (called the covariate) and one categorical. The following are some examples in which this
kind of relationship is important:
Pre-clinical studies. Ten xenograft rodents are
treated with a range of doses of an anti-tumor
agent and the tumor weight for each animal
decreases as dose increases. The objective is to
quantify the animal to animal differences in dose
response profile. Here tumor weight is the dependent variable, rodent identity is the categorical
variable, and dose is the covariate.
Process scale-up. Active pharmaceutical ingredient (API) concentration is measured over time
in three chemical reactors. The reactors differ
in size (scale). The objective is to estimate scale
effects on the rate of API synthesis. Here, API
concentration is the dependent variable, scale
is the categorical variable, and dose level is the
covariate.
A nalytical methods. An assay measures the
concentration of an analyte in plasma samples
based on a florescence response. Samples are
tested in duplicate. Each test provides a blank
response and a test response. The objective is to
compare analyte concentrations among samples,
while correcting each for the effect of the blank. In
this case, the test response is the dependent variable, sample identity is the categorical response,
and blank is the covariate.
Pharmaceutical product stability. The drug
potency, related substance (a degradation product), and moisture level are measured over time
in multiple batches of product stored in a temperature- and humidity-controlled chamber. The
objective is to estimate the shelf life of the product. Here, the potency, related substance, and
moisture levels are the dependent variables, batch
identity is the categorical variable, and storage
time is the covariate.
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Notice the following distinctions in these examples.

The relative importance of the covariate and categorical variable differs. Sometimes, as in the pharmaceutical stability example, the primary interest may be
on the effects of the covariate (i.e., stability over time)
where the categorical variable, batch, is merely an
unavoidable nuisance variable. In other cases, as in the
analytical methods example, differences among levels
of the categorical variable, sample, are of primary
interest, while the effects of the covariate, blank, is an
unavoidable nuisance variable. In other cases, as with
the pre-clinical studies or process scale-up examples,
both the differences between the categorical variable
(rodent or scale) and the effects of the covariate (dose
or reaction time) may be of equal interest.
The covariate may or may not be truly independent. Sometimes the covariate may be a truly independent variable whose value is well controlled and
known with certainty, such as dose level or time.
In other cases, the covariate is actually a measured
value, such as an analytical blank. This violates one
of the assumptions of regression, that the predictor
variables are known without error (1). We still often
use regression in these cases as long as the covariate
is measured relatively accurately.
The experiment may include all or only some of the
categorical variable levels of interest. Sometimes we
include all levels of a categorical variable that are of
interest, such as with the analytical methods example
where we are concerned only with the samples being
tested. In other cases, the categorical variable levels in our experiment are merely a sampling of all
possible levels drawn from a larger population, such
as all possible rodents or all possible manufactured
batches. In these later cases, we must remember that
the methods we discuss here do not allow us to make
strong inferences about that larger population; our
conclusions will be limited primarily to the categorical
levels (e.g., rodents, batches) we have tested. To make
stronger inferences about the larger population, more
advanced statistical methods are required.
This article focuses on the important example of
pharmaceutical product stability. Thus our categorical variable will be batch and our covariate will be
storage time. Design and analysis of stability studies
is a mature discipline and such studies may include
additional continuous covariates such as dosage
strength, storage temperature, or excipient levels as
well as additional categorical variables such as excipient lot or packaging type. These more complex studies
are referred to as multi-factor stability studies (2).
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David LeBlond, Daniel Griffith, and Kelly Aubuchon.

Figure 1: Multiple-batch models of instability: CICS (common intercept and common slope),
SICS (separate intercept and common slope), SISS (separate intercept and separate slope).

The analysis of such studies is beyond the scope of

this article.
Because batches often differ in stability, stability
studies on a single batch of product are not of interest.
The number of batches in such studies is often small,
yet the objective is inevitably a shelf-life estimate to be
applied to the population of all future manufactured
batches. This is somewhat troubling. In light of the
last distinction mentioned above, we advise caution
and encourage the reader to discuss with a statistician the possibility of using mixed model or Bayesian
approaches (2) where appropriate. We will proceed
with our description of the traditional approach without apologies because it is common industry practice.

MODELS OF INSTABILITY
We will assume here that, for a given batch, the change
in level over time can be approximated by a straight
line. Chemists refer to this as pseudo zero-order kinetic mechanism. The real kinetic mechanism is almost
certainly more complex, but this linear assumption
is often found to be adequate. In any real application
this linear assumption should be justified. In some
cases, the response measurements or the time scale
can be altered using appropriate transformation(s) to
obtain a linear stability profile.
Consider the case where stability data are available for three batches of product. Figure 1 illustrates
possible models, or scenarios, of product instability
where the response is, for instance, the level of some
related substance or degradation product of the active
drug. However, the models described in Figure 1 apply
equally well for decreasing responses (e.g., potency)
or for responses that may rise or fall over time (e.g.,
moisture). In Figure 1, the mean response level for
each batch is indicated by a different colored line.
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Each line can be defined by its intercept (i.e., response

level at time zero) and slope (i.e., rate of change in
response over time).
The common intercept and common slope (CICS)
model represents a scenario were the stability profiles
of all batches have a common intercept and common
slope. This might be the result of a well controlled
manufacturing process where the initial levels of all
components, as well as their stability over time, are
uniform across batches. The CICS model generally
will result in a longer estimated product shelf life
because it allows tighter estimates of the mean slope
and intercept that are common to all batches.
The separate intercept and common slope (SICS)
model represents a scenario where batches have separate intercepts but a common slope. This could result
from a manufacturing process in which the initial
level of the component of interest is not well controlled batch to batch. However, other aspects of the
process that govern batch stability are uniform such
that the rate of change in the level of the component
of interest is the same for all batches.
The separate intercept and separate slope (SISS)
model represents a scenario where batches have separate intercepts and separate slopes. This could result
from an uncontrolled manufacturing process in which
neither the initial level nor the stability of the component of interest is well controlled batch to batch.
Clearly the CICS model is most desirable. The SICS
model may be acceptable as long as the initial level
non-uniformity is controlled within acceptance limits.
However, the SISS model is the least desirable scenario
because batches may become increasingly less uniform over time. The presence of large batch-to-batch
variability makes it difficult to accurately estimate
a shelf life for the process from only a few batches.
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Table I: Model comparisons made in the ANCOVA F-tests.

ANCOVA F-test

Simple model

More complicated model

Test for common slopes

SICS

SISS

Test for common intercepts

CICS

SICS

Some readers may notice that a CISS (common

intercept and separate slopes) model is missing from
Figure 1. Certainly there is no scientific reason to
exclude a manufacturing process in which initial levels of batches are very well controlled but that other
components (such as stabilizers) or process settings
that affect batch stability might not be well controlled.
However, while the initial levels may be relatively well
controlled they are unlikely to be identical, at least for
batches derived from blended powders or unit-dose
filling processes. So, unless there are compelling scientific reasons to consider the CISS model, we must
use the stability data to choose either the SISS, SICS,
or CICS models.
A model that is important in building the analysis
of covariance (ANCOVA) table but is not considered in
the evaluation of stability data is what we might call
the common intercept, no slope (CINS) model. The
CINS model assumes that the common slope of all
batches is zero. This implies a perfectly stable product.
While very stable pharmaceutical products do exist,
we never make an assumption of perfect stability in
evaluating stability data.

ANCOVA MODEL SELECTION

Well controlled processes that follow a CICS model
will more likely result in a longer shelf-life estimate
than those that follow the SICS or SISS models.
Because the estimate of shelf life depends on the
model choice, the first task is to choose the model.
While there may be development experience or theoretical reasons to expect one model over another, the
traditional approach is to let the stability data themselves guide us to the most appropriate model. The
ANCOVA is the statistical procedure for selecting the
most appropriate of the three models. ANCOVA is a
close cousin of the analysis of variance (ANOVA) associated with simple linear regression (1). Like ANOVA,
ANCOVA partitions the variance in the observed measurement in a specific way. This partitioning allows
us to make two statistical F-tests for batch differences
among slopes and intercepts.
The algebra behind the ANCOVA F-tests is complicated. But it is not necessary to understand the
algebra because the calculations are easily handled by
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statistical software packages such as Minitab Statistical

Software (3). However, it is necessary to understand
what these F-tests are comparing, what the criteria for
test acceptance or rejection are, and to be familiar with
the ANCOVA table that statistical software produces.
The ANCOVA F-tests make a comparison between
two models: a simple (null or reduced) model and
a more complicated (alternative or full) model. The
p-value associated with the test F statistic is used
to decide whether the portion of response variance
attributable to the extra features of the more complicated model is larger than can be explained by measurement variation alone. If so, we reject the simpler
model in favor of the more complicated one. Table I
shows the models being compared in the ANCOVA
F-tests.
The p-value obtained from either ANCOVA test in
Table I is the probability of obtaining an F statistic
that is as or more extreme than the one we observed,
given that the null hypothesis (i.e., the simpler model)
is true. If the p-value is below some fixed value, we
should select the more complicated model; otherwise we choose the simpler model. This fixed value
is referred to as the alpha or type I error level. In
many applications, we choose a limit value of 0.05 for
our hypothesis tests. However, in the case of pharmaceutical product stability, it is traditional to use the
more conservative limit of 0.25 for the p-value (4).
The 0.25 limit is controversial because it implies that
25% of the time we will incorrectly choose the more
complicated (and less desirable) model.
The rational for choosing this more conservative
limit has to do with the safety and efficacy. If we
incorrectly choose the more complicated model, the
estimated shelf-life estimate will likely be too short.
The consumers of this drug product will likely not
suffer side effects if a manufacturer establishes a shelf
life that is shorter than necessary. On the other hand,
if we incorrectly choose the simpler model, the estimated shelf life estimate will likely be too long. In that
case, consumers that use product near the end of its
shelf-life may be under medicated (if potency declines
with time) or be exposed to higher levels of harmful
degradation products. Consequently, regulatory agencies have established the more conservative p-value
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David LeBlond, Daniel Griffith, and Kelly Aubuchon.

limit of 0.25 to reduce the likelihood of establishing a

shelf life that is too long. This practice seems undesirable
from a manufacturers point of view, but remember that
the shelf life is meant to apply to the population of all
future batches. Establishing a shelf life that a process
cannot support adds to the cost of operations due to
out-of-specification investigations of batches on stability
and potential product recalls.
The ANCOVA decision process is diagrammed in Figure 2. It starts with the stability data at the top. There
are of course many ways to organize stability data. The
format shown in Figure 2 is what is required for input into
most statistical packages, such as Minitab, for ANCOVA
analysis. In this format, there are three columns: the
response level column, the time (covariate) column, and
the batch (categorical variable) column. For brevity, only
the first four and last observations are shown.
We start with the worst-case presumption that slopes
and intercepts vary among batches. The F-test for separate
slopes is examined first. If this test is statistically significant (i.e., p-value < 0.25) then the ANCOVA process
concludes with the selection of SISS as the final stability
model. As discussed previously, unless there is a compelling scientific argument, an F-test comparing the SISS
and CISS models is not made at this point.
If the F-test for separate slopes is not statistically significant (i.e., p-value 0.25), there is no evidence in the
data for a difference in slopes among the batches, and
we can presume a common slope model, SICS. Next, we
perform the second F-test in Table I that tests for separate
batch intercepts, assuming that batch slopes are common. This test is a comparison of model SICS and CICS.
If the test is statistically significant, then the ANCOVA
concludes with the selection of SICS model. If the test is
not statistically significant, then the remaining model,
CICS, is selected.
An ANCOVA table that is produced by the Minitab
stability macro is shown in Table II. It consists of five rows
and five columns of statistical quantities. The quantities
in each column, how they are obtained, and what they
represent are described as follows.

Source. A label indicates the variable or interaction

that contributes variation to the measurement. This label
also indicates the particular F-test that this row represents. The Time source provides an F test that tests the
hypothesis that the common slope is zero in the CICS
model. A low p-value suggests that some instability is
present, but is of no interest to us in model selection
here because we never entertain a model with zero slope.
The Error source does not include an F-test but provides
an estimate of total analytical variance (the quantity
mean squre error [MSE]), assuming that the SISS model
is appropriate. The degrees of freedom (DF) and the Seq
SS in the Total source row are merely the sum of those
quantities in the rows above.
The Batch and Batch*Time sources provide the ANCOVA F-tests for intercept and slope, respectively, that are
of interest to us here. The p-values from these F-tests are
used to make the model choice as described in Table I
and Figure 2.
Figure 2: The ANCOVA model selection process.

Table II: ANCOVA table output from the Minitab stability macro.
Source

DF

Seq SS

Seq MS

Time

DF T=1

SST=SSECINS-SSECICS

MST=SST/DF T

F T=MST/MSE

p-valueT

Batch

DFB=B-1

SSB=SSECICS-SSESICS

MSB=SSB/DFB

FB=MSB/MSE

p-valueB

Batch*Time

DFBT=B-1

SSBT=SSESICS-SSESISS

MSBT=SSBT/DFBT

FBT=MSBT/MSE

p-valueBT

Error

DFE=N-2*B

SSE=SSESISS

MSE=SSE/DFE

Total

DFtot=N-1

SStot=SSECINS

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DF. This gives the degrees of freedom associated

with each source. This is a measure of the amount
of information available in the data to estimate the
statistics associated with this source. B is the number
of batches in the data set, and N is the total number
of independent measurements in the data set. Notice
how the DF for the Total source equals the sum of the
values above it.
Seq SS. This is the sum of squares associated with
this source. Larger Seq SS values represent sources that
contribute more to variation in the data. This quantity
is obtained from the ANOVA error sum of squares (see
Reference 1) from the multiple regression fit to models
CINS, CICS, SICS, and SISS. The error sum of squares
is indicated as SSEmodel where the subscript gives the
fitted model. Notice how the Seq SS for the Total source
equals the sum of the values above it.
Seq MS. Seq MS gives the mean square (or variance)
associated with the source. This is simply the respective
Seq SS divided by the DF.
F. This gives the F-value for this source that is simply a ratio of the respective SS MS to some measure of
error variance. In the case of pharmaceutical stability
ANCOVA, it is common to use MSE as the error variance for all F-tests, but in a traditional ANCOVA table,
the quantity SSESICS/(N-B-1) is used as the measure of
error variance for the test for common intercept (the
Batch source). MSE is used because it is smaller than
the traditional quantity. This leads to a larger F-value,
which is more likely to lead to statistical significance
and a more conservative final model choice.
P. This gives the p-value for the F-test associated
with this Source. This p-value is the complement of
the cumulative F-distribution with quantile = FSource,
numerator degrees of freedom = DFSource, and denominator degrees of freedom = DFE.
To summarize, p-valueB and p-valueBT in Table II are
calculated from the stability data and are used to test for
common intercept and slope, respectively, as described
in Table I and Figure 2. The outcome of the ANCOVA
process is a final stability model that is used to estimate
the product shelf life. It is important to remember that
the model selected through the ANCOVA process may
change if data are re-analyzed after additional stability
time points are acquired.

DETERMINATION OF SHELF LIFE

Shelf life for a pharmaceutical product is based on
measurements of one or more stability indicating
responses for which upper or lower acceptance limits
have been established. The responses are measured
on a few (typically three) batches of product that are
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stored under carefully controlled temperature and

humidity in the intended packaging. Traditionally, a
pharmaceutical product shelf life for a batch is based
on the 95% confidence limit for the mean response
level over time, as estimated from the available stability batch data. The 95% confidence limit for a mean
regression line is described briefly in Reference 1. The
shelf life (S) is based on the shortest Time at which the
estimated 95% confidence bound crosses an acceptance limit. Shelf-life estimation for a single batch in
three common situations is illustrated in Figure 3.
The left panel of Figure 3 illustrates an increasing response level over time (such as a degradation
product) for which only an upper acceptance limit
is set. In this case, it is common to use a one-sided
upper confidence bound. The middle panel illustrates
a decreasing response level (such as tablet potency)
with only a lower acceptance limit set. In this case,
it is common to use a one-sided lower confidence
bound. The right panel illustrates the situation for
a response level (such as moisture) that may either
increase or decrease on storage and for which both
upper and lower limits have been set. Cases do
exist where lower (or upper) limits are in place for
responses expected to increase (or decrease) over
time. In such cases, it may be desirable to employ
two-sided limits. One-sided confidence limits will
lead to longer shelf-life estimates so their use must
be risk justified.
Usually, multiple response data from multiple batches are used to set shelf life. ANCOVA is
employed to identify the appropriate stability model
for the batches at hand. Regression procedures (1)
based on the selected ANCOVA model are used
to obtain 95% confidence bounds for each batch.
Assignment of shelf life for a product is based on
worst-case: the response, batch, and side (upper
or lower) giving the shortest shelf life is used to set
the shelf life for the product. Shelf life estimates are
often based on extrapolation beyond storage periods
of available stability batches. International Conference on Harmonisation (ICH) Q1E guidelines state
that the maximum allowable shelf life is two times
the maximum storage period of available stability
data (4).
Because our focus here is on the ANCOVA decision
process, we will emphasize this aspect of the computer
output in the examples below. The shelf life estimation
process involves simple or multiple regression and
results in additional tables of computer output. This
consists of prediction equations and stability profile
graphs for all batches (CICS model) or for each batch
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Figure 3: Illustration of shelf-life determination for a single batch. Red horizontal lines indicate
upper (U) or lower (L) acceptance limits. The solid straight line is the mean regression line, and the
dashed line is the upper or lower confidence interval. The maximum batch shelf life is indicated by S.

(SICS or SISS models), model summary statistics, and

an ANOVA that may include a lack-of-fit (LOF) test of
nonlinearity in the stability profile. This output will
be illustrated in the examples that follow. Interested
readers can learn more details about multiple regression from standard statistical textbooks (5).

ANCOVA DATA ANALYSIS USING THE

MINITAB STABILITY MACRO
ANCOVA and regression analysis for shelf-life estimation can be obtained using many commercially available
statistical packages. We illustrate this process here using
a convenient Minitab macro that may be downloaded
and saved (3). Once saved, an analysis is made as follows:
1. Start Minitab.
2. Enter stability data into a worksheet using the format
given in Figure 2.
3. Select Edit, then Command Line Editor.
4. Type a short script into the Command Line Editor
(syntax described below) that describes the type of
analysis desired.
5. Choose Submit Commands to execute the script.
The script will invoke the macro, and the ANOVA
and regression results, including stability profile
graphs and shelf-life estimates, will be produced. A
typical stability macro script is given as follows:

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%stability ycol tcol bcol;

store out.1-out.n;
itype it;
confidence cl;
life c.1 c.z;
xvalues xpredt xpredb;
nograph;
criteria alpha.

The script syntax consists of a main command

(%stability), given in the first line, and a set of
optional subcommands, each given on subsequent
lines. The order of appearance of the subcommands
is not important. All commands and subcommands
must end in a semicolon except the last subcommand, which must end in a period. Each command
and subcommand consists of a key word followed by
user-specified input parameters whose values tell the
macro what worksheet columns to use for data and
calculated predictions, and the kind of confidence
interval to employ. In the %stability command, ycol
indicates the column in your worksheet containing
your response (e.g., potency, related substance, or
moisture level,), tcol indicates column for storage
time, and bcol indicates the particular batch. The bcol
worksheet column can be formatted as either numeric
(i.e., 1, 2, 3,) or text (i.e., A, B, C, ). The macro
has a limit of up to 100 batches in the worksheet.
The other subcommands are explained in Table III.
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STABILITY ANALYSIS
The following illustrates five stability analyses using
this macro. The potency data used was obtained
from an actual literature example (6). The related
substance and moisture data are realistic, but artificially constructed.

Example One: Potency Stability

(CICS Model, One- or Two-Sided Limit)
Table IV provides potency stability data (%LC)
obtained over a 24-month period from B=3 batches
(batches numbered 2, 5, and 7) of a drug product. A
total of N=31 independent measurements are available. The first three columns of this table are in the
format required by the Minitab macro. Notice that
independent replicate measurements on each batch
are available for months 3-24. Such independent
replicates provide a test of the linearity assumption
as described below. Note also that we are assuming
independence of each measurement here (as discussed
in Reference 1), but independence is a key assumption

that must be justified. The lower acceptance limit for

potency for this product is 95% LC.
We can use the following script to analyze these
data and obtain an estimate for the product shelf life:
%stability c1 c2 c3;
store c4 c5 c6;
itype -1;
confidence 0.95;
life 95;
criteria 0.25.

Table V provides the ANCOVA and other computer

output. Compare the ANCOVA output in Table V to that
shown in Table II and to the ANCOVA decision process
shown in Figure 2. The p-value associated with the test
for separate slopes (Source = Batch*Time) is 0.797 which
is > 0.25, so the data provide no evidence for separate
slopes among the batches. The p-value associated with
the test for separate intercepts (Source = Batch) is 0.651,
which is > 0.25, so the data provide no evidence for sepa-

Table III: Stability macro subcommands.

Subcommand

Input Parameters

Definition

LIFE

c.1 c.z

Required in order to obtain shelf-life estimation. Specifies the acceptance limit(s) of

your response as constants. If you have only an upper or lower spec limit, indicate this
using only c.1. Use both c.1 and c.2 for two-sided limits.

STORE

out.1-out.n

Specifies storage columns for the fitted values and confidence/prediction limits for
each row of data. Either 3 or 5 columns for one- or two-sided limits, respectively.
These may be separated by spaces (c4 c5 c6 ) or given as a range using a dash
(c4-c6). When using the xvalues subcommand, fits and limits are provided only for the
batches/ times in the columns specified in the xvalues subcommand, and not the fits
and limits for every value in the dataset.

ITYPE

it

Defines the type of confidence limit.

It = 1 for an upper confidence bound
It = 0 for a two-sided confidence interval
It = -1 for a lower confidence bound
If ITYPE is not used, the LIFE subcommand parameters are used to select it. If both
c.1 and c.z are specified, it is set to 0, if only c.1 is given, it is set to -1..ITYPE must be
used if an upper bound is desired.

CONFIDENCE

cl

Cl is the confidence level used to estimate confidence/prediction intervals. By default,

the cl = 0.95. The type of interval depends on it:
it=0 produces a two-sided 100*cl% confidence central interval
it= -1 (or 1) produces a single-sided lower (or upper) bound.

XVALUES

xpredt xpredb

Requests fitted values and limits for batch/time combinations that were not included in
your stability data set. The desired times and batchs are entered into columns xpredt
and xpredb, respectively, prior to invoking the macro. The xvalues subcommand always needs to be used in conjunction with the store subcommand.

NOGRAPH

N/A

Suppresses the output of graphs.

CRITERIA

alpha

Defines the significance level used in the ANCOVA F-tests. By default, the significance
level is 0.25.

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rate intercepts among the batches. Consequently, we take

CICS as an appropriate stability model for estimating shelf
life. As seen in Table V, the Minitab macro output refers
to the CICS model as Model 1.
The output in Table V provides the regression equation with common intercept (100.567 %LC) and slope
(-0.192994 %LC/month). The negative slope indicates
that potency is decreasing with time. The output includes
the following summary statistics.
S. Root mean square estimate of the final model 1 fit.
This estimates total analytical standard deviation.
PRESS. Prediction sum-of-squares (PRESS). This gives
a robust estimate of your models predictive error. In general, the smaller the PRESS value, the better the models
predictive ability.
R-Sq(pred). A robust version of Adjusted R-Sq useful for comparing models because it is calculated using
observations not included in model estimation. Predicted
R-Sq ranges between 0 and 100%. Larger values of predicted R-Sq suggest models of greater predictive ability.
R-Sq(adj). A robust version of R-Sq, the percentage
of response variation that is explained by the model,
adjusted for the complexity of the model.
The output in Table V also includes a ANOVA table.
This ANOVA table is similar to that described previously
(1), but has a few additional statistical tests. Interested
readers are referred to standard statistical text books for
more information on complex ANOVA (5). One useful
feature of the ANOVA in Table V is the LOF test. Simply
put, this LOF test compares a models residual variance
to that available from pure replication to form an F ratio.
If this ratio is large and the p-value is significant (i.e., <
0.05), either there is evidence for non-linearity, or the
replicates are not truly independent. Such is the case in
this example (p-value = 0.0000037). If it is determined
that this nonlinearity is impacting the shelf-life estimation, it may be advisable to alter the model, transform
the response, or analyze replicate averages rather than
individual replicates. We will assume in this example
that the LOF has no impact and, for illustration, will use
this model to estimate shelf life.
The shelf-life estimate for this example is given at the
bottom of Table V as 26 months. This estimate is illustrated in Figure 4. This plot shows the individual measurements for each batch as separate colors. The solid black
line is the best-fit regression line for the mean potency of
all three batches. The red dashed line gives the one-sided
lower 95% confidence bound of the mean potency. It
can be seen that this line intersects the lower acceptance
limit for the product (95% LC) at about 26 months. It is
common practice to round a shelf-life estimate down to
the nearest whole month.
gxpandjv t.com

Table IV: Example one potency stability data

and estimated fits and limits.
c1

c2

c3

c4

c5

c6

Potency

Month

Batch

Fit

Lower CL

Lower PL

101.0

100.567

100.215

99.1808

102.0

100.567

100.215

99.1808

101.3

100.567

100.215

99.1808

101.3

100.374

100.043

98.9928

101.4

100.374

100.043

98.9928

101.5

100.374

100.043

98.9928

100.8

100.181

99.869

98.8043

99.8

99.988

99.693

98.6152

100.2

99.988

99.693

98.6152

10

100.2

99.988

99.693

98.6152

11

99.2

99.988

99.693

98.6152

12

99.7

99.988

99.693

98.6152

13

99.8

99.988

99.693

98.6152

14

99.5

99.409

99.154

98.0442

15

98.8

99.409

99.154

98.0442

16

99.0

99.409

99.154

98.0442

17

97.8

99.409

99.154

98.0442

18

98.5

99.409

99.154

98.0442

19

98.5

99.409

99.154

98.0442

20

97.4

12

98.251

97.994

96.8857

21

98.0

12

98.251

97.994

96.8857

22

98.5

12

98.251

97.994

96.8857

23

97.2

12

98.251

97.994

96.8857

24

97.1

12

98.251

97.994

96.8857

25

97.4

12

98.251

97.994

96.8857

26

96.9

24

95.935

95.436

94.5045

27

96.6

24

95.935

95.436

94.5045

28

96.6

24

95.935

95.436

94.5045

29

96.0

24

95.935

95.436

94.5045

30

96.1

24

95.935

95.436

94.5045

31

96.4

24

95.935

95.436

94.5045

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Figure 4: Example one potency stability profile for all

batches based on a CICS model and a one-sided lower
acceptance limit.

Notice the additional numbers in columns c4-c6 of

Table IV. The stability macro will place these numbers in
the worksheet as a result of the store subcommand (see
the script above used for this analysis). The Fit and Lower
CL (columns c4 and c5) correspond to the black and red
dashed lines, respectively, in Figure IV. The Lower PL in
Table IV is the Lower 95% prediction limit for individual
observations. This limit is more conservative (lower)
than the 95% confidence for the mean (red line) and
reflects the scatter of individual values about the fitted
line (see Reference 1 for more description). Notice in Table
IV that this prediction limit is below the acceptance limit
at 24 months. Thus in this case, while a 26-month shelf
life for the product may be acceptable from a regulatory
point of view, a sponsor may want to consider the risk
of out-of-specification results for this product near the
end of shelf life.
So far we have assumed a one-sided lower limit of
95%LC. If the product had an upper limit of 105%LC

Table V: Example one ANCOVA, regression, ANOVA, and estimated shelf-life output from the Minitab stability macro.
ANCOVA
Source

DF

Time

1 80.359 80.359 117.167 0.000

Batch

2 0.598 0.299 0.436 0.651

Batch*Time 2

Seq SS

0.314

Seq MS

0.157

Error

25 17.146 0.686

Total

30 98.417

0.229

0.797

Seq MS

Model 1 Analysis
Regression Equation
y = 100.567 - 0.192994 time
Summary of Model
S = 0.789106

R-Sq = 81.65%

R-Sq(adj) = 81.02%

PRESS = 20.4369 R-Sq(pred) = 79.23%

Analysis of Variance
Source

DF

Seq SS

Adj SS

Regression 1

80.3588 80.3588 80.3588 129.051

0.0000000

Time

80.3588 80.3588 80.3588 129.051

0.0000000

Error

29 18.0580 18.0580 0.6227

Lack-of-Fit

13.1613

13.1613

2.6323

Pure Error

24

4.8967

4.8967

0.2040

Total

30 98.4168

12.901 0.0000037

Estimated shelf-life for all batches: 26

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as well and there is risk of batches exceeding the upper

limit, then we might want a shelf life based on a twosided 95% confidence interval. In that case we could use
the following analysis script:

Figure 5: Example one potency stability profile for all

batches based on a CICS model and a two-sided acceptance
limit.

%stability c1 c2 c3;
life 95 105.

The resulting stability profile is shown in Figure 5.

Notice in this case that the shelf-life estimate is slightly
lower (25.5 months which we would likely round down
to 25 months). This is because two-sided limits will be
wider than a one-sided bound and will thus intersect
the limit sooner.

Example Two: Potency Stability

(SICS Model Two, One-Sided Lower Limit)
Another set of potency stability data is given in columns
C1-C3 of Table VI. As before, we will assume a one-sided
lower acceptance limit of 95%LC.
We will use the following script to estimate the product
shelf life based on these data:
%stability c1 c2 c3;
store c4 c5 c6;
itype -1;
confidence 0.95;
life 95;
criteria 0.25.

The ANCOVA and other statistical output from this

analysis are given in Table VII.
There is no evidence for separate slopes (p-value
= 0.834). However, there is evidence for separate
intercepts (p-value < 0.001). A comparison with the
ANCOVA decision process of Figure 2 shows that the
SICS model is appropriate in this case. The regression
equations in Table VII show that the estimated slope
(-0.213121 %LC/month) is common to each batch, but
the intercepts differ. As in example one, the LOF test is
significant (p-value = 0.0258), but we will assume that
the straight-line assumption is adequate for illustration
purposes here.
Figure 6 provides the separate stability profiles for each
batch. Because the intercepts differ, the macro produces
a separate plot for each batch. The shelf life estimated
for each batch, based on when its 95% confidence lower
bound crosses the acceptance limit of 95%LC, is given
on the upper right corner of each plot. Batch 5 has the
lowest estimated shelf life (23.4 months). Therefore, by
the worst-case logic of pharmaceutical shelf-life estimation, limits the shelf life for the product to 23.4 months
gxpandjv t.com

as is also indicated in Table VII. In practice, we would

likely round this down to 23 months. As described in
Example one, columns C4-C6 of Table VI provide the
numeric Fit and interval estimates based on the store
subcommand request.

Example Three: Potency Stability

(SISS Model, One-Sided Lower Limit With
Predictions)
Yet another set of potency stability data is provided in
columns C1-C3 of Table VIII.
These data are analyzed using the following script:
%stability c1 c2 c3;
store c4 c5 c6;
itype -1;
confidence 0.95;
life 95;
criteria 0.25.

Table IX shows the ANCOVA and other statistical output from this analysis.
There is evidence for both separate slopes (p-value
= 0.17) and intercepts (p-value < 0.01). Both p-values
are below the regulatory limit of 0.25. A comparison with
the ANCOVA decision process of Figure 2, shows that the
SISS model is appropriate in this case. The regression equations for each batch are given in Table IX, and the slopes
and intercepts differ for each batch as expected. We note
that in this case, the LOF test is not statistically significant
(p-value = 0.100568). For this test we use the traditional
Type I error rate of 0.05 to judge statistical significance.
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Figure 6: Example two potency stability profiles for each batch on a SICS model and a onesided lower acceptance limit.

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Table VI: Example two potency stability data and estimated fits and limits.
C1

C2

C3

C4

C5

C6

Potency

Month

Batch

Fit

Lower CL

Lower PL

104.8

102.176

101.434

100.192

104.0

104.255

103.463

102.252

102.0

100.820

100.163

98.866

101.4

100.607

99.971

98.660

100.8

100.394

99.777

98.453

103.0

101.536

100.857

99.575

103.2

103.616

102.887

101.637

100.2

100.181

99.581

98.245

101.2

101.536

100.857

99.575

10

99.7

100.181

99.581

98.245

11

100.8

100.897

100.261

98.950

12

102.8

102.976

102.295

101.014

13

98.8

99.541

98.977

97.617

14

99.2

100.897

100.261

98.950

15

103.3

102.976

102.295

101.014

16

98.5

99.541

98.977

97.617

17

98.6

12

99.618

98.999

97.677

18

102.4

12

101.698

101.045

99.745

19

98.0

12

98.263

97.688

96.335

20

97.2

12

99.618

98.999

97.677

21

101.2

12

101.698

101.045

99.745

22

97.1

12

98.263

97.688

96.335

23

97.6

24

97.061

96.215

95.035

24

99.1

24

99.140

98.291

97.113

25

96.6

24

95.705

94.853

93.677

26

98.0

24

97.061

96.215

95.035

27

99.5

24

99.140

98.291

97.113

28

96.1

24

95.705

94.853

93.677

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Table VII: Example two ANCOVA, regression, ANOVA, and estimated shelf-life
output from the Minitab stability macro.
ANCOVA
Source

DF

Time

1 74.489 74.489 60.008 0.000

Batch

2 53.968 26.984 21.738 0.000

Batch*Time 2

Seq SS

0.455

Seq MS

0.227

Error

22 27.309 1.241

Total

27 156.221

0.183

0.834

Model 2 Analysis
Regression Equation
batch
3

y = 102.176 - 0.213121 time

y = 104.255 - 0.213121 time

y = 100.82 - 0.213121 time

Summary of Model
S = 1.07556

R-Sq = 82.23%

R-Sq(adj) = 80.01%

PRESS = 37.3301 R-Sq(pred) = 76.10%

Analysis of Variance
Source

DF

Seq SS

Adj SS

Seq MS

Regression

128.457

128.457

42.8191

37.0144 0.0000000

time

74.489

88.734

74.4895

64.3914

0.0000000

batch

53.968

53.968

26.9839

23.3259

0.0000024

3.3602

0.0258372

Error

24 27.764 27.764 1.1568

Lack-of-Fit

13

22.179

22.179

1.7061

Pure Error

11

5.585

5.585

0.5077

Total

27 156.221

Overall minimum estimated shelf-life: 23.4

Stability profiles for each batch are given in Figure 7.

As seen in Figure 7 and Table IX, the product shelf
life estimated by these data is limited by Batch 8 to
15.6 months. We would likely round this down to 15
months in practice. However, it would be interesting in
this case to see what potencies the model would predict
for these batches at 15 months. No real stability testing
was done at 15 months of storage, but we can use the
stability model to obtain estimates by including the
desired times and batch numbers in columns c4 and
c5, respectively, prior to the analysis and employing
the following script:
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%stability c1 c2 c3;
itype 0;
confidence 0.95;
life 95 105;
xvalues c4 c5;
store C6 c7 c8 c9 c10.

For illustration, we are requesting two-sided 95%

confidence limits (it=0). This amounts to requesting
a 97.5% confidence lower bound, which is more conservative than a 95% confidence lower bound. The
same result could be obtained using it= -1 and cl =
97.5. Columns C4 and C5 contain the time points and
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David LeBlond, Daniel Griffith, and Kelly Aubuchon.

batches for which we want predictions. The above macro

performs the fit as given previously in Table IX and the
xvalues subcommand produces the predictions in columns C6-C10 of Table X. Note that the lower confidence
bound is still within the limit of 95%LC, although the
lower prediction bound, which reflects individual result
variation, is below the acceptance limit.

Table VIII: Example three potency data and

estimated fits and limits.

Example Four: Related Substance

Stability (SISS Model Three, One-Sided
Upper Limit)
To illustrate estimation of shelf life for a response whose
level increases on storage, we will use the data for a related
substance (degradation product of the active ingredient)
given in columns C1-C3 of Table XI. The levels in column
C1 are expressed as a percent of label claim for the active
ingredient and the upper limit for this particular related
substance is assumed to be 0.3%LC.
We can obtain the shelf life based on this response by
using the following script:

C1

C2

C3

C4

C5

C6

Potency

Month

Batch

Fit

Lower CL

Lower PL

104.0

104.071

103.402

102.729

102.0

100.782

100.280

99.515

101.6

101.259

100.375

99.798

101.4

100.573

100.101

99.318

100.8

100.365

99.919

99.119

103.2

103.482

102.921

102.191

100.2

100.156

99.736

98.919

100.0

100.269

99.618

98.936

99.7

100.156

99.736

98.919

10

102.8

102.894

102.419

101.637

11

98.8

99.530

99.164

98.311

12

99.0

99.278

98.754

98.002

%stability c1 c2 c3;
store c4 c5 c6;
itype 1;
confidence 0.95;
life 0.3;
criteria 0.25.

13

103.3

102.894

102.419

101.637

14

98.5

99.530

99.164

98.311

15

102.4

12

101.717

101.302

100.482

16

98.0

12

98.279

97.897

97.054

17

97.8

12

97.297

96.514

95.895

Notice in this case that we are requesting a one-sided

upper confidence limit (it=1) of 95% (cl=0.95). The output
from this analysis is shown in Table XII.
As in example three, the ANCOVA output in Table XII
indicates an SISS model. The separate slopes and intercepts are given in Table XII along with an LOF test that
is not statistically significant, and an estimated shelf life
of 15.61 months (which we would usually round down
to 15 months).
Stability profiles for these batches are given in Figure 8,
which confirms that batch 8 is the stability limiting batch
for the product shelf life. Numeric predictions, requested
using the STORE subcommand are given in columns
C4-C6 of Table IX.

18

101.2

12

101.717

101.302

100.482

19

97.1

12

98.279

97.897

97.054

20

97.0

12

97.297

96.514

95.895

21

99.1

24

99.363

98.605

97.975

22

96.6

24

95.775

95.027

94.392

23

99.5

24

99.363

98.605

97.975

24

96.1

24

95.775

95.027

94.392

Example Five: Moisture Stability (CICS

Model, Two-Sided Limits)
As a final example of a response that may either increase or
decrease on storage, we examine the moisture data given
in columns C1-C3 of Table XIII. The moisture measurements in column C1 have units of %(w/w). We will take the
acceptance limits for this product to be 1.5 to 3.5 %(w/w).
We can analyze these data using the following script.
Notice that we have specified both the lower and upper
gxpandjv t.com

acceptance limits using the life subcommand and

requested two-sided confidence limits using the itype
subcommand.
%stability c1 c2 c3;
itype 0;
confidence 0.95;
life 1.5 3.5;
criteria 0.25.

The results of this analysis are provided in Table XIV.

Notice in this case, the ANCOVA analysis leads to the
CICS model because neither the test for separate slopes
nor the test for separate intercepts is statistically significant
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Table IX: Example three ANCOVA, regression, ANOVA, and estimated shelf-life
output from the Minitab stability macro.
ANCOVA
Source

DF

Seq SS

Time

45.451 45.451 100.994 0.00

Batch

2 64.918 32.459 72.124 0.00

Batch*Time 2

Seq MS

1.760

0.880

Error

18 8.101 0.450

Total

23 120.230

1.955

0.17

Model 3 Analysis
Regression Equation
batch
4

y = 104.071 - 0.196151 time

y = 100.782 - 0.208609 time

y = 101.259 - 0.330208 time

Summary of Model
S = 0.670850

R-Sq = 93.26%

R-Sq(adj) = 91.39%

PRESS = 13.5446 R-Sq(pred) = 88.73%

Analysis of Variance
Source

DF

Regression 5

Seq SS

Adj SS

Seq MS

112.129

112.129

22.4258 49.831 0.000000

time

45.451

45.950

45.4513

100.994

0.000000

batch

64.918

21.635

32.4588

72.124

0.000000

time*batch

1.760

1.760

0.8800

1.955

0.170420

2.532

0.100568

Error

18 8.101 8.101

0.4500

Lack-of-Fit

10

6.156

6.156

0.6156

Pure Error

1.945

1.945

0.2431

Total

23 120.230

Overall minimum estimated Shelf-Life: 15.6

Table X: Example three fit, confidence limit, and prediction limit estimates for
time and batch combinations not present in the stability data.

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Journal

C4

C5

C6

C7

C8

C9

C10

Xvalue_Month

Xvalue_Batch

Fit

Lower CL

Upper CL

Lower PL

Upper PL

15

101.128

100.574

101.683

99.6139

102.643

15

97.653

97.110

98.195

96.1426

99.163

15

96.306

95.036

97.577

94.4088

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Figure 7: Example three potency stability profiles each batch on a SISS model and a one-sided
lower acceptance limit.

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Table XI: Example four related substance stability data and estimated fits and limits.

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C1

C2

C3

C4

C5

C6

Related

Month

Batch

Fit

Upper CL

Upper PL

0.030

0.027881

0.047950

0.068139

0.054

0.045534

0.062376

0.084284

0.066

0.063188

0.077421

0.100878

0.051

0.063188

0.077421

0.100878

0.078

12

0.098495

0.110942

0.135547

0.114

12

0.098495

0.110942

0.135547

0.177

24

0.169110

0.191852

0.210765

0.165

24

0.169110

0.191852

0.210765

0.090

0.126544

0.141610

0.164556

10

0.108

0.132802

0.146984

0.170472

11

0.126

0.139060

0.152420

0.176429

12

0.144

0.145319

0.157933

0.182427

13

0.159

0.145319

0.157933

0.182427

14

0.186

0.164093

0.175072

0.200678

15

0.195

0.164093

0.175072

0.200678

16

0.210

12

0.201643

0.213096

0.238373

17

0.237

12

0.201643

0.213096

0.238373

18

0.252

24

0.276742

0.299188

0.318236

19

0.267

24

0.276742

0.299188

0.318236

20

0.102

0.112219

0.138754

0.156060

21

0.150

0.141938

0.161447

0.181919

22

0.180

0.171656

0.187385

0.209936

23

0.216

12

0.231094

0.254586

0.273163

24

0.240

12

0.231094

0.254586

0.273163

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Table XII: Example four ANCOVA, regression, ANOVA, and estimated shelf-life
output from the Minitab stability macro.
ANCOVA
Source

DF

Seq SS

Seq MS

Time

0.041

0.041

100.994 0.00

Batch

2 0.058 0.029 72.124 0.00

Batch*Time 2

0.002

0.001

Error

18 0.007 0.000

Total

23 0.108

1.955

0.17

Seq MS

Model 3 Analysis
Regression Equation
batch
4

y = 0.0278806 + 0.00588454 time

y = 0.126544 + 0.00625826 time

y = 0.112219 + 0.00990625 time

Summary of Model
S = 0.0201255

R-Sq = 93.26%

R-Sq(adj) = 91.39%

PRESS = 0.0121902 R-Sq(pred) = 88.73%

Analysis of Variance
Source

DF

Seq SS

Regression

0.100916 0.100916

0.0201832 49.831 0.000000

time

0.040906

0.041355

0.0409062

100.994

0.000000

batch

0.058426

0.019472

0.0292129

72.124

0.000000

time*batch

0.001584

0.001584

0.0007920

1.955

0.170420

18

0.007291 0.007291 0.0004050

Lack-of-Fit

10

0.005540

0.005540

0.0005540

2.532

0.100568

Pure Error

0.001751

0.001751

0.0002188

Error

Total

Adj SS

23 0.108207

Overall minimum estimated shelf-life: 15.61

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Statistical Viewpoint.

Figure 8: Example four related substance stability profiles

for each batch on a SISS model and a one-sided upper
acceptance limit.

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Table XIII: Example two moisture

stability data.
C1

C2

C3

Moisture

Month

Batch

2.20059

1.70372

3.32395

2.75907

2.43192

1.76331

1.56801

2.19423

12

3.22311

12

10

3.16325

24

11

1.54837

24

12

2.81078

13

1.94915

14

2.49058

15

2.00485

16

3.30700

17

2.99309

18

3.30159

19

2.72512

12

20

1.88341

12

21

2.77215

24

22

1.69048

24

23

2.45301

24

2.16138

25

2.26631

26

2.12853

27

2.51775

28

2.31034

29

3.36915

30

2.32070

12

31

2.72001

12

32

2.19393

24

33

3.45895

24

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David LeBlond, Daniel Griffith, and Kelly Aubuchon.

Table XIV: Example five ANCOVA, regression, ANOVA, and estimated shelf-life
output from the Minitab stability macro.
ANCOVA
Source

DF

Time

1 0.012 0.012 0.033 0.858

Batch

2 0.251 0.125 0.354 0.705

Batch*Time 2

Seq SS

0.531

Seq MS

0.265

Error

27 9.573 0.355

Total

32 10.366

0.748

0.483

Model 1 Analysis
Regression Equation
y = 2.45678 + 0.0022724 time
Summary of Model
S = 0.577939

R-Sq = 0.11%

R-Sq(adj) = -3.11%

PRESS = 12.0483 R-Sq(pred) = -16.23%

Analysis of Variance
Source

DF

Seq SS

Adj SS

Seq MS

Regression

0.0116

0.0116

0.011599

0.034726 0.853386

0.0116

0.0116

0.011599

0.034726

0.853386

0.589613

0.707875

time
Error

31 10.3544 10.3544 0.334013

Lack-of-Fit

1.0545

1.0545

0.210898

Pure Error

26

9.2999

9.2999

0.357689

Total

32 10.3660

Estimated shelf life for all batches: 45.35

(i.e., p-values of 0.483 and 0.705, respectively). The stability

profile given in Figure 9 indicates a shelf life for all batches
of 45.35 months, which agrees with the estimate at the
bottom of Table XIV. In this case, it is the 95% confidence
upper bound that crosses the upper limit earliest and that,
therefore, governs the product shelf life.

CONCLUSION
We have illustrated here the ANCOVA process that is used
to set product shelf life for pharmaceutical products. We
have also illustrated the use of a convenient Minitab macro
that can be used to perform the ANCOVA analysis, choose
the appropriate stability model, and execute the multiple
regressions to estimate shelf life and produce other useful
statistical tests and statistics. The macro is flexible enough
to handle a variety of common situations and produces
graphics that serve as useful regression diagnostics.
gxpandjv t.com

It is essential to stress here the critical aspect of software validation. Validation is a regulatory requirement
for any software used to estimate pharmaceutical product
shelf life. Reliance on any statistical software, whether
validated or not, carries with it the risk of producing
misleading results. It is incumbent on the users of statistical software to determine, not only that the statistical
packages they use can produce accurate results, given a
battery of standard data sets, but also that the statistical
model and other assumptions being made apply to the
particular data set being analyzed, and that data and command language integrity are maintained. It is not uncommon for a computer package to perform differently when
installed on different computing equipment, in different
environments, or when used under different operating
systems. In our hands, using a number of representative
data sets, the Minitab Stability macro performs admirably
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67

Statistical Viewpoint.

Figure 9: Example five moisture stability profile for all batches based on a CICS model and a
two-sided acceptance limit.

compared to other statistical packages such as JMP, SAS,

and R. However, we can make no general claim that it will
not be found lacking in other environments. Readers are
advised to enlist the aid of local statisticians to assure that
the statistical packages they use are properly validated.

REFERENCES
1. Hu Yanhui, Linear Regression 101, Journal of Validation
Technology 17(2), 15-22, 2011.
2. LeBlond D., Chapter 23, Statistical Design and Analysis of
Long-Term Stability Studies for Drug Products, In Qui Y, Chen
Y, Zhang G, Liu L, Porter W (Eds.), 539-561, 2009.
3. Minitab Stability Studies Macro (2011), A technical support document describing the use of the Macro in Minitab
version 16 is available from the Minitab Knowledgebase
at http://www.minitab.com/support/answers/answer.
aspx?id=2686.
4. International Conference on Harmonization. ICH Q1E,
Step 4: Evaluation for Stability Data, 2003. http://www.ich.
org/products/guidelines/quality/article/quality-guidelines.
html
5. Neter J, Kuntner MH, Nachtsheim CJ, and Wasserman W,
Applied Linear Statistical Models, Chapter 23. 3rd edition,
Irwin Chicago, 1996.
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6. Schuirmann, DJ, Current Statistical Approaches in the

Center for Drug Evaluation and Research, FDA, Proceedings of Stability Guidelines, AAPS and FDA Joint Conference,
Arlington, VA, Dec 11-12, 1989. JVT

ARTICLE ACRONYM LISTING

ANCOVA
ANOVA
API
CICS
CL
DF
LOF
%LC
MSE
PL
PRESS
RMSE
R-Sq
R-Sq(adj)
R-Sq(pred)
SICS
SISS

Analysis of Covariance
Analysis of Variance
Active Pharmaceutical Ingredient
Common Intercept and Common Slope
Confidence Limit
Degrees of Freedom
Lack of fit
Percent of Label Claim
Mean Square Error
Prediction Limit
Predicted Residual Sum of Squares
Root Mean Squared Error
R-square
Adjusted R-square
Prediction R-square
Separate Intercept and Common Slope
Separate Intercept and Separate Slope
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