BIOAVAILABILITY AND

BIOEQUIVALENCE DATA

ABSTRACT
To ensure a therapeutic equivalence between a pharmaceutically equivalent test drug and a
generic drug or test drug.

Aditya Kekan
Drug Regulatory Affairs
1.INTRODUCTION
1.1 BIOAVAILABILITY-
A) Bioavailability is a measurement of the rate and extent to which a therapeutically active
chemical is absorbed from a drug product into the systemic circulation and becomes available
at the site of action.
B) For most drugs that are taken orally, the active ingredients are released in the
gastrointestinal (GI) tract and arrive at their site of action via the systemic circulation.
C) Blood concentrations of the active ingredients and/ or their active metabolites thereby
provide a marker for the concentration at the site of action and a valid measure of
bioavailability.
D) A blood concentration-time curve (achieved by serial measurements over time) reflects
not just the release of the active ingredient from the drug and its absorption from the GI tract,
but also other factors including pre-systemic metabolism, distribution, and elimination.

Bioavailability is assessed using two main pharmacokinetic variables:

• The area under the blood concentration versus time curve (AUC).
• The maximum blood concentration (C max).

1.2 BIOEQUIVALENCE-
A) Bioequivalence is a term in pharmacokinetics used to assess the expected in vivo
biological equivalence of two proprietary preparations of a drug. If two products are said to
be bioequivalent it means that they would be expected to be, for all intents and purposes, the
same.
If two drugs are bioequivalent, there is no clinically significant difference in their
bioavailability.
B) Although bioequivalence is most commonly discussed in relation to generic drugs, it is
important to note that bioequivalence studies are also performed for brand name drugs in
some situations such as:
• Between early and late clinical trial formulations or between the formulations used in
clinical trials and the product to be marketed for new drugs
• When changes in formulation have occurred after a brand name drug has been approved; for
example, a change in one or more excipients (inactive ingredients). Bioequivalence studies
are a surrogate marker for clinical effectiveness and safety data, as it would not normally be
necessary to repeat clinical studies for generic products.
. For oral drugs, it is accepted that if blood concentrations of the active ingredient of the
generic and brand name drugs are the same, then their concentration at the site of action and
therefore their safety and effectiveness will also be the same.
. For other dosage forms (e.g., drugs for inhalation, topical, or parenteral use), bioequivalence
can be demonstrated through other comparative testing (e.g., comparative pharmacodynamic
studies, pharmaceutical properties) in addition to or in view of comparative bioavailability to
support the safety and efficacy of the proposed product.

1.2.1 ACCEPTANCE CRITERIA FOR BIOEQUIVALENCE-

a) The standards for bioequivalence in different parts of the world are built upon
internationally recognized standards and criteria and are amongst the highest in the
world.

b) Health scientists ensure that the standards are adhered to and kept up to date as they
work closely with an expert panel of scientists, physicians, and pharmacists from
across the world.

c) For oral drugs, bioequivalence is determined by comparing the relative bioavailability

of the brand name drug v/s the generic drug.

d) There must be no more than a 20% difference between the AUC and Cmax of brand
name versus generic products.

e) This is based on international agreement that differences below this percentage rate
are not clinically significant.

f) In order to establish bioequivalence, the AUC and Cmax for the generic drug are
compared with that of the brand name drug.

g) Bioequivalence is based on a comparison of ratios where the ratio of generic to brand

name for each pharmacokinetic variable does not differ by more than 8:10.

h) This is how the range for the confidence intervals is defined: • 8/10 = 0.8 (gives the
lower limit of 80%) • 10/8 = 1.25 (gives the upper limit of 125%).

i) The ratio of the Cmax and the 90% confidence interval for the ratio of the AUC
should be contained within the limits of 0.8 to 1.25.

j) Thus, bioequivalence is based on ratios where the nominal equality is 1 (or 100%). It
is not based on differences in absolute values.
 k) In practice, for a generic product to demonstrate bioequivalence, the ratio of the mean
values must be close to 100% in order for the upper and lower limits to be contained
within the accepted range.

l) If the observed ratio is closer to 80% or 125%, then the data would have to contain
little or no variation from the mean for the 90% confidence intervals of the ratio to lie
within the 80% to 125% range necessary to demonstrate bioequivalence.

m) This applies to generic drugs and aforementioned situations cited for new
formulations of brand drugs.

2. BCS (Biopharmaceutical Classification

System) classification of drugs
2.1 DEFINITION-
“The Biopharmaceutical Classification System is a scientific framework for classifying a
drug substance based on its aqueous solubility & intestinal permeability & dissolution
rate”.
To save time, fast screening is required so drug substances are classified on basis of solubility
and permeability. This classification is called Biopharmaceutical Classification System.
2.2 FACTORS AFFECTING ON BIOPHARMACEUTICAL CLASSIFICATION
SYSTEM-
The Biopharmaceutical Classification System has been developed to provide a scientific
approach to allow for prediction in vivo pharmacokinetics of oral immediate release (IR)
drug product by classifying drug compound based on their:
1. SOLUBILITY
2. PERMEABILITY
3. DISSOLUTION

2.2.1 SOLUBILITY-

a) The Maximum Amount of solute dissolved in a given solvent under standard

conditions of temperature, pressure and pH.

b) Solubility is the ability of the drug to be solution after dissolution ( ).

c) The higher single unit dose is completely soluble in 250 ml at pH 1- 6.8 (37˚C).

2.2.2 PERMEABILITY-
 a) Permeability of the drug to pass the biological membrane which is the lipophilic.

b) Permeability is indirectly based on the extent of absorption of a drug substance.

c) Drug substance is considered to be highly permeable, when the extent of absorption in human
determined to be 90% or more of administered drug or compare to in vivo reference dose.

2.2.3 DISSOLUTION –

a) It is process in which solid substance solubilises in given solvent i.e mass transfer from solid
surface to liquid phase.

b) Using USP apparatus I (Basket type) at 100 rpm or USP apparatus II (Paddle type) at 50 rpm.

c) Dissolution Media [900 ml],

1. 0.1 N HCl or simulated gastric fluid (pH 1.2) without pepsin enzyme.
2. pH 4.5 acetate buffer & pH 6.8 phosphate buffer.
3. Simulated intestinal fluid without peptidase enzyme.

2.3 BCS CLASSIFICATION FOR DRUGS – EXAMPLES-

SR. CHARACTERISTICS CLASS I CLASS II CLASS III CLASS IV
NO
1) ROUTE OF Ideal for oral Suitable for Ideal for oral Poorly
ADMINISTRATION route of oral route of route for absorbed by
administration administration. administration. orally
administration
2) ABSORBANCE Drug absorbed Drug absorbed Drug absorbance Both solubility
rapidly. rapidly. is limited. & permeability
limited.

3) NATURE Amphiphilic Lipophilic Hydrophilic Riskphilic

4) THERAPEUTIC Rapid therapeutic Drug dissolves Drug dissolves Low
ACTION action. slowly. rapidly. dissolution
rate.
Slow or low
therapeutic
action.

5) BIOAVAILABILITY Bioavailability Bioavailability Bioavailability is An alternate

problem not is controlled incomplete if route of
expected for by dosage drug is not release administration
immediate release form and rate or dissolve in may be
drug product. of release of absorption needed.
the drug window.
substance.
6) CATEGORY OF Anti- NSAIDs, CNS Anti-diabetic, Diuretics,
DRUGS hypertensive, drugs, etc. anti-histamine, anticancer
adrenergic drugs, etc. drugs, etc.
etc.
7) EXAMPLES OF Metoprolol, Nifedipine, Cimetidine, Taxol,
DRUGS Propranolol, Naproxen. Metformin, Chlorthioxazol
Diltiazem. Insulin. e, Cefixime
trihydrate.

2.4 APPLICATIONS-
a) To predict in vivo performance of drug product using solubility and permeability
measurements.
b) Aid in earliest stages of drug discovery research.
c) To use in biowaiver considerations.
d) For research scientist to decide upon which drug delivery technology to follow or
develop.
e) Used also for the regulation of bioequivalence of the drug product during scale up and
post approval.
2.5 CLASS BOUNDARIES-
A] HIGHLY SOLUBLE-
The highest dose strength is soluble in < 250 ml water over a pH range of 1 to 7.5. The
volume estimate - a glassful (8 ounce).

B] HIGHLY PERMEABLE-
When the extent of absorption in humans is determined to be > 90% of an administered
dose.

C} RAPIDLY DISSOLVING-
When > 85% of the labelled amount of drug substance dissolves within 30 minutes using
USP apparatus I or II in a volume of < 900 ml buffer solutions.

3.REGULATORY REQUIREMENTS
FOR BIOEQUIVALENCE STUDY
3.1 The basic requirements for bioequivalence study are as follows-

BIOEQUIVALENCE STUDY

LINEAR STEREOCHEMISTRY IS
PHARMACOKINETICS NOT AN ISSUE

NON-NARROW NON-HIGHLY VARIABLE

THERAPEUTIC DRUG DRUG

DECISION BASED DECISION BASED ON

ON PARENT DRUG PLASMA DRUG
DATA CONCENTRATION
3.2 Regulatory requirements for BA & BE studies -
The regulatory requirements for BA/BE studies are based upon mainly two factors –
3.2.1 For IND/NDAs –
1. To establish equivalence between –
a) Early and late clinical trial formulations.
b) Formulations used in clinical trial and stability studies.
c) Clinical trial formulations and to be marketed drug product
d) Any other comparisons if appropriate

3.2.2 For ANDA –

a) ANDA for a generic drug product.
b) Change in components. compositions, manufacturing processes
c) Change in dosage forms.
3.3 Design and Conduct of studies –
The studies are mainly performed on 4 various criteria -
3.3.1 Pharmacokinetic Studies
3.3.2 Pharmacodynamic Studies
3.3.3 Comparative clinical Studies
3.3.4 Dissolution Studies

3.3.1 PHARMACOKINETIC STUDIES–

Clinical Pharmacokinetic Studies are performed to examine the absorption, distribution,
metabolism and excretion of a drug under investigation (IND) in healthy volunteers and/or
patients.

PHARMACOKINETIC STUDY

STUDY STUDY SELECTION STUDY STATISTICAL

DESIGN POPULATION CRITERIA CONDITION EVALUATION

A) Study Design
The Basic design of an in vivo bioavailability study is determined by the following:
 What are the scientific questions to be answered?
 The nature of the reference material and the dosage form to be tested.
 The availability of the analytical methods.
 Benefit-Risk ratio considerations in regards to testing in humans.

B) Study Population

1) The number of subjects to be included in the study based on an appropriate sample

size is estimated by:
 Pilot experiment
 Previous studies
 Published data
 2) The number of subjects recruited should be sufficient to allow for possible
withdrawals or removals (dropouts) from the study.
3) The significance level desired is usually 0.05.
4) Power of the study, normally 80% or more.
5) Minimum 12 subjects are required unless ethical justification.
6) If the drug product is intended for use in both males and females, the sponsor attempt
to include similar proportions of males and females in the study.

C) Selection of Subjects

1) The criteria for selection of subjects are as follows-

2) Healthy adult volunteers
3) Age: 18 yrs or older
4) Age/Sex representation corresponding to therapeutic & safety profile.
5) Non-smokers/without a history of alcohol or drug abuse.
6) Medical history/Clinical lab test values must be within normal ranges.
7) Weight within normal limits i.e (BMI 18.5-30 kg/m 2).
8) Women should be required to give assurance that they are neither pregnant, nor likely
to become pregnant until after the study.
9) Drug use intended in elders i.e the sponsor must attempt to include as many subjects
of 60 yrs of age or older or possible.
10) For teratogenic drugs mostly male volunteers are chosen.

D) Study condition

1) Selection of Blood Sampling Points/Schedules-

2) For at least three elimination half-lives (cover >80% of AUC)
 Absorption phase: 3-4 points
 Around Tmax: 3-4 points
 During elimination: 4 points
3) Intervals not longer than the half-life of the drug
4) If urine tested, collect it for at least 5 half-lives
5) The lot Numbers of both the reference listed products and the expiration date for the
reference product would be stated.
6) The drug content of the test product cannot differ from that of reference product by
more than ± 5%.
7) The sponsor can include a statement of the composition of the test product and if
possible, a side-by-side comparison of the compositions of the test and reference
product.
8) Samples of the test and reference listed product must be retained for 5 years.
E) Fasting or fed conditions

Fasting state:

a) A single dose study should be conducted after an overnight fast (at least 10 hours),
with subsequent fast of 4 hours following dosing.
b) For multiple dose fasting state studies, when an evening dose must be given, two
hours of fasting before and after the dose is considered acceptable.

Fed State:
Required when:
a) Drug recommended with food
b) Modified release product
c) Assessment of Cmax and Tmax difficult with fasting state study
d) Requires consumption of a high fat food, 30 minutes before dosing
e) Provide 800-1000 kcals with about 50% of calories derived from fat
f) Fat- 500-600, Proteins - 150, Carbohydrate- 250 Kcal
g) Ethnic & cultural variation considered
h) Specified in protocol

In general BE Study conducted under fasting condition-

 Where the SmPC (Summary of product characteristics) recommends intake of the
reference medicinal product on an empty stomach or irrespective of food intake, BE
study conducted under fasting condition.
 Where SmPC recommends intake of RMP (Risk management plan) only in fed state,
BE Study conducted under fed state.
 For specific formulation both studies are recommended, unless product taken only in
fasting or fed state.

Where information is required in both Fasting and fed state, it is acceptable to conduct either:
a. Two separate two-way cross-over.
b. Four way cross-over study.

F) Steady state/ Multiple Dose studies

a) Long elimination half life→ Accumulation in the body

b) Toxic drugs requiring multiple dose therapy
c) Some Modified-release drugs
d) Combination products
e) Drugs inducing own metabolism
f) Drugs showing non-linear pharmacokinetics
Disadvantages:
• Difficult to conduct
• Costly
• Longer monitoring
• Longer exposure to drug

3.3.1.1 Characteristics to be measured-

Evaluations of BA/BE will be based upon the measured concentrations of the active drug
substance(s) in the biological matrix.
The measurements of an active or inactive metabolite may be necessary:
a) Where the concentrations of the drug(s) may be too low to accurately measure in the
biological matrix
b) Limitations of the analytical method
c) Unstable drugs
d) Drugs with a very short half-life
e) In the case of prodrugs, racemates should be measured using an achiral assay method.
f) The plasma-time concentration curve is mostly used to assess the rate and extent of
absorption of the study drug. This include Cmax, Tmax, AUC0-t and AUC0-∞.

Pharmacokinetic Parameters measured are:

• Plasma conc. and time points
• Subject, period, sequence, treatment
• Cmax, Tmax, AUC0-t, AUC0-∞
• Partial AUC only for early exposure of drug content.

For steady state studies:

• Swing
• Cav (Avg. concentration)
• Cmin
• Degree of fluctuation

Measurement of Individual enantiomers in BE Studies is recommended if:

1. Exhibit different Pharmacodynamic characteristics
2. Exhibit different Pharmacokinetics characteristics
3. Primary efficacy and safety activity resides with minor enantiomers
4. Nonlinear absorption is present
3.3.1.2 Statistical Evaluation-
a) Primary concern of bioequivalence is to limit Consumer’s & Manufacturer’s risk.
b) Cmax & AUC analysed using ANOVA (Analysis of variance).
c) ANOVA is an analysis tool used in statistics that splits an observed aggregate
variability found inside a data set into 2 parts: Systematic factors and Random factors.
d) The systematic factors have a statistical influence on the given data set, while the
random factors do not.
e) Tmax analysed by non-parametric methods.
f) Use natural log transformation of Cmax and AUC.
g) Calculate 90% confidence interval for this GMR for Cmax.
h) Similarly calculate GMR (Geometric Mean Ratio) for AUC.
Criteria for bioequivalence-
To establish Bioequivalence, the calculated 90% confidence interval for AUC and Cmax
should fall within the bioequivalence range, usually 80-125%.
Tighter limits for permissible differences in bioavailability may be required for drugs that
have:
• A narrow therapeutic index.
• A serious, dose-related toxicity.
• A steep dose/effect curve,
• A non-linear pharmacokinetics within the therapeutic dose range.

3.3.1.2.1 Two-Period Crossover Design-

2 formulations (1 standard and 1 control), even number of subjects, randomly divided
into 2 equal groups
a) First period, each member of one group receives a single dose of the test formulation;
each member of the other group receives the standard formulation.
3.3.1.2.2 Latin Square Design-
a) Includes more than 2 formulations.
b) A group of volunteers will receive formulations in the sequence as shown below.

3.3.1.2.3 Balance Incomplete Block Design (BIBD)-

a) Includes more than 3 formulations, latin square design will not be ethically advisable
because each volunteer may require drawing of too many blood samples.
b) If each volunteer expected to receive at least two formulations, then such a study can
be carried out using BIBD.
c) Conditions required for conducting BIBD are as follows-
• If all treatment comparisons are of equal importance and we wish to estimate
each treatment effect with the same precision, then the design needs to be
balanced
• Each block contains the same no. of units.
• Each treatment occurs the same no. of times in total.
• Each pair of treatments occurs together the same no. of times in total.
3.3.1.2.4 Parallel-Group Design-
a) Requires even number of subjects in two groups.
b) Each receive a different formulation.
c) No washout necessary.
d) Washout period is the time frame allotted for an administered drug to be eliminated
from the body or for a previously administered intervention to become ineffective.
e) Mainly suitable for drugs with longer half life.

3.3.1.2.5 Replicate Crossover-Study Design-

a) Mostly used for highly variable drugs (showing poor mode of action).
b) Allows comparisons of within-subject variances.
c) Reduce the number of subjects needed.
d) Four-period, two-sequence, two-formulation design is recommended or three-
sequence, three-period, single-dose, partially replicated.
3.3.1.2.6 Pilot Study-
a) If the sponsor chooses, this is performed in a small number of subjects.
b) To assess variability, optimize sample collection time intervals & provide other
information, validate analytical methodology.
Example:
 Immediate-release products: careful timing of initial samples→ avoid a subsequent
finding that the first sample collection, occured after the plasma concentration peak.
 Can be appropriate, provided its design & execution are suitable & sufficient number
of subjects have completed the study.

3.3.1.3 Biowaivers-
A Biowaiver means that in vivo bioavailability and/or bioequivalence studies may be waived
(not considered necessary for product approval/ exempted).
In vitro studies, i.e dissolution studies can be used in place of in vivo bioequivalence under
certain conditions, called BIOWAIVERS.
The drug product differs only in strength of active substance, provided the following
condition hold:
a) Pharmacokinetics is linear.
b) The qualitative composition is same.
c) The ratio between active substance and excipient is same.
d) Both products are produced by same manufacturer at same site with same
manufacturing process.
The drug has been slightly reformulated or manufacturing method has been slightly modified
by same manufacturer in ways that can be argues irrelevant for BA.
The product meets following requirement:
a) The product is in form of solution or solubilised form (elixir, syrup) etc.
b) The product contains active ingredient in same conc. as approved drug.
c) The product contains no excipient known to significantly affect absorption of active
ingredient.
d) The product is administered by inhalation as gas or vapor.
e) The product is for oral administration but not intended for absorption (antacid or radio
opaque medium).
f) The product is intended for topical administration (ointment, creams, gels etc,) for
local effect.
3.3.2 PHARMACODYNAMIC STUDIES-
These studies include measurement of effect on a Patho-physiological process as a function
of time, after administration of 2 different products
Necessity-
1. Quantitative analysis in plasma or urine is not possible with sufficient accuracy &
sensitivity
2. Drug concentrations are not surrogate endpoints e.g Topical formulations without
systemic absorption.
3. In situations of ‘Superiority Claims’.
4. In case only Pharmacodynamic data is collected→ other methods tried & why they
were unsuitable should be mentioned.

3.3.3 COMPARATIVE CLINICAL STUDIES-

Comparative clinical study means the controlled study of product by the administration of
product to human beings where the principal purpose of such studies is to provide
confirmatory/comparative study, efficacy and immunogenicity data necessary to support the
filing for regulatory approval of product.
Necessity-
1. Both pharmacokinetic & pharmacodynamic parameters not properly measurable or
not feasible.
2. Mention which methods were tried & found unsuitable.

Following critical points need to be defined in advance on case to case basis-

1. Clinical end points (Target parameters) → intensity & onset of response.
2. Size of equivalence range→ case-to-case basis (depending on natural course of
disease, efficacy of available treatments, target parameter).
3. Statistical confidence interval.
4. Placebo must be included when appropriate.
5. Safety end-points in some cases.

3.3.4 DISSOLUTION STUDIES-

1. Dissolution is a process in which solid substance solubilizes in a given solvent i.e
mass transfer from the solid surface to the liquid phase.
2. Suitable to confirm unchanged product quality with minor changes in formulation /
manufacturing after approval→ SUPAC (Scale-Up & Post- Approval Changes).
3. Different strengths of drug manufactured by same manufacturer where:
 Qualitative composition is same.
 Ratio of active ingredients & excipients is same
 Method of manufacture is same
  BE study has been performed on 1 strength
 Linear pharmacokinetics
 Assess batch-to-batch quality
 More than 1 batch of each formulation tested
Design should include:
a) Individually testing of at least 12 dosage units of each batch →Mean & Individual
results with Sd or SE.
b) Measurement of percentage of content released at suitably spaced time points eg. At
10, 20 & 30 mins or appropriate for complete dissolution).
c) Dissolution profile in atleast 3 aqueous media with pH range of 1.0-6.8 Or 1.0-8.0
wherever necessary.
d) Conduct tests on each batch with same apparatus & on same or consecutive days.

Dissolution testing should be carried out in:

1) USP Apparatus I at 100 rpm or Apparatus II at 50 rpm using 900 ml of the following
dissolution media:
2) 0.1N HCl or Simulated Gastric Fluid USP without pepsin enzymes.
3) A pH 4.5 acetate buffer.
4) A pH 6.8 phosphate buffer or Simulated Intestinal Fluid USP without peptidase
enzymes.
5) For capsules and tablets with gelatin coating- Simulated Gastric and Intestinal Fluids
USP (with enzymes) can be used.

3.3.4.1 COMPARISON OF DISSOLUTION PROFILE BY DIFFERENT

METHODS & IVIVC (In-Vitro and In-Vivo Correlation)-

It is graphical representation [in terms of concentration vs. time] of complete release of A.P.I.
from a dosage form in an appropriate selected dissolution medium. i.e. in short it is the
measure of the release of A.P.I from a dosage form with respect to time.

3.3.4.1.1 Objective-

a) To Develop in vitro-in vivo correlation which can help to reduced costs, speed-up
product development and reduced the need of perform costly bioavailability human
volunteer studies.
b) Demonstrating equivalence after change in formulation of drug product.
c) Establish the similarity of pharmaceutical dosage forms, for which composition,
manufacture site, scale of manufacture, manufacture process and/or equipment may
have changed within defined limits.

3.3.4.1.2 Importance-

a) Dissolution profile of an A.P.I. reflects its release pattern under the selected condition
sets. i.e. either sustained release or immediate release of the formulated formulas.
 b) For optimizing the dosage formula by comparing the dissolution profiles of various
formulas of the same A.P.I.
c) The most important application of the dissolution profile is that by knowing the
dissolution profile of particular product of the BRAND LEADER, we can make
appropriate necessary change in our formulation to achieve the same profile of the
BRAND LEADER.

3.3.4.1.3 METHODS TO COMPARE DISSOLUTION PROFILE-

3.3.4.1.3.1 Graphical method-

a) In this method we plot graph of Time V/S concentration of solute (drug) in the
dissolution medium or biological fluid.
b) The shape of two curves is compared for comparison of dissolution pattern and the
concentration of drug at each point is compared for extent of dissolution.
c) If two or more curves are overlapping then the dissolution profile is comparable.
d) If difference is small then it is acceptable but higher differences indicate that the
dissolution profile is not comparable.
3.3.4.1.3.2 Model dependent methods-

A) Zero order kinetics (osmotic system, transdermal system)-

Zero order API release contributes drug release from dosage form that is independent of
amount of drug in delivery system. i.e constant drug release

A0 - At = Kt
where ,
A0 = Initial amount of drug in the dosage form.
At = Amount of drug in the dosage form at time ‘t’.
Kt = Proportionality constant.

Application:
Transdermal systems as well as matrix tablets with low solubility drugs in coated forms,
osmotic systems, etc.

B) First order kinetics (Water soluble drugs in porous matrix)-

Using Noyes Whitney’s equation, the rate of loss of drug from dosage form (-dA/dt) is
expressed as;
-dA/dt = k (Xs– X)
Assuming that,
sink conditions = dissolution rate limiting step for in-vitro study
absorption = dissolution rate limiting step for in-vivo study.
Then (1) turns to be:
-dA/dt = k(Xs ) = constant
So it becomes,
A = Ao × e-k

Applications:
This relationship can be used to described the drug dissolution in pharmaceutical dosage
forms containing water soluble drugs in porous matrices.

C) Higuchi Model (Diffusion matrix formulation)-

Higuchi in 1961 developed models to study the release of water soluble and low soluble
drugs incorporated in semisolid and solid matrices.
To study the dissolution from a planer system having a homogeneous matrix the relation
obtained was,
A = [D.(2C – Cs).Cs × t]1/2
where,
A is the amount of drug released in time ‘t’ per unit area,
C is the initial drug concentration,
Cs is the drug solubility in the matrix media
D is the diffusivity of drug molecules in the matrix substance.

Applications:
Modified release dosage forms, transdermal systems and matrix tablets with water soluble
drugs.
D) Baker-Lonsdale model (Microspheres, microcapsules)-
In 1974 Baker-Lonsdale (Baker and Lonsdale, 1974) developed the model from the Higuchi
model and describes the controlled release of drug from a spherical matrix that can be
represented as:
3/2 [1-(1-At/A∞)2/3]-At/A∞ = (3DmCms) / (r02C0) t
where,
At is the amount of drug released at time ’t’
A∞ is the amount of drug released at an infinite time,
Dm is the diffusion coefficient,
Cms is the drug solubility in the matrix,
ro is the radius of the spherical matrix
Co is the initial concentration of the drug in the matrix.

E) Hixon- Crowell model (Erodible matrix formulation)-

To evaluate the drug release with changes in the surface area and the diameter of the particles
/tablets.
The rate of dissolution depends on the surface of solvent - the larger is area the faster is
dissolution.
Hixon-Crowell in 1931 (Hixon and Crowell, 1931) recognized that the particle regular area is
proportional to the cubic root of its volume, designed an equation as,
Wo1/3-W1/3 = Kt
where,
Wo= original mass of API particles
K = cube-root dissolution rate constant
W = mass of the API at the time ‘t’.
This model is called as “Root law”.

3.3.4.1.3.3 Model independent methods-

3.3.4.1.3.3.1 Difference factor (f1) and Similarity factor (f2)-

A) The difference factor (f1)-

The difference factor as defined by FDA calculates the % difference between 2 curves at each
time point and is a measurement of the relative error between 2 curves.

where,
n= no. of time points
Rt= % dissolved at time (t) of reference product (pre-changes)
Tt= % dissolved at time (t) of test product (post-changes)
B) The similarity factor (f2)-
The similarity factor (f2) as defined by FDA is logarithmic reciprocal square root
transformation of sum of squared error and is a measurement of the similarity in the
percentage (%) dissolution between the two curves.

3.3.4.1.3.3.2 Limits for similarity and difference factors-

DIFFERENCE FACTOR SIMILARITY FACTOR INFERENCE
(f1) (f2)
0 100 Dissolution profile are
similar
< 15 > 50 Similarity or equivalence of
2 profiles

3.3.4.1.3.3.3 Advantages-
a) They are easy to produce
b) They provide single number to describe the comparison of dissolution profile data

3.3.4.1.3.3.4 Disadvantages-

a) The values of f1 and f2 are sensitive to the number of dissolution time point used.
b) If the test and reference formulation are interchanged, f2 is unchanged but f1 is not,
yet difference between two mean profiles remains same.
c) The basis of criteria for deciding the difference or similarity between dissolution
profile is unclear.

3.3.4.1.3.3.5 The evaluation of similarity between dissolution profile is based on

following conditions-

a) Minimum of three dissolution time points are measured

b) Number of drug products tested for dissolution is 12 for both test and reference
c) Not more than one mean value of >85% dissolved for each product
d) Standard deviation of mean of any product should not be more than 10% from 2nd to
last dissolution time point

3.4 DOCUMENTATION-
With respect to the conduct of bioequivalence/bioavailability studies following important
documents must be maintained:

A) Clinical Data:
All relevant documents as required to be maintained for compliance with GCP Guidelines.

B) Details of the analytical method validation including the following:

 System suitability test-
a) System suitability test is to prove that system is working perfectly before the
analysis on HPLC, GC, TOC (Total organic carbon) analyser or any other system.
b) It is required to be done before every sample analysis.
c) In the HPLC technique, a liquid sample is passed over an absorbent material to
test its efficacy.

 Linearity range-
The linearity of an analytical method is its capability to elicit check consequences
which might be at once, or with the aid of well described mathematical adjustments,
proportional to the concentration of analytes in within a given range.

 Lowest limit of quantitation-

The quantitation limit of an individual analytical procedure is the lowest amount of
analyte in the sample which can be quantitatively determined with suitable precision
and accuracy.

 QC sample analysis

 Stability sample analysis-

a) During a stability study, materials are stored at various temperature and humidity
conditions and samples are pulled at predetermined time points and subjected to a
battery of tests that may include: an identification test, assay, physical tests,
microbiological limits, and preservative effectiveness testing, using appropriately
validated methods and/or recognized compendial methods.
b) Acceptance criteria are stipulated before initialization of the study, and if a product
fails to meet specifications at any time point, the stability study may be halted and
restarted after reformulation or other modifications have occurred.

 Recovery experiment result-

a) The recovery is the ratio of the concentration of analyte found to that stated to be
present.
b) Results obtained on test materials of the same matrix could be corrected for recovery
on the basis of recovery found for the reference material.
c) Recovery studies are a classical technique for validating the performance of an
analytical method.
d) However their use in clinical laboratories has been fraught with problems due to
improper performance of the experiment, improper calculation of the data, and
improper interpretation of the results.

C) Analytical data of volunteer plasma samples which should include the following:
  Validation data of analytical methods used,
 Chromatograms of all volunteers,
 Inter-day and intra-day variation of assay results,
 Details including chromatograms of any repeat analysis performed,
 Calibration status of the instruments.

D) Raw data

E) All comments of the chief investigator regarding the data of the study submitted for
review

F) A copy of the final report