The document discusses the design and requirements of bioequivalence studies, essential in demonstrating that a new drug product performs similarly to an approved one. It outlines types of equivalences (chemical, pharmaceutical, bioequivalence, and therapeutic) and details both in vivo and in vitro study methodologies along with their protocols. Additionally, it highlights the challenges in establishing bioequivalence for highly variable drugs and the importance of adhering to regulatory guidelines during study design.

1. BIOEQUIVALENCE STUDIES &
DESIGN
Submitted by:-
Shiva Kant Thakur
M. Pharm 1st year ( pharmaceutics)
Submitted To :-
Dr .Tejpal Yadav
Assistance professor(pharmaceutics)

2. CONTENT
 Definition
 Requirement /objectives of BIOEQUIVALENCE
 Type of bioequivalence
I. in vivo
II. In vitro
 Study designs

3. Definition
 It refers to the drug substance in two or more identical
dosage forms , reaches systemic circulation at he same
rate and to the same relative extent .
 Their plasma concentration –time profile will be identical
without significant statistical differences.

4. Advantages & Disadvantages
 Minimizes the effect of inter
subject variability.
 It minimizes the carry over effect .
 Requires less number of subjects to
get meaningful results
 Requires longer time to time to
complete the studies.
 Completion of studies depends of
formulations evaluated in the
studies.
 Increase in study period leads to
high subject
 Medical ethics does not allow too
many trials on subject
continuously for a longer time.

5. Objectives
 If a new product is intended to be a substitute for an
approved medicinal product as a pharmaceutical equivalent
or alternative, the equivalence with this product should be
shown or justified .
 In order to ensure clinical performance of such drug
products, bioequivalence studies should be performed.
 Bioequivalence studies are conducted if there is:
 A risk of pharmacotherapeutic failure or diminished clinical
safety.

6. Some of the important terms
 Equivalence :- it is a relative term that compares drug
products with respect to a specific characteristic or function
or to a defined set of standards.
 These are several types of equivalences.
A. Chemical equivalence
B. Pharmaceutical equivalence
C. Bioequivalence
D. Therapeutic equivalence

7. Types of equivalences
A. Chemical Equivalence:- it indicates that two or more drug
products contain the same labelled substance as an active
ingredient in the same amount .
B. Pharmaceutical Equivalence :-the term implies that two or
more drug products are identical in strength , quality ,purity
,content uniformity and disintegration and dissolution
characteristics .they may, however, differ in containing
different excipients.

8. Types of equivalence
C . Bioequivalence :- it is a relative term which denotes that the
drug substance in two or more identical dosage forms, reaches the
systemic circulation at the same relative rate and to the same
relative extent i.e. their plasma concentration –time profiles will be
identical without significant statistical differences..
 When statistically significant differences are observed in the
bioavailability of two or more drug products, bio -inequivalence
is indicated.
D . Therapeutic :- this term indicates that two or more drug
products that contain the same therapeutically active ingredient
elicit identical pharmacological effects and can control the disease
to the same extent.

9. Types of bioequivalence studies
Bioequivalence can be demonstrated either:-
o In-vivo
o In -vitro

10. In vivo bioequivalence studies
 The following sequence of criteria is useful in assessing the
need for in vivo studies:
1. Oral immediate – release products with systemic action-
 Indicated for serious conditions requiring assured response.
 Narrow therapeutic margin.
 Pharmacokinetics complicated by absorption < 70% or
absorption window ,nonlinear kinetics ,presystemic
elimination>70%.
 Unfavorable physiochemical properties ,e.g.low solubility,
metastable modification ,instability , etc.

11. In vivo bioequivalence studies
 Documented evidence for bioavailability problems.
 No relevant data available , unless justification by applicant
that in vivo study is not necessary .
2. Non –oral immediate – release products.
3. Modified – release products with systemic action .
In vivo bioequivalence studies are conducted in the usual
manner as discussed for bioavailability studies, i.e. the
pharmacokinetic and the pharmacodynamics methods.

12. 1. Pharmacokinetic methods
a) Plasma level – time studies
b) Urinary excretion studies
2. Pharmacodynamics methods
a) Acute pharmacological response
b) Therapeutic response

13. In vitro bioequivalence studies
If none of above criteria is applicable, comparative in
vivo dissolution studies will suffice.
In vitro studies, i.e. dissolution studies can be used in lieu
of in-vivo bioequivalence under certain circumstances ,
called as bio waivers (exemption).
1. The drug product differs only in strength of the active
substance it contains, provided all the following
conditions hold-
 Pharmacokinetics are linear.
 The qualitative composition is the same .

14. In vitro bioequivalence studies
 The ratio between active substance and the excipient is the same ,
or(in the case of small strengths) the ratio between the excipients is
the same.
 Both products are produced by the same manufacture at the same
production site.
 A bioavailability or bioequivalence study has been performed with a
original product.
 Under the same test conditions, the in vitro dissolution rate is the
same.
2. The manufacturing method has been slightly modified by
the original manufacture in ways that can convincingly be
argued to be irrelevant for the bioavailability.

15. In vitro bioequivalence studies
3. The drug product meets all of the following requirements –
 The product is in the form of solution or solubilized
from(elixir, syrup, tincture, etc).
 The product contains active ingredient in the same
concentration as the approved drug product.
 The product contains no excipients known to significantly
affect absorption of the active ingredient

16. In vitro bioequivalence studies
4.The in vitro dissolution rate of the new product is
equivalent with that of the already approved medicinal
product.
 The product is intended for topical administration (
cream, ointment ,gel , etc.) for local effect .
 The product is for oral administration but not intended to
be absorbed ( antacid or radio- opaque medium ).
 The product is administered by inhalation as a gas vapor .
 The criteria for drug products listed above indicate that
bioavailability and bioequivalence are self-evident.

17. Bioequivalence protocol
1. Experimental design
2. Wash out period
3. Drug products:
(a) Test product
(b) Recognized standard
4. Route of administration
5. Dosage regimen
6. Frequency and duration of sampling
7. Randomization of drug administration
8. Single-versus multiple-dose study design

18. Bioequivalence protocol
9. Subjects
(a) Healthy subjects versus patients
(b) Subject selection
(i) medical history,
(ii) Physical examination,
(iii) Laboratory tests
(c) Study conditions
10. Analysis of biological fluids

19. Bioequivalence protocol
C. Methods of Assessment of Bioavailability
1. Plasma data
2. Urine data
3. Acute pharmacological effect
4. Clinical response
D. Analysis and Presentation of Data
1. 1. Statistical treatment of data-Analysis of variance (AN OVA)
2. 2. Format of data

20. Bioequivalence design study
The FDA provides the guidance for the performance of in
vitro dissolution and in vivo bioequivalence studies which
include ( solid oral dosage form):
1. Fasting study
2. Food intervention study
3. Crossover study designs
4. Sprinkle BE study(extended release capsules having
beads)

21. 1. Fasting study
a. The study is required for all immediate and modified
release oral dosage forms.
b. Both male and female subjects are included.
c. Overnight fasting is required (at least 10 hrs).
d. After admn. Of drug fasting continued up to 4 more hours.
e. Blood sampling is performed before dose and at diff.
intervals after dose
f. Plasma drug concentration –time profile is obtained.
g. No other medication given at least 1 week prior to study.

22. 2. Food intervention study
1. It uses single dose, randomized ,2 treatment , 2period crossover study.
2. Conducted using meal conditions that have greatest effect on GI
physiology.
3. Meal containing high calories (50% of total caloric content) and fat (800-
1000cal)is taken.
4. After a overnight fast of 10hrs , meal is given 30min prior to dosing.
5. The meal is consumed over 30min with admn. of drug(with 240ml of
water) immediately after meal.
6. No food is allowed 4hrs after dosing.
7. Study on drug like ibuprofen and naproxen which is affected by food.

23. Crossover study designs
 Latine –square crossover design for a bioequivalence study of three
drug products in six human volunteers .
Subject Study period1 Study period2 Study period3
1 A B C
2 B C A
3 C A B
4 A C B
5 C B A
6 B A C

24. Crossover study designs
Latine –square crossover design for a bioequivalence study of four drug
products in 16 human volunteers .

25. Crossover study designs
Subject Study period1 Study period2 Study period3 Study period4
1 A C C D
2 B A D A
3 C D A B
4 D B B C
5 A C D C
6 C D C A
7 B B A B
8 A D B D
9 C C B D
10 B A D A
11 D B A C
12 A A C B
13 C C D A
14 D D B C
15 C B A D
16 B A C A

26. Crossover study designs
Period 1 Period 2
Sequence 1 T R
Sequence 2 R T
Period refers to the time period in which a study is performed.
A two-period study is a study that is performed on two different days (time periods)
separated by a washout period during which most of the drug is eliminated from the body-
generally about 10 elimination half-lives.
A sequence refers to the number of different orders in the treatment groups in a study. For
example, a two-sequence, two-period study would be designed as above: where R=
reference and T = treatment.
The same reference and the same test are each given twice to the same subject. Other
sequences are possible. In this design, Reference-to-Reference and Test-to-Test comparisons
may also be made.

27. Replicated crossover study designs
1. When the no. of study subjects <80, it is difficult to achieve with highly
variable drugs and drug product (%CV> 30).
2. These drugs have wide therapeutic window and despite high variability,
have been demonstrated to be safe and effective.
3. Replicate designs for these drugs require smaller no. of subject and avoid
exposure of large no. of healthy subjects.
4. Used for determining individual BE, to estimate within subject variance
for both test and reference.
5. Provide an estimate of the subject-by- formulation interaction variance.

28. Replicated crossover study designs
Period 1 Period 2 Period 3 Period4
Sequence 1 T R T R
Sequence 2 R T R T
A four , two sequence ,two formulation design is recommended by FDA
Where R= reference and T = treatment
The same reference and the same test are each given twice to the same
subject .other sequence are possible .
In this design ,Reference –to reference and test to test comparisons may also
be made.

29. Sealed average bioequivalence
1. 3 sequence, 3 period, 2 treatment partially replicated crossover design.
2. This design allows the estimation of within -subject variance and subject-
by- formulation interaction for reference product.
3. Completion time of this study is shorter than fully replicated four way
crossover.
4. If the test has lower variability than reference product, the study will need
smaller no. of subjects.
5. This is evaluated for both AUC and Cmax.

30. Sealed average bioequivalence
Period 1 Period 2 Period 3
Sequence 1 T R R
Sequence 2 R T R
Sequence 2 R R T

31. Non replicate parallel study design
1. For the drugs having long elimination half life or depot injection in
which the drug is slowly released over weeks and month.
2. Two separate groups of volunteers are used.
3. One group will have the test product while the other will have the
reference product.
4. Blood sample collection time should be adequate to ensure completion
of GI transit(2-3days).
5. Cmax and AUC, 72 hrs after dose admn. can be used to characterize
peak and total drug exposure.
6. This design is not for drugs that have high intra-subject variability in
distribution and clearance.

32. 4.Multiple dose (steady state)
1. Multiple doses of same drug are given consecutively to reach steady
state plasma drug levels.
2. The multiple dose study is designed as steady state, randomized, 2
treatment ,2 way crossover study comparing equal dose of test
reference.
3. To ascertain that the subjects are at steady state, three consecutive
trough concentrations(Cmin) are determined.
Pharmacokinetic analyses include calculation of following parameters
for each subject:
AUC Area under the curve during a dosing intervals.
tmax Time to Cmax during interval.

33. Multiple dose (steady state)
Cmax Maximum drug concentration during dosing interval
Cmin Drug concentration at end of a dosing interval.
Cavg The average drug concentration during a dosing interval
Degree of fluctuation = (Cmax- Cmin)/Cmax ;
Swing =( Cmax – Cmin)/Cmin

34. Clinical endpoint BE study
1. This consists of randomized double-blind ,placebo –
controlled , parallel – designed study comparing test
product , reference product, and placebo in patients.
2. The primary analysis for bioequivalence is determined by
evaluating the difference between the proportion of patients
in the test and reference treatment groups who are
considered a “ therapeutics” at the end of study.
3. The superiority of the test and reference products against
the placebo is also tested during the same dichotomous end
point of “therapeutic cure”.

35. Special Concerns In Bioavailability
And Bioequivalence Studies.
1. For certain drug and dosage forms , systemic bioavailability and
bioequivalence are difficult to ascertain for eg. Cyclosporine
,verapamil, are considered to be highly variable.
2. The number of subjects required to demonstrate bioequivalence for
these drug products may be excessive, requiring more than 60
subjects.
3. The intrasubject variability may be due to the itself or to the drug
formulation or to both.
4. The FDA has held public forums to determine whether the current
bioequivalence guidelines need to be changed for these highly
variable drugs.

36. Thank You