BIOAVAILABILITY

“Rate & extent to which the active moiety is

absorbed from a drug product and
becomes available at the site of action”
OR
“Drug product performance studies are
used to define the effect of changes in
physicochemical properties of the drug
substance, the formulation of the drug, and
the manufacture process of the drug
product(dosage form).”
WHY BIOAVAILABILITY IS IMPORTANT?
•Dynamic increase in utilization of generics
faced by the pharmacists

•Selection of appropriate drug products

•FDA guidelines
 TYPES OF BIOAVAILABILITY
Bioavailabilty comprises of two types:

1.Absolute bioavailabilty

2.Relative bioavailabilty
 ABSOLUTE BIOAVAILABILITY
•Systemic availability of a drug after
extravascular administration compared to IV
dosing.

•“F” , is the fraction of an administered dose

and ranges from F=0 to F=1

•Comparison between the AUC for the test

product and that the AUC of the IV
( STANDARD)
DETERMINATION OF ABSOLUTE BIOAVAILABILITY
Absolute availability=F=[AUC]EV / DoseEV
[ AUC]IV / DoseIV

WHERE “F” may be expressed as a fraction or a

percent by multiplying F*100.

Absolute availability Using urinary drug excretion

data:
F =[Du]EV / DoseEV
[Du]IV / DoseIV
• For all the drugs given by IV BOLUS INJECTION , F=1 and for
all the drugs given extravascularly administered F may not
exceed 100 percent.
 RELATIVE BIOAVAILABILITY
“Availability of the drug product as
compared to another dosage form or
product of the same drug given in the same
dose”

•The relative bioavailability of product A

compared to product B, both products
containing the same dose of the same
drug, is obtained by comparing their
respective AUCs.
 DETERMINATION OF RELATIVE
BIOAVAILABILITY
Relative bioavailability =[AUC]A / Dose A
[AUC]B / Dose B
Where , drug product B is the reference standard. When the
bioavailability of the generic drug is considered, It is usually
the relative bioavailability that is referred to. A more general
form of the equation results from considering the possibility
of different doses,

Comparative Bioavailability =
[AUC]Generic / Dose Generic
[AUC]Brand / Dose Brand
 DIFFERENCE BETWEEN ABSOLUTE AND
RELATIVE BIOAVAILABILTY
DRUG PRODUCT AREA UNDER THE
CURVE
A. Intravenous injection 100

B. Oral dosage form, brand or 50

reference standard

C. Oral dosage form, generic product 40

The “F” for product B and C is 50%
(F=0.5) and 40%(F=0.4),respectively.
However when the two oral products
are compared, the relative
bioavailability of product C as
compared to product B is 80%.
 Methods for assessing BA of drugs
• BA testing is a means of predicting the clinical efficacy of a
drug, the estimation of the BA of the drug in a given dosage
form is direct evidence of the efficiency with which a dosage
form performs its intended therapeutic function.

• BA determinations are performed by drug manufacturers to

ensure that a given drug product will get the therapeutic agent
to its site of action in adequate concentration.

• Used to compare the availability of a drug substance from

different dosage forms or from the same dosage form produced
by different manufacturers.
 Methods

Both direct and indirect methods maybe used to

asses drug bioavailability from a drug product.

Following are the methods used in BA studies

1. Plasma drug concentration
2. Urinary drug excretion
3. Acute pharmacodynamic effect
4. Clinical oberservations
5. In-vitro bioavailability studies
 Plasma drug concentration
• Commonly used
• Direct method to asses the clinical performance of a drug involves
measurement of the drug concentrations in the blood, plasma or serum.
• In such studies a single dose of the drug product is administered to a panel
of normal, healthy adult subjects.
• Blood samples are collected over a period of time following administration
and are analyzed for drug content.
• Based on the blood concentration as a function of time inferences are
drawn regarding the rate and extent of absorption of the drug.
• These studies are relatively easy to conduct and require a limited number
of subjects.
• Blood level studies 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.
• By monitoring the concentration in the blood, it is thus possible to obtain an
indirect measure of drug response.
• Following the administration of single dose of a medication, blood samples
are drawn at specific time intervals and analyzed for drug content. A profile is
constructed showing the concentration of drug in blood at the specific times
the samples were taken.
• The key parameters to note are:

AUC:
• The area under the plasma level time curve, AUC, is a measurement of the
extend of drug bioavailability.
• The AUC reflects the total amount of active drug that reaches the circulation.
• It is the area under the drug plasma level-time curve from t=0 to t= ∞, and is
equal to the amount of unchanged drug reaching the general circulation
divided by the clearance.
[AUC]0∞= Fd0 = Fd0

clearance kVD
 Methods of determination of AUC
1. Planimetric method: this method involves plotting the data for the
reference as well as for different samples in the form of graphs. Then cutting
out the graphs in pieces which are joined together to form a measurable
shape such as a triangle, rectangular, square, etc. later on mearsurement is
performed using a planimeter or ruler which is used for the calculation of the
AUC for the reference and the sample specimens
2. Cut and weight method: this method also involves plotting of the data for
different samples on the same scale, then cutting out the curves and
weighing them. The areas may be calculated from the weights if weight of
one unit of the paper is known.
3. Trapezoidal method: the trapezoidal rule is frequently used in
pharmacokinetics to calculate the areas under the plasma drug
concentration time curve (AUC). In this case the AUC is estimated by
dividing the curve into several sections that approximate a series of
trapezoids with a triangle at each end of the curve. The individual area of
each trapezoid and triangle is calculated with help of
4- Numerical method or Definite integral method:
this method also employees the trapezoidal rule and
utilizes the sum of individual areas under the curve.
B- Peak height or Cmax:
The peak plasma drug concentration, Cmax, represents the maximum
plasma drug concentration obtained after oral administration of drug. Cmax
will increase with an increase in the dose, as well as with an increase in the
absorption rate. For many drugs a relationship is found between the
pharmacodynamic drug effect and the plasma drug concentration.
Units are ng/mL, ug/mL

C- Time of Occurrence or tmax:

The time of peak plasma concentration, tmax, corresponds to the time
required to reach maximum drug concentration after drug administration.
For a given dose and bioavailability fraction, tmax is inversely dependent on
absorption rate, hence the tmax reflects the rate of drug absorption, and
decreases as the absorption rate increases.
Units are hours,minutes.
 Urinary Drug Excretion
 An alternative and indirect method for
assessment of bioavailability of drugs is to use
the urinary excretion data.
 These studies involve collection of urine
samples and the determination of the total
quantity of drug excreted in the urine as s
function of time.
 These studies are based on the premise that
urinary excretion of the unchanged drug is
directly propotional to the plasma concentration
of total drug.
 Thus, the total quantity of drug excreted in
the urine is a reflection of the quantity of drug
absorbed from the gastrointestinal tract.
 Example
• Two products A and B , each containing
100mg of the same drug are
administered orally. A total of 80mg of
drug is recovered in the urine from
Product A, but only 40mg is recovered
from Product B.

• This indicates that twice as much drug

was absorbed from product A as from
product B.
 Continued….

• This technique of studying bioavailability is

most useful for those drugs that are not
extensively metabolized prior to urinary
elimination.
• As a rule-of-thumb, determination of
bioavailability using urinary excretion data
should be conducted only if atleast 20% of a
dose is excreted unchanged in the urine after
an IV dose.
• Other condition which must be met for this
method to give valid results include:
• The fraction of drug entering the bloodstream and being
excreted intact by the kidneys must remain constant

• Collection of the urine has to continue until all the drug has
been completely excreted(five times the half- life)

• Urinary excretion data are primarily useful for assessing

extent of drug absorption, although the time course for the
cumulative amount of drug excreted in the urine can also be
used to estimate the rate of absorption.

• In practice, these estimates are subject to a high degree of

variability , and are less reliable than those obtained from
plasma concentration-time profile.

• Thus, urinary excretion of drug is not recommended as a

substitute for blood concentration data, rather , these studies
should be used in conjunction with blood level data for
confirmatory purpose
• A)
Du : The cumulative amount of the drug
excreted in urine is related directly to the total
amount of drug absorbed. Experimentally,
urine samples are collected periodically after
administration of a drug product. Each urine
sample is analyzed for free drug using a
specific assay procedure .A graph is
constructed that related cumulative drug
excreted to the collection-time interval
Between The Cumulative Amount Of Drug Excreted In Urine
And The Relationship The Plasma Drug Level-time Curve

• When the drug is almost completely eliminated, the plasma concentration approaches
zero and the maximum amount of drug eliminated, the plasma concentration
approaches zero and the maximum amount of drug excreted in urine(DU) is obtained
 B) Rate of drug excretion in urine
(Ddu/dt):
• Most of the drugs are eliminated by
a first-order rate process, the rate
of drug excretion is dependant on
the first order elimination rate
constant k and the concentration of
drug in plasma Cp. The maximum
rate of drug excretion is 71g/ml
after a period of 70hours.
C) Time for maximum urinary drug
excretion:
• The slop of the curve is related to
the rate of drug absorption,
whereas the point at the 70th hour
time is related to the total time
required for the drug to be
absorbed and completely excreted
after its administration.
 Acute pharmacodynamic or pharmacologic effect:
• In some cases, the quantitative measurement of a drug is not available, or it
lacks sufficient accuracy and/or reproducibility.
• In such cases the an acute pharmacodynamic effect, such as effect on pupil
diameter, heart rate, or blood pressure, can be used as an index of drug
bioavailability.
• This method is based on the assumption that a given intensity of response
is associated with a particular drug concentration at the site of action; e.g.,
variation of meiotic response intensity can be directly related to the oral
dose of chlorpromazine. In this case, an acute pharmacodynamic effect-
time curve is constructed.
• However, monitoring of pharmacologic data is often difficult, precision
and reproducibility are difficult to establish, and there are only a limited
number of pharmacologic effects (e.g. heart rate, body temperature, blood
sugar levels)that are applicable to this method.
• Measurement of the pharmacodynamic effect should be made with
sufficient frequency to permit a reasonable estimate of the AUC for time
period at least three times the half-life of the drug
• . Pharmacodynamic parameters that are obtained include maximum
pharmacodynamics effect-time curve and onset time for
pharmacodynamics effect.
 Clinical observations:
• One method for assessing the bioavailability of a drug product
is through the demonstration of a clinically significant effect.
Well-controlled clinical trials in humans can establish the
safety and effectiveness of the drug product.
• However, this approach is the least accurate, least sensitive
and least reproducible of the general approaches for
determining in vivo bioavailability. Moreover, such clinical
studies are complex, expensive and time-consuming.

• They also require a sensitive and quantitative measure of the

desired response. Furthermore, response is often quite
variable, requiring a large test population. Practical
considerations therefore preclude the use of this method
except in initial stages of development while proving the
efficacy of a new chemical entity.
 Single-dose versus multiple -dose:
• Most bioavailability evaluations are made on the basis of single-dose
administration. The argument has been made that single doses are not
representative of the actual clinical situation, since in most instances,
patients require repeated administration of a drug.
• When a drug is administered repeatedly at fixed intervals, with the
dosing frequency less than five half-lives, drug will accumulate in the
body and eventually reach a plateau, or a steady-state.
• The amount of drug eliminated from the body during one dosing
interval is equal to the available dose(rate in = rate out); therefore, the
area under curve during a dosing interval at steady-state is equal to the
total area under the curve obtained when a single dose is administered.
This AUC can therefore be used to assess the extent of absorption of the
drug, as well as its absolute and relative bioavailability.
• Multiple-dose administration has several advantages over single-dose
bioavailability studies, as well as some limitations.
 Advantages:
1. Eliminates the need to extrapolate the plasma
concentration profiles to obtain the total AUC after a
single dose.
2. Eliminates the need for a long wash out period
between doses.
3. More closely reflects the actual clinical use of the
drug.
4. Allows blood levels to be measured at the same
concentrations encountered therapeutically.
5. Because blood levels tend to be higher than in the
single-dose method, quantitative determination is
easier and more reliable.
6. Saturable pharmacokinetics, if present, can be more
readily detected at steady-state.
 Limitations:
1. Requires more time to complete.
2. More difficult and costly to
conduct(requiring prolonged monitoring
of subjects).
3. Greater problems with compliance control.
4. Greater exposure of subjects to the test
drug, increasing the potential for adverse
reactions.
 In vitro dissolution studies
and bioavailability:
Drug dissolution studies may under certain
conditions give an indication of drug
bioavailability. Ideally the in vitro drug
dissolution rate should correlate with the in
vivo drug bioavailability .Dissolution studies
are often performed on several test
formulations of the same drug. The test
formulation that demonstrates the most rapid
drug dissolution in vitro will generally have the
most rapid rate of drug bioavailability in vivo.
 Continued…
• Pharmaceutical scientist have for many years been attempting to
establish a correlation between some physicochemical property of a
dosage form and the biological availability of the drug from that
dosage form.

• The term commonly used to describe this relationship is “in-vitro/in-

vivo correlation”. Specifically , it is felt that if such a correlation could
be established, it would be possible to use in-vitro data to predict a
drug’s in-vivo bioavailability .This would drastically reduce ,or in some
cases ,completely eliminate the need for bioavailability test.

• The desirability for this becomes clear when one considers the cost
and time involved in bioavailability studies as well as the safety issues
involved in administering drugs to healthy patients or subjects.

• It would certainly be preferable to be able to substitute a quick

,inexpensive in-vitro test for in-vivo bioavailability studies .This would
be possible if in-vitro tests could reliably and accurately predict drug
absorption and reflect the in-vivo performance of a drug in humans.
 Disintegration tests:
• The early attempts to establish an indicator of drug bioavailability
focused on disintegration as the most pertinent in-vitro parameters
.

• The first official disintegration test appeared in the United State

Pharmacopoeia (USP) in 1950.

• However ,while it is true that a solid dosage form must disintegrate

before significant dissolution and absorption can occur, meeting
the disintegration test requirement only insures that the dosage
form(tablet) will break up into sufficiently small particles in a
specified length of time. It does not ensure that the rate of solution
of the drug is adequate to produce suitable blood levels of the
active ingredient.

• Therefore, while the test for tablet disintegration is very useful for
quality control purposes in manufacturing , it is a poor index of
bioavailability.
 Dissolution test:
• Since a drug must go into solution before it can be absorbed , and since
the rate at which drug dissolves from a dosage form often determines its
rate and /or extent of absorption ,attention has been directed at the
dissolution rate. It is currently considered to be the most sensitive in-
vitro parameter most likely to correlate with bioavailability.
• There are two official USP dissolution methods:
• Apparatus 1 (basket method), Apparatus 2 (paddle method ).
• Dissolution tests are extremely valuable tool in ensuring the quality of a
drug product.
• Product to product variations are due to formulation factors, such as
particle size differences, excessive amount of lubricants and coatings.
These factors are reactive to dissolution testing. Dissolution test are
very effective in discriminating between and within batches of drug
product(s).These tests can exclude any unacceptable product.
• There are some problems with in-vitro dissolution testing which should
be noted, problems which make correlation with in-vivo availability
difficult. The first is related to instrument variance and the absence of
standard method. The tests describes in the USP are but few of the
large number of dissolution methods proposed to predict bioavailability.
The dissolution rate of a dosage form is dependent on the methodology
used in dissolution test, changes in apparatus ,dissolution medium etc
can dramatically modify the results.
 Continued…
• Another significant problem is related to the difference between in-
vitro and in –vivo environments in which dissolution occurs. In-vitro
studies are generally carried out under controlled conditions in one,
or perhaps two, standardized solvents. The in-vivo environment (the
gastrointestinal tract), on the other hand, is a continuously changing,
complex environment. There are many variables which can affect the
dissolution rate of the drug in the GIT , including pH, enzyme
secretions, surface tension, motility, presence of other substances
and absorption surfaces. Thus, drugs frequently dissolve in the body
at rates quite different from those observed in an in-vitro test
situation. Most of the official dissolution tests tend to be acceleration
dissolution tests which bear limited or no relationship with in-vivo
dissolution.

• Adding to the complexity of the correlation dissolution with in-vivo

absorption are factors such as drug-drug interactions, age , food
effects, health ,genetic background, biorhythm and physical activity.
All these factors may have an effect on the rate and extent of
absorption of a drug. Thus, the in-vivo environment is for more
complex ,variable , and unpredictable than any in-vitro test
environment , making in-vitro/in-vivo correlation very difficult .A simple
dissolution test in a standardized vehicle cannot reflect the in vivo
absorption of a drug across a population.
 Continued…
• Proper selection of the in-vitro and in-vivo parameters to be
correlated is critical in achieving a meaningful correlation.
• The in-vitro parameter should be selected that has the
greatest effect on the absorption characteristics of the drug.
There are several approaches to establishing a correlation
between the dissolution of the drug in-vitro and the
bioavailabilty of the drug in- vivo.
• The in-vitro – in-vivo correlative methods used most often are
of the single-point type where the dissolution rate(expressed
as the percent of drug dissolved in a given time , or the time
required for a given percent of the drug to dissolve) is
correlated to a certain parameter of the bioavailability.
• Examples of in-vivo parameters used include Cmax, AUC,
time to reach half-maximal plasma concentration, the average
plasma concentration after 0.5 or 1 hour, maximum urinary
excretion rate and cumulative percent excreted in urine after
a given time.
• According to Wagner, the best in vitro variable to use is the
time for 50 percent of the drug to dissolve , and the best
variable from in vivo data to use is the time for 50 percent of
the drug to be absorbed (Wagner,1970).
In-vitro/in-vivo correlation studies:
• There have been many attempts to establish in vitro / in vivo
correlations for a large variety of drugs.
• While there are many published examples of satisfactory
correlations between absorption parameters and in vitro dissolution
tests, most studies have resulted in poor, or moderate, in-vitro – in-
vivo correlations often involving agreement with only one of the
critical bioavailability parameters. Moreover , the positive
correlations that have been found generally apply only to the
specific formulation studied.
• There have been instances where the dissolution rates or various
formulations of the same drug have been significantly different , yet
little or no difference was observed in their bioavailability
parameters. There have also been cases where a drug has failed to
meet compendia dissolution standards but has demonstrated
adequate bioavailability.

• Welling states: “To the writer’s knowledge ,there have been no

studies that have accurately correlated in-vitro and in-vivo data to
the point that the use of upper and lower limits for in-vitro dissolution
parameters can be confidently used to predict in-vivo behavior and
therefore, to replace in-vivo testing”(Welling,1991).
 Continued…
• Even if an in-vitro test could be designed that would
accurately reflect the dissolution process in the GIT,
dissolution is only one of the many factors that affect a drug’s
bioavailability. For example, saturable presystemic
metabolism may affect the extent of drug absorption , but this
would not be predicted by an in-vitro test . Dissolution studies
also would not predict poor bioavailability due to instability in
gastric fluid or complication with another drug or food
component.

• Thus , the ultimate evaluation a drug product’s performance

under the conditions expected in clinical therapy must be an
in-vivo test ;a dissolution test is unlikely to entirely replace
bioavailability testing (FDA,1988;Kottke and Rhodes,1991).

• In-vitro methods are important in the development and

optimization of dosage forms while in-vivo tests are essential
in obtaining information on the behavior of medication in living
oraganisms. One cannot be substituted for the other
(HUttenrauch,1985)