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Bioavailability
The in- vitro methods of evaluating
dosage forms provide only indirect
evidence of the therapeutic utility of the
drug in a given dosage form.
It would be more desirable to employ a
direct means of ascertaining the utility of
a particular dosage form.
In order to this in-vivo methods are
needed which will reflect the true safety
and efficiency of the dosage forms.
The estimation of the bioavailability of a
drug in a given dosage form is direct
evidence of the efficiency with which a
dosage form performs its intended
function.
The bioavailability of a drug is controlled
by three principle factors:
1. The rate and extent of release of the
drug from the dosage form.
2. The subsequent absorption from
solution state
3. The biotransformation during the
process of absorption.
Plasma concentration following the oral
administration of a drug assumes four
sequential phases:
Absorption >elimination
Absorption = elimination
Absorption<elimination
Absorption = 0, elimination> 0
Figure 1
The estimation of the bioavailability
from plasma concentration profiles
requires a thorough understanding of
the nature of these profiles.
Graphically bioavailability of a drug is
portrayed by conc-time curve of
administered drug in appropriate
tissue system, e.g. plasma.
Bioavailability data are used to
determine :
1. The amount or proportion of drug
absorbed from a formulation or
dosage form.
2. The rate at which the drug was
absorbed
3. The duration of the drug presence in
the biologic fluid or tissue, and when
correlated with patient response.
4-The relationship between drug blood
levels and clinical efficacy and toxicity.
Blood (or serum or plasma) concentration
time curve:
Following
oral
administration
of
medication if blood samples are drawn
from the patient at specific time intervals
and analyzed for drug content, the
resulting data may be plotted on ordinary
graph paper to yield the type of blood
level curve presented in the figure.
Figure 2
When the drug is first administered the
blood concentration of the drug should be
zero.
As the drug pass into the stomach or
intestine, it released from the dosage
form and absorbed.
As the sampling and analysis continue
the blood sample increasing concentration
of drug until the maximum (peak) conc.
(Cmax) is reached.
Then the blood level of the drug
decrease until reach to zero.
The diminished blood level of drug
after peak indicates the rate of the drug
elimination, and it is greater than the
drug absorption.
The drug absorption does not
terminate after the peak blood level
reached, but may continue for some
time.
Similarly the process
of
drug
elimination is continuous one. It begins
as soon as the drug first appear in the
blood stream and continuous until all
the drug has been eliminated.
A urine analysis for drug or its
metabolites may be used to indicate the
extent of drug absorption and/or rate
of drug elimination from the body.
Parameters
for
assessment
and
comparison of bioavailability from blood
level curves
The parameters used for comparison of
oral administration of single doses of two
formulations of the same drug entity are
the following:
1. The peak height concentration (Cmax)
2. The time of peak concentration (Tmax)
3. The area under blood concentration –
time curve (AUC)
Peak height concentration (Cmax)
It the maximum drug conc. observed in
blood plasma following a dose of the
drug
For conventional dosage forms, as
tablets and capsules, the maximum will
usually occur at only single time point is
Tmax
Figure 3
This figure represent conc- time curve
showing different peak height conc. for
equal amounts of drug from two
different formulations A,B following oral
administration.
The horizontal line represent minimum
effective conc (MEC) for drug substance
(4 ug/ml)
It is apparent from the figure that
formulation A will achieve the required
blood level to produce pharmacologic
effect, while formula B will not.
If the minimum toxic conc. (MTC) is 4
ug/ml) and MEC is 2 ug/ml ,then the
formula A will result in toxic effects while
formula B give desired effect.
The objective in individual dosing of
patient is to achieve the MEC but not MTC.
The size of the dose administered
influence the blood level conc. and Cmax
for the drug substance.
Figure4
 In this example it is assumed that all
doses are completely absorbed and
eliminated at the same rates.
It is evident that as the dose
increases, the Cmax is higher and
AUC increases. The peak time Tmax is
the same for each dose.
Time of peak (Tmax)
The second parameter of importance in
assessing the comparative bioavailability
of two formulations is time required to
achieve the maximum level of drug in
blood (Tmax)
This parameter reflects the rate of
drug absorption from a formulation.
It is the rate of drug absorption that
determine the time needed for MEC to
be reached and thus for the initiation
of the desired pharmacologic effect.
The rate of drug absorption also
influence the period of time over which
the drug enters the blood stream, and
therefore affect the duration of time
looking at figure, formulation A allows
the drug to reach the MEC within 30
min., and a peak in 1 hour.
Figure 5
Formulation B has slower rate of drug
release, it reached MEC 2hour, and its
peak conc. after 4 hours.
Thus formulation A permits the
greater rate of drug absorption, it
allows drug to reach both MEC, and its
peak height
sooner than drug
formulations B.
On the other hand, formulation B
provides greater duration of action, and
drug concentration maintained above
MEC. 8 hours from (2-10) to 5.5 (306h) for formulation A.
Thus if rapid onset of action is desired
a formulation similar to A would be
preferred ,but if longer duration action
is desired than rapid onset , a
formulation B would be preferred.
In sum, changes in the rate of drug
absorption will result in changes in the
values of both Cmax,Tmax.
Each
product
has
its
own
characteristics rate of absorption.
When rate of absorption decreased the
Cmax is lowered and Tmax occur at
later time.
If the doses of the drug are the same
and completely absorbed, the AUC for
each is essentially the same.
For example: a higher or earlier peak
does not necessarily mean greater
overall absorption than from a product
giving a smaller or delayed peak.
The total absorption of drugs is
proportional not only to the plasma
conc. achieved but also to the length of
time these conc. persist in the blood or
plasma.
Area under the serum conc. –time curve
The AUC is considered representative
of total amount of drug absorbed into
circulation flowing the administration of
a single dose of that drug.
Equivalent doses of a drug when fully
absorbed into the circulation would
produce the same AUC.
Thus two curves much unalike in terms
of Cmax and
One parameter that characterizes this
aspect is the area under the plasma
conc. vs time profile.
Many methods are available for the
measuring of AUCs.
Three of these are described as
follows.
The trapezoidal rule
This is the simplest of all the method
and involves the breaking up of the
plasma conc. vs time profile into
several trapezoids, calculating the area
of individual trapezoids, and the adding
up these areas to arrive at a cumulative
AUC:
AUC1 = (Co + C1) (t1-t0) + C1+C2 (t2-t1) +
2
2
-----------+ (Cn-1+Cn) (tn- tn-1)
= Σ (Ci-1+ Ci) (ti – ti-1)
2
The units for AUC are : conc × time e.g. µg.hr /ml or
mg.min/liter.
Example
Time(hrs) C,µg/ml trapezoidal area cumulative area
0
0
_
1
2
1
1
2
15
17
18
3
5
20
38
4
1
6
44
5
The AUC is proportional to the dose
absorbed only when the calculation are
extended to the point where the plasma
conc. approaches zero.
2-Integration Method
The rate of change of plasma conc
(c ) is described as
dc/dt = rate of absorption – rate of
elimination
= KaXa – KX
Where Ka and K are absorption and
elimination rate constants
Xa and X are the amounts of drug in
GIT and body respectively
An integration of this equation between
limits of time for which the drug remains
in the body, as reflected by plasma conc.
gives
C= A ( e-kt - e-kat)
And the total area under the curve
(AUC), for which the total integral between
time zero and infinity is given by:
AUC = A{ 1/K -1/Ka}
3-Physical methods
A variety of methods which utilize
physical properties can be used to
calculate AUCs
For example, if plasma conc. profiles
are plotted on smooth paper, these can
be cut out and weighed on an
electronic balance.
Area under the serum conc- time curve
The area under the curve (AUC) is
considered representative of total
amount of drug absorbed into
circulation following the administration
of a single dose of that drug
Equivalent doses of a drug when fully
absorbed into the circulation would
produce the same AUC
Thus two curves much unalike in
terms of Cmax, Tmax, but may be much
alike in AUC and thus in the amount of
drug absorbed.
From the last figure, formula A with
AUC = 34.4, and formula B with AUC =
34.2 are essentially the same
If equivelent doses of drug in different
formulations produce different AUC
values.
The difference exist in the extent of
absorption between the formulations.
F = (AUC)oral / (AUC)IV
F : is the fraction of dose absorbed
In practice, it would rare for a drug to
be completely absorbed in the circulation
followed oral administration.
As noted earlier many drugs undergo
the first- pass effect lead to some
metabolic degradation. In addition factors
of drug product formulation, drug
dissolution,
chemical
and
physical
interactions with the GI contents, gastric
emptying time, intestinal motility and
others
contribute
the
incomplete
absorption of administered dose of a
drug.
The absolute bioavailability following
oral dosing is generally compared to IV
dose.
Urinary excretion data
Cumulative urinary drug excretion
curves
Measurement of the conc. of intact drug
and/or its metabolites in the urine can
also be used to assess bioavailability
The assessment of bioavailability by
urinary excretion is based on the
assumption that the appearance of
drug and its metabolites in the urine is
a function of the rate and the extent of
absorption.
This assumption is only valid when the
drug and / or its metabolites are
extensively excreted in the urine, and
where the rate of urinary excretion is
proportional to the conc. of the intact
drug in the blood plasma. This
proportionally is not hold if:
The drug and/or metabolites are
excreted by active transport process
into distal kidney tubule.
The intact drug and/or its metabolites
are weakly acidic or weakly basic ( i.e.
excretion is dependent on urine pH).
The excretion rate depends on urine
flow.
The important parameters in urinary
excretion studies are: the cumulative
amount of intact drug and its
metabolite and the rate at which this
excretion takes place.
Figure
The
segment
(x-y)
absorption phase
reflect
the
The slope of this segment related to
the rate of absorption of the drug.
The total amount of intact drug or its
metabolite excreted in the urine at point
Z.
The total amount of drug excreted at
point Z may be quite different from the
total amount of drug administered
(dose), either because of incomplete
absorption or because of drug being
eliminated by process other than
urinary excretion.
Absolute and relative bioavailability
Absolute bioavailability
It is the fraction (or percentage) of the
administered dose which is absorbed
intact into systemic circulation.
•Absolute
bioavailability
may
be
calculated by comparing the total amount
of intact drug that reaches the systemic
circulation after administration of a
known dose of the dosage form via a
route of administration with total amount
that reaches the systemic circulation after
the administration of an equivalent dose
of drug in the form of IV injection bolus.
•IV injection bolus is used as a
reference to compare the systemic
availability of the drug administered
via different routes, because when a
drug is delivered IV the entire
administration dose is introduced
directly into systemic circulation, i.e. it
has no absorption barrier to cross and
it is considered to be totally
bioavailable.
•The absolute bioavailability of a given
drug using plasma data
may be
calculated by comparing the total AUC
following
the
administration
of
equivalent doses of the drug via
absorption site and via IV route in the
same subject on different occasions.
For equivalent doses of administered
drug:
( AUC) abs
Absolute bioavailability = ------------(AUC ) IV
(AUC)abs : area under the curve
following administration of single dose
via absorption site.
(AUC)Iv : area under the curve following
administration of single dose via rapid IV
injection.
Figure
If different doses of the drug are
administered by both routes, a correction
for the sizes of the doses can be made as
follows
( AUC) abs/ Dabs
Absolute bioavailability = ----------(AUC ) IV/ DIV
Where
Dabs is the size of single dose of the
drug taken via absorption site
DIV : is the size of single dose of the
drug taken via IV injection
Absolute bioavailability using urinary
excretion data may be determined by
comparing the cumulative amount of
unchanged drug excreted in urine
following administration of drug via
absorption site and that of IV route,
respectively on different occasions to
the same subject.
For equivalent doses of administered
drug
( Xu) abs
Absolute bioavailability = -----------(Xu) IV
(Xu) abs, (Xu)Iv are the total cumulative
amounts of unchanged drug excreted in
the urine after administration of
equivalent single dose via absorption
site and IV route.
For equivalent doses of administered
drug
( Xu) abs
Absolute bioavailability = -----------(XU) IV
(Xu) abs, (Xu)iv are the total cumulative
amounts of unchanged drug excreted in
the urine after administration of
equivalent single dose via absorption
site and IV route.
( Xu) abs/Dabs
Absolute bioavailability = ---------(XU) IV / DIv
It should be noted that the value
calculated for absolute bioavailability
will be only valid if the drug kinetics
of
elimination
and
distribution
independent of the route and time of
administration and also the dose of
size administered.
Relative bioavailability
In the case of drugs that cannot be
administered by IV bolus injection, the
relative bioavailability is determined
rather than absolute bioavailability.
In this case the bioavailability of a given
drug from a (test) dosage form is
compared to that of the same drug
administered in a standards dosage
form,
which
is
either
orally
administered solution or established
commercial preparations of proven
clinical effectiveness.
Hence the relative bioavailability is a
measure of the fraction of a given drug
that is absorbed intact into the systemic
circulation from a dosage form relative to
the recognized (i.e. clinically proven)
standards dosage form of the drug.
The Relative bioavailability of a given
drug administered as equal doses of a
test dosage form and a recognized
standard dosage form
the corresponding plasma conc-time
curve as follows
( AUCT) test
Relative bioavailability = ----------(AUC T ) stand
When different doses of the test and
standard dosage form are administered a
correction for the size of dose is made as
follows:
( AUCT) test/ D test
Relative bioavailability = ------------(AUC T )stand/ D stand
where D test and D stand and the sizes of
the single doses of the test and
standard dosage form respectively .
urinary excretion data may also be
used to measure relative bioavailability
(Xu) test
Relative bioavailability = ----------(Xu ) standard
where Xu test and Xu stand and are the
total cumulative amounts of unchanged
drug excreted in the urine for test and
stand drug
when different doses are used from test
and standard
(Xu) test / D test
Relative bioavailability = ----------------(Xu ) stand/ D stand
Relative
bioavailability
measurements are often used to
determine the effects of dosage form
differences on systemic bioavailability
of a given drug.
Numerous dosage form factors can
influence the bioavailability of a drug.
These factors include
1-the type of dosage form (tablets,
capsules, solution, and suspension),
differences in the formulation of a
particular type of dosage form,
2and
manufacturing
variables
employed the production of a particular
type of dosage form.
Bioequivalence of drug products
It has become well established that the
rate and extent to which a drug in a
dosage form become available for
biologic
absorption
or
utilization
depends on great measure upon the
materials utilized in the formulation and
also on the method of manufacture.
Thus the same drug when formulated in
different dosage forms may be found to
possess
different
bioavailability
characteristics and hence exhibit different
clinical effectiveness.
Further two identical or equivalent
products, of the same drug in the same
dosage form type but differing in
formulation
materials
or
method
manufacture
may
vary
widely
in
bioavailability and other single or multiple
dose.
According
to
USP,
significant
bioavailability
and
bioequivalence
problems may be revealed through
dissolution testing and are generally the
result of one or more of the following
casual factors:
1. The drug particle size
2. Excessive amount of the lubricant
mg.stearate in the formulation
3. Coating materials especially shellac
4. Inadequate amounts of tablets and
capsule disintegrants
Pharmaceutical equivalents
Are drug product that contain identical
amounts of identical active drug
ingredient i.e. the same salt or ester of
the same therapeutic moiety in
identical dosage forms but not
necessarily
containing
inactive
ingredients.
They meet the identical compendia or
other applicable standard of identity,
strength quality and purity including
potency and where applicable content
uniformity,
disintegration
times
and/or dissolution rates.
Pharmaceutical alternatives
Are drug product that contain the
identical moiety or its precursor, but
not necessarily in the same amount or
dosage form or as the same salt or
ester.
Each such drug product individually
meets either the identical or its own
respective compendia standard (as
above).
Bioequivalent drug products
Are pharmaceutical equivalents or
pharmaceutical alternatives whose rate
and extent of absorption do not show a
significant
differences
when
administered at the same molar dose of
the therapeutic moiety under similar
experimental conditions
Some pharmaceutical equivalents or
pharmaceutical alternatives may be
equivalent in the extent of their
absorption but not in the rate of
absorption
Therapeutic equivalents has been
used
to
indicate
pharmaceutical
equivelents which when administered to
the same individuals in the same
dosage
regimens,
will
provide
essentially the same therapeutic effect.
The most common experimental plan
to compare the bioavailability of two
drug products in the simple crossover
design study.
In this method each of the 12-24
individuals in the group of carefully
matched subjects (usually healthy) adult
males between 18-40 years of age of
similar height and weight.
Once the first assigned product is
administered samples of blood or
plasma are drawn from the subjects at
predetermined times and analyzed for
active drug moiety and its metabolites
as a function of time.
The same procedure is then repeated
(crossover) with the second product
after an appropriate interval of time
i.e. a washout period to ensure that
there is no residual amount of the first
drug.
After that the patient population
data tabulated and parameters as
Cmax, Tmax and AUC, are then
analyzed with statistical procedures.
Absolute Bioequivelence
It is rarely occur between drug products
Such absolute equivelency would yield
serum concentration time curves for the
products involved that would be exactly
superimposable.
This simply is not expected of products
which are made at different times,
different batches ,or by different
manufactures.
In most studies of bioavailability, the
originally marketed product known as
pioneer or innovator is recognized as the
established product of the drug and is
utilized as the standard for bioavailability
comparative study.
According to the FDA, a generic
drug is considered bioequivalent if
the rate and extent of absorption do
not show a significant difference
from that of the pioneer drug when
administered at the same molar dose
of the therapeutic ingredient under
the same experimental conditions
To gain FDA approval a generic drug product
must:
1- Contain the same active ingredients as the
pioneer drug
2- Be identical in strength, dosage form and
route of administration
3- Have the same indications and precautions
for use and other labeling instructions
4- Be bioequivalent
5- Meet the same batch-to-batch
requirements for identity, strength, purity,
and quality.
6- Be manufactured under the same strict
standards of FDA’s for pioneer products
In-vitro demonstration of bioavailability:
Dissolution testing:
Dissolution testing the most important
way to study, under in-vitro conditions,
the release of drug from solid dosage
form, and thus represents an important
test to assess factors that affect the
bioavailability of a drug from solid
preparation
Dissolution studies are carried out for
several reasons
To evaluate the potential effect of
formulations and process variables on the
bioavailability of a drug
To ensure that preparations comply with
product specifications
To indicate the performance of the
preparation under in-vivo conditions
•This last point requires that in-vitro
dissolution data correlate with the in-vivo
performance of the dosage form, which
must be experimentally varied
•The term in-vitro/in-vivo correlation in
this context is related to the correlation
between in-vitro dissolution and the
release or uptake of the drug in-vivo