Download BIOAVAILABILITY AND BIOEQUIVALENCE STUDIES

 DEFINITION
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Bioavailability is defined as the rate and the
absorption of drug that reaches the biological
system in an active form, capable of exerting
the desired pharmacological effect, including
its onset, intensity and duration of its action.
Objectives of bioavailability studies:
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Development of new formulation.
Determination of influence of excipients, patient
related factors and possible interaction with other
drugs on the efficiency of absorption.
Control of quality of a drug product during the early
stages of marketing in order to determine the
influence of processing factors, storage, stability on
drug absorption.
Primary stages of the development of a suitable
dosage form for a new drug entity.
• CONSIDERATIONS IN IN-VIVO BIOAVAILABILITY
STUDY DESIGN:
 Bioavailability—Absolute versus Relative
• ABSOLUTE BIOAVAILABILITY:
• The systemic availability of a drug administered orally is determined in
comparison to its iv administration.
• Characterization of a drug's absorption properties from the e.v. site.
• F = AUCev
AUCiv
• RELATIVE BIOAVAILABILITY:
• The availability of a drug product as compared to another dosage form or
product of the same drug given in the same dose.
• Characterization of absorption of a drug from its formulation.
• Fr=AUCA
AUCB
Route
Intravenous
(IV)
Bioavailability (%)
100 (by definition)
Intramuscular
(IM)
75 to ≤ 100
Subcutaneous
(SC)
75 to ≤ 100
Oral (PO)
5 to < 100
Characteristics
Most rapid onset
Large volumes often possible; may be
painful
Smaller volumes than IM; may be painful
Most convenient; first pass effects may be
significant
Rectal (PR)
30 to < 100
Less first-pass effects than oral
Inhalation
5 to < 100
Often very rapid onset
Transdermal
80 to ≤ 100
Usually very slow absorption; used for
lack of first-pass effects; prolonged duration
of action
 Single Dose versus Multiple Dose Studies
• The dose to be administered for a bioavailability study is determined from
preliminary clinical experiments.
• Single dose bioavailability studies are very common. They are easy, offer
less exposure to drugs and are less tedious.
Limitation: Difficult to predict the steady-state characteristics of a drug and
inter-subject variability.
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multiple dose study :
the bioavailability is determined at steady-state.
Better evaluation of a controlled-release formulation is possible.
Small inter-subject variability is observed,
Requires collection of fewer blood samples.
Limitations:
– Poor compliance by subjects.
– Greater exposure of subjects to the test drug, increasing the potential
for adverse reactions.
In multiple dose study the drug should be administered for 5 to 6 elimination
half-lives until the steady-state has been reached, before collecting the
blood samples.
• ASSESSMENT OF BIOAVAILABILITY
 Pharmacokinetics method – This method is more practical and
discriminative. Pharmacokinetic methods are of two types.
a) Determination of whole blood, plasma or serum concentration
b) Urinary excretion method
 Pharmacodynamic methods:
• Acute Pharmacologic Response Method : When bioavailability
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measurement by pharmacokinetic method is difficult, an acute
pharmacologic effect such as effect on pupil diameter, heart rate or BP can
be useful as an index of drug bioavailability.
Disadvantage: It tends to be complex, expensive, time-consuming and
require a sensitive and quantitative measure of the desired response.
Therapeutic Response Method: Clinical response of the drug for which it
is intended to be used is measured.
E.g.: heart rate, body temperature, blood sugar levels, and for antiinflammatory drugs, reduction in inflammation is determined.
Drawbacks: quantification of observed response is too improper to allow for
reasonable assessment of relative bioavailability between two dosage forms
of the same drug.
• A) The blood (or serum or plasma)
concentration-time curve -
• These are based on the assumption that there is a direct relationship
between the concentration of drug in blood or plasma and the
concentration of drug at the site of action.
• If the drugs are given to the volunteers through iv dose, the blood
samples should be withdraw after 5min. And the frequency of sampling
should be 15min.
• In case of one compartment open model – at elimination phase 3 points
are required to describe he
kE value.
• In case of two compartmental model – 5 to 6 points are required to
describe kE value.
• If Oral dose is given – 3 points are required to describe ka value.
• The key parameters for determining bioavailability
AUC: The AUC is proportional to the total amount of drug reaching the
systemic circulation, and thus characterizes the extent of absorption.
Cmax: Gives indication whether drug is sufficiently absorbed systemically to
provide a therapeutic response.
Tmax: The Tmax reflects the rate of drug absorption, and decreases as the
absorption rate increases.
MEC: The minimum plasma concentration of the drug required to achieve a
given pharmacological or therapeutic response
MSC: plasma concentration of the drug beyond which adverse effects are
likely to happen.
• THERAPEUTIC RANGE-The range of plasma drug concentration in which the
desired response is achieved yet avoiding adverse effect. The aim is clinical
practice is to maintain plasma drug concentration within the therapeutic
range.
• ONSET OF ACTION-On set of action is the time required to achieve the
minimum effective plasma concentration following administration of drug
formulation.
• DURATION OF ACTION-Duration of action of the therapeutic effect of the
drug is defined as the time period during which the plasma concentration of
the drug exceeds the minimum effective level.
• INTENSITY OF ACTION-In general, the difference between the peak plasma
concentration and the minimum effective plasma concentration provides a
relative measure of the intensity of the therapeutic response of the drug.
B) Urinary Excretion Data:
• These studies are based on the principle that urinary
excretion of the unchanged drug is directly proportional
to the plasma concentration of total drug.
• This technique of studying bioavailability is most useful
for those drugs that are not extensively metabolized
prior to urinary elimination.
• The three major parameters examined in urinary excretion
data are as follow:
1.(dXu/dt)max : maximum urinary excretion rate,
gives the rate of appearance of drug in the urine is
proportional to its concentration in systemic circulation. Its
value increases as the rate of and/or extent of absorption
increases
2. (tu)max : time for maximum excretion rate, is
analogous to the of plasma level data, its value decreases
as the absorption rate increases.
3. Xu : cumulative amount of drug excreted in the
urine is related to the AUC of plasma level data and
increases as the extent of absorption increases.
• These studies used for certain thiazide diuretics and
sulfonamides and for drugs that have urine as the site of
action-for example, urinary antiseptics.
BIOEQUIVALENCE STUDIES
 DEFINITIONS
 Equivalence – Equivalence is more relative term that compares
one drug product with another or with a set of established
standards. Equivalence may be defined in several ways:
 Chemical equivalence indicates that two or more dosage forms
contain the labeled quantities of drug.
 Clinical equivalence occurs when the same drug from two or
more dosage forms gives identical in vivo effects as measured
by a pharmacological response or by control of a symptom or a
disease.
 Therapeutic equivalence implies that one structurally different
chemical can yield the same clinical result as another chemical.
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The design and conduct of the study should follow ICH
regulations on Good Clinical Practice, including reference to
an Ethics Committee.
A bioequivalence study is basically a comparative
bioavailability study designed to establish equivalence
between test and reference products.
1. Design:
The various types of test designs that are usually employed in
bioequivalence stdies;
i. Completely randomised designs
ii. Randomised block designs
iii. Repeated measures, cross-over and carry-over designs
iv. Latin square designs
i. Completely randomised designs
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Method of randomisation
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Label all subjects with the same number of digits, for e.g., if there are 20
subjects, number them from 1 to 20.
Randomly select non-repeating random numbers (like simple
randomisation) with among these labels for the first treatment, and then
repeat for all other treatments.
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Advantages
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The design is extremely easy to construct.
It can accommodate any number of treatments and subjects.
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Disadvantages
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Although the design can be used for any number of treatments, it is best
suited for situations in which there are relatively few treatments.
Any unrelated sources of variability will tend to increase the random error
term, making it difficult to detect differences among the treatment (or
factor level) mean responses.
ii. Randomised block designs
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First, subjects are sorted into homogeneous groups, called blocks and the
treatments are then assigned at random within the blocks.
Method of Randomisation
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Subjects having similar background characteristics are formed as blocks.
Then treatments are randomised within each block, just like the simple
randomisation.
Randomisations for different blocks are done independent of each other.
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Advantages
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It can accommodate any number of treatments or replications.
Different treatments need not have equal sample size.
The statistical analysis is relatively simple. The design is easy to construct.
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Disadvantages
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Missing observations within a block require more complex analysis.
The degrees of freedom of experimental error are not as large as with a
completely randomised design.
iii. Repeated measures, cross-over and carry-over designs
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This is essentially a randomised block design in which the same subject serves as a
block.
The administration of two or more treatments one after the other in a specified or
random order to the same group of patients is called a crossover design or
change-over design
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Method of Randomisation
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Complete randomisation is used to randomise the order of treatments for
each subject. Randomisations for different subjects are independent of
each other.
Advantages
They provide good precision for comparing treatments because all sources
of variability between subjects are excluded from the experimental error.
It is economic on subjects. This is particularly important when only a few
subjects can be utilized for the experiments.
Disadvantages
There may be an order effect, which is connected with the position in the
treatment order.
There may be a carry-over effect, which is connected with the preceding
treatment or treatments.
iv. Latin square designs
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A Latin square design is a two-factor design (subjects and treatments are the two
factors) with one observation in each cell.
Such a design is useful to compare the earlier ones when three or more
treatments are to be compared and carry-over effects are balanced.
In a Latin square design, rows represent subjects, and columns represent
treatments.
Advantages
It minimizes the inter-subject variability and carry-over effect in plasma
drug levels.
Minimizes the variations due to time effect.
Treatment effects can be studied from a small-scale experiment. This is
particularly helpful in preliminary or pilot studies.
Disadvantages:
The randomisation required is somewhat more complex than that for
earlier designs considered.
The study takes a long time since an appropriate washout period between
two administrations is essential which may be very long if the drug has a
long t½.
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WASHOUT PERIOD
The time interval between the 2 treatments is
called as WASHOUT PERIOD
It is required for the elimination of the
administered dose to avoid the carry over effect
Large no. of drugs have been found to have half –
life between 1-10 hrs, awashout period of 1 week
was usually found suitable in most cases.
2. Subjects:
A. Selection of subjects:
 Aim to minimize variability and permit detection of
differences between pharmaceutical products.
 The studies should normally be performed with healthy
volunteers.
 They should be screened for suitability by means of clinical
laboratory tests, review of medical history, and medical
examination.
B. Standardization of the study:
 The test conditions should be standardized in order to
minimize the variability of all factors involved.
 Standardization of the diet, fluid intake and exercise is
recommended.
 The subjects should not take other medicines during a
suitable period before and during the study.
C. Inclusion of patients:
 If the investigated active substance is known to have adverse
effects and the pharmacological effects or risks are considered
unacceptable for healthy volunteers it may be necessary to use
patients instead, under suitable precautions and supervision.
D. Genetic pheno-typing:
Pheno-typing of subjects should be considered as well in
crossover studies.
 If a drug is known to be subject to major genetic polymorphism,
studies could be performed in panels of subjects of known
phenotype or genotype.
3. Characteristics to be investigated:
 In most cases evaluation of bioavailability and bioequivalence
will be based upon the measured concentrations of the parent
compound.
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In some situations, measurements of an active or inactive metabolite
is carried out.
 The plasma concentration versus time curves are mostly used to
assess extent and rate of absorption.
 The use of urine excretion data may be advantageous in determining
the extent of drug input in case of products predominately excreted
renally.
 Specificity, accuracy and reproducibility of the methods should be
sufficient.
4.Chemical analysis:
 It is conducted according to the applicable principles of Good
Laboratory Practice (GLP).
 Determination of the active moiety and/or its biotransformation
product(s) in plasma, urine or any other suitable matrix must be well
characterized, fully validated and documented to yield reliable results
that can be satisfactorily interpreted.
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5 Reference and test product:
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The choice of reference product should be justified by the
applicant and agreed upon by the regulatory authority.
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The test products used in the biostudy must be prepared in
accordance with GMP-regulations.
6.Data analysis:
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To quantify the difference in bioavailability between the
reference and test products and to demonstrate that any
clinically important difference.
A.Statistical analysis:
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The statistical analysis (e.g. ANOVA) should take into
account sources of variation that can be reasonably
assumed to have an effect on the response variable.
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Pharmacokinetic parameters derived from measures of
concentration, e.g. AUC, Cmax should be analyzed using
ANOVA.
B. Acceptance range for pharmacokinetic parameters:
1.AUC-ratio:
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It should lie within an acceptance interval of 0.80-1.25.
2.Cmax-ratio:
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It should lie within an acceptance interval of 0.80-1.25.
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The wider interval must be 0.75-1.33.
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For tmax if there is a clinically relevant claim for rapid release or action or
signs related to adverse effects. The interval should lie within a clinically
determined range.
C. Handling deviations from the study plan:
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The protocol should also specify methods for handling drop-outs.
D.A remark on individual and population bioequivalence:
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Bioequivalence studies are designed for population and individual is
limited.
7.In vitro dissolution for bioequivalence study:
 The term commonly used to describe is "in-vitro/in-vivo
correlation".
 The specifications for the in vitro dissolution of the product
should be derived from the dissolution profile of the batch.
A. Official dissolution tests:
Apparatus 1, (basket method).
 Apparatus 2 (paddle method).
B.Parameters used:
1. Degree of agitation
2. Size and shape of container
3. Composition of dissolution medium
• pH, ionic strength, viscosity
4. Temperature of dissolution medium
5. Volume of dissolution medium
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8.Reporting of results:
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The report of a bioequivalence study should give the complete
documentation of its protocol, conduct and evaluation
complying with GCP-rules and ICH guideline.
Drop-out and withdrawal of subjects should be fully
documented.
The method used to derive the pharmacokinetic parameters
should be specified.
The analytical report should include the results for all standard
and quality control samples.
APPLICATIONS FOR PRODUCTS CONTAINING
APPROVED ACTIVE SUBSTANCES:
A. Bioequivalence studies:
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In vivo bioequivalence studies are needed when there is
a risk that possible differences in bioavailability may
result in therapeutic in equivalence.
oral immediate release drug formulations with
systemic action
Non-oral and non-parenteral drug formulation
designed to act by systemic absorption.
Sustained or otherwise modified release drug
formulation.
Fixed-dose combination products.
 REFERENCES:
 Brahmankar.D.M,Sunil.B.Jaiswal,Vallabh Prakashan
Biopharmaceutics and Pharmacokinetics-A Treatise
,page no.282-305.
 LeonShargel & Andrew Yu Applied Biopharmaceutics &
pharmacokinetics, page no 247-260.
 Madan.PL Biopharmaceutics & pharmacokinetics page
no 118-127.
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