1. Presentation on : Evaluation of Pharmaceutical Formulation in vitro - in vivo
Department of Pharmaceutics , School Of Pharmacy .
B.I.T. By Pass Road Partapur Meerut
Presented to : Presented By :
Dr. Upendra Nagaich Promila Sharan
Associate Professor M.Pharma -1 sem
Pharmaceutics (Pharmaceutics)

2. Evaluation
of
Pharmaceutical Formulation
in vitro in vivo

3. CONTENTS :
 INTRODUCTION
 IN VITRO DISSOLUTION STUDIES
 OFFICIAL DISSOLUTION APPARATUS
 DISSOLUTION PARAMETER
 IN VITRO EVALUATION
 SOLID
 LIQUID
 SEMI SOLID
 FACTOR AFFECTING DISSOLUTION
 BIOAVAILABILITY
 MEASUREMENT OF BIOAVAILABILITY
 PHARMACOKINETIC PARAMETERS

4. INTRODUCTION
The physicochemical property of most drugs that has greatest influence on their ADME characteristics from
the GIT is dissolution rate. The best way to determine the therapeutic efficacy of drug with a slow
dissolution rate is in-vivo determination of bioavailability of which is usually done for any formulation. The
best available tool today which can at least quantitatively assure about the biological availability of a drug
from its formulation is its in vitro dissolution test . IVIVC is the link between in vitro and in vivo
performance of the drug product[B].It has wide application in drug delivery at various stages of
development
to setting dissolution specifications. The IV-IV studies enables dissolution test to serves as a surrogate for
drug bioavailability studies in human beings.

5. SOLID
DOSAGE
DOSAGE FORMS
FORM
LIQUID
DOSAGE
SEMISOL FORMS
ID
DOSAGE
FORMS

6. IN VITRO DISSOLUTION STUDIES
DISSOLUTION APPRATUS[IVDT]

7. IN VITRO (DISSOLUTION STUDIES)
Dissolution testing is a requirement for all solid oral dosage forms and is used in all phases of
development for product release and stability testing. It is a key analytical test used for detecting
physical changes in an active pharmaceutical ingredient (API) and in the formulated product.[IVD]
The specific dissolution technique employed is determined by the dosage form characteristics and the
intended route of administration. For solid dosage forms, industry standard dissolution testing
methodologies are the United States Pharmacopoeia (USP) Apparatus:
 Apparatus 1 (basket) [Floating capsules and tablets ]
 USP Apparatus 2 (paddle) [Immediate-release, modified-release and extended release tablets].
Other dissolution techniques and equipment include
 USP 3 (reciprocating cylinders)
 USP 4 (flow-through-cell)
 USP 5 (paddle-over-disk)
 USP 6 (cylinder)
 USP 7 (reciprocating holders).

8. OFFICIAL USP DISSOLUTION
APPARATUS[B]
ROTATING BASKET ROTATING PADDEL
APPARATUS APPARATUS
RECIPROCATING
CYLINDER FLOW THROUGH
APPRATUS CELL APPARATUS
CYLLINDER
PADDEL OVER APPARATUS
DISC APPARATUS
APPARATUS 7 RECIPROCATING DISC APPARATUS

9. FACTORS AFFECTING DISSOLUTION
DISSOLUTION PARAMETER
USP APPARATUS 1 AND 2 .
•Acid (HCl 0.1 – 0.001 N)
MEDIA • Buffers: Acetate (pH 4.1 – 5.5, 0.05 M), Phosphate (pH 5.8 – 8.0, 0.05 M)
• Simulated Fluid: Gastric Fed and Fasted, Intestinal Fed and Fasted
• 900 mL, 500 mL (for low dosage strengths)
MEDIA VOLUME • 1000 mL, 2 L or 4 L (for increased sink)
• 200 mL or smaller volumes (as justified)
Paddle
SPEED • 50 rpm (preferred for BCS[Bio pharmaceutics classification system])
• 75 rpm (to eliminate coning/variability)
• 25 rpm (for suspensions)
• 100 rpm (needs justification for IR, common for ER)
Basket
• 50 - 100 rpm
• 37 °C ± 0.5 °C.
TEMPERATURE

10. As in one of my previous slide I have mentioned about three main
dosage forms:
SOLID DOSAGE FORMS
 TABLET
 CAPSULES
LIQUID
 SYRUPS
 SUSPENSIONS
 EMULSIONS
SEMI SOLID DOSAGE FORMS
 GELS
 OINTMENTS

11. In Vitro evaluation for various dosage forms
As in one of my previous slide I have mentioned about three main dosage forms:
TABLETS and CAPSULES –
Basically for both these dosage forms we use USP type 1&2 dissolution apparatus. Where the
tablet or the capsule is introduced to the basket containing media (900ml as per USP) and
temperature is maintained at 37+/- 0.5 degree C. Speed is selected (50 rpm) as per BCS. Time
interval is selected for the withdraw of the sample , the amount of sample is withdrawn the same
quantity of fresh media is added to maintain the volume and the absorbance is taken by UV
Spectrophotometer. The concentration of the drug mg/ml or ug/ml is found and using these
readings drug content release is found at various time interval and the time of maximum release
is determined.

12. Solid dosage forms non ionic drugs
On disintegration drug in solution at
the absorption site non ionic drugs
Granules
/aggregates dissolution
De-aggregation ionic drug
Fine particles ………….
……………
ionic drugs
Solid disintegration solid dissolution drug in permeation across Drug in
dosage drug solution at body
form de-aggregation particle absorption bio-membrane
site
RDS
for lipophilic drugs
RDS for Hydrophilic drugs

13. DISSOLUTION OF LIQUID DOSAGE FORMS :
Similar to the tablets and capsules The USP 2 (paddle) has been used at a
rotation speed between 25-50 rpm and as this apparatus has gained acceptance
for suspension because it provides mild laminar liquid agitation , and it also
functions as an in situ non blocking filter. Sufficient media of dissolution media
should be using to maintain the sink condition (about 900-1000) , and
temperature of 37’C should be maintained and at particular time intervals and
absorbance is taken and drug release is found

14. DISSOLUTION OF SEMI SOLID DOSAGE FORMS.( USP DISSOLUTION APPARATUS 6)
The in vitro dissolution of the semi solid preparations is based on an open
chamber
diffusion cell system e.g. Franz diffusion cell or the Modified diffusion cell
apparatus
fitted with a synthetic semi permeable membrane . The test product is placed on
the upper side of the membrane inside the donor compartment of the diffusion
cell
and a sampling fluid is placed on other side of the membrane
in the receptor compartment diffusion of drug from semi solid topical product to
and across the membrane is monitored by assay of sequentially withdrawn
sample

15. SCHEMATIC REPRESENTATION OF MODIFIED DIFFUSION
CELL APPARATUS FOR SEMI SOLID PREPARATIONS
Diagram:
DONOR
COMPARTMENT
JACKETED WATER
BATH
RECEPTOR
COMPARTMENT
.1 N HCl
SEMI
PERMEABLE
MEMBRANE
MAGNETIC
STIRRER

16. In previous discussed in vitro evaluation of solid , liquid and semi solid dosage forms we
have obtained the various drug release profile for the dosage forms. By studying this
release profile we can found the time at which the rate of release is maximum and an
arbitrary value for Cmax and tmax is found.
FACTOR AFFECTING RATE OF DISSOLUTION :
Particle size
Polymorphism
Hydrates/Solvates(pseudo polymorphs)
Salt form of drug
Drug pKa and Lipophilicity and GI pH-pH partition hypothesis
Lipophilicity and drug absorption
Drug stability

17. In vivo evaluation of the Pharmaceutical Dosage forms
IN VIVO STUDIES : This is a Latin word which it self means in life are the studies done to determine the
bioavailability of any drug/ dosage form on the required animal model.
BIOAVILABILITY: It is defined as the rate and extent (amount)of absorption of unchanged drug from its
dosage form to the systemic circulation.
BIOAVAILABLE FRACTION(F) = Bioavailable dose/Administered dose.
bioavailability
ABSOLUTE
RELATIVE
BIOAVAILABILI
BIOAVAILABILI
TY
TY (Fr)
(F)
Absolute bioavailability : It is done when systemic availability of a drug administered orally is determined
in
comparison to its intra venous administration.
F=[AUC]oral Div
[AUC]iv Doral ……………..(1)

18. Relative bioavailability:
When the systemic availability of the drug after oral administration is compared with that of an oral standard of the
same drug is referred to as relative or comparative bioavailability.
Fr=[AUC]test D std
C
[AUC]std D iv Cmaxa
AUC
Plasma
drug
Conc.
tmax
time
This curve represent the bioavailability of the drug in the form of AUC i.e. the maximum drug release
Cmax is obtained at t max in the systemic circulation.

19. MEASUREMENT OF BIOAVAILABILITY
BIOAVAILABILITY
PHARMACOKINET PHARMACODYNAMI
IC METHODES C METHODES
Urinary
Plasma level- Acute
excretion Therapeutic
time studies pharmacologica
studies response
l response
C
max
(dXu/dt)
max
t
max (tu)max
AUC Xu(infinite)

20. PHARMACOKINETIC METHODES:
These are the methods based on assumptions that the pharmacokinetic profile reflect
the therapeutic effectiveness of a drug hence also called indirect methods:
1-Plasma level-time studies :
Done on animal model.
More reliable method than urinary excretion rate.
After drug administration blood sample is withdrawn and concentration of
drug required to show the therapeutic response at any time is determined.

21. Plasma level-time curve
C max
The peak plasma concentration that gives an
indication whether the drug is sufficiently
absorbed systemically to provide therapeutic
response.
t max
It is the peak time that gives an indication of the
rate of absorption. It decrease as the rate of
absorption increase.
AUC
The area under curve –the area under the
plasma level-time curve that gives a measure of
the extent of absorption or the amount of the
drug reaches to the systemic circulation

22. 2-URINARY EXCRETION RATE:
Urinary excretion of unchanged drug is directly proportional to the plasma drug concentration.
Genraly used for the drugs :
Excreted in urine e.g. diuretics and sulphonamides.
Having site of action as urine e.g. nitrofurantoin in UTI
Determination of
Drug drug excreted in
administered to each interval and
the animal modal cumulative
excretion rate
Withdraw urine Analysis of
sample till 7 half sample for
lives unchanged drug

23. Method :
(dXu/dt)– The maximum excretion rate, it is obtained from the peak of plot between
rate of excretion Vs mid point time of urine collection period. It increase with the
extent of absorption.
(tu)max – the time for maximum excretion rate . It decrease as the absorption rate
increase.
Xu~ -- The cumulative amount of drug excreted in the urine it is related to the AUC of
plasma level data and increase as the extent of absorption increases.

25. IN VIVO STUDIES FOR THE SOLID/LIQUID DOSAGE FORMS:
 TABLETS
 CAPSULES
LIQUID (SYRUP /SUSPENSION )
In case of tablets crushed tablets or encapsulate the capsules and the powder is taken .
The diseased condition has to be induced by PTZ. Swiss albino Rat(150 gm) .
Powdered drug dissolved in water and in case of liquid formulation by feeding needle
administered.

26. Experiment
In case of in vivo studies of Anti epileptic drug the
epilepsy is induced by administering PTZ i.v. injection
to the rat and intensity of seizures was observed
(tonic or clonic).
Seizures and rat become un conscious then required
amount of dose ofPhenobarbitone.Sod.200mg
inj/60mg
tab was administered and the effect was observed.
For the study purpose the blood sample was
withdrawn at particular time intervals till the rat again
become conscious but majorly till 3-5 half life of the
drug and the plasma drug concentration was
determined at different time by their analysis for
drug concentration and making a plot of
concentration Vs corresponding time of sample of
collection to obtain the plasma level- time profile.

27. IN VIVO EVALUATION FOR THE SEMI SOLID DOSAGE FORM/TOPICAL
PREPARATION.
 The skin permeation of freshly excised hairless abdominal skin of male Wistar rat (250 gm) in
a non fasting condition was sacrificed .
 Surgically the hairless abdominal skin was removed and cleaned of muscles fat and
vasculature and kept 4’C for 24 hrs.
 Skin was mounted on Franz diffusion cell with dermal skin surface towards the receiver and
corneum remained in contact wit the donor compartment
 The reciver phase was phosphate buffer saline 7.4 pH at 32’+/- 0.1’C and amount of drug
taken for evaluation was 0.4g was taken in donor compartment.
 3-5ml of reciver phase was withdrawn after 1,2,4,6,8,12 hrs and equivalent volume of
phosphate buffer was added to reciever compartment to maintain volume and the withdrawn
 Sample was analyzed under HPLC.

28. APPLICATION OF IVIV EVALUATION
(1)Providing process control and quality assurance.
(2)Determining stable release characteristics of the product over time.
(3)Facilitating certain regulatory determinations (e.g., absence of effect of minor
formulation changes or of change in manufacturing site on performance).
FACTORS AFFECTING IVIV :
Stereochemistry:
 Due to the stereoselectivity, one enantiomer may have more
affinity towards receptor than other.
 Results in difference in Pharmacokinetics and
pharmacodynamics behaviour of two enantiomers of same
drug.
 In such conditions dissolution data of the racemate will not be
 Useful for development of IVIVC.

29. First pass effect:
First pass effect decreases the systemic
availability of parent drug. Therefore
the amount of drug reaching to systemic
circulation will not match with amount
of drug release in GIT. Hence use of
Plasma concentration data of parent
drug will not be appropriate to
calculate in-vivo drug release

30. Food effect:
Presence of food make may alter dissolution behavior of drug and hence it becomes
an important factor that should be considered in IVIVC development. Presence of
food in stomach alters the pH, ionic strength, enzymes level, gastric emptying time
etc.
Burst Release :
In the case of polymer-based delivery systems, the underlying issue with developing
IVIVC is drug release during the initial period called burst release, which results in
biphasic plasma profiles. The bi-phasic profile is believed to occur due to the loosely
associated drug particles with the surface of the (polymer) particles. Because the
burst release is unpredictable and unavoidable , sophisticated modeling techniques
are needed to correlate the in vitro and in vivo data.

31. Other than bioavailability there is one more parameter i.e. Bioequivalency testing.
BIOEQUIVALENCE : It is a relative term which denotes that the two drug
substance in two or more identical dosage forms , reaches to the systemic circulation
at
the relative rate and to the same relative extent i.e. their plasma concentration – time
profiles will be identical without significance difference.
Bioequivalence
.
chemical clinical therapeutic pharmaceutica
equivalence equivalence equivalence l equivalence
Bioequivalence comparise these four main types of equivalence i.e..

32. Chemical equivalence: It indicates that two or more drug products contain the same labeled chemical
substance as an active ingredient in the same amount.
Pharmaceutical equivalence: This term implies that two or more drug products are identical in:
 Strength
 Quality
 Purity
 Content Uniformity
 Disintegration Characterstic
 Dissolution Characterstic
Therapeutic Equivalence: Two brands of a drug product are expected to yield the same clinical
result in the terms of potency and efficacy.
Clinical equivalence : When the same drug from two or more dosage forms gives identical in vivo
effects as measured by pharmacological response or by control of a symptom or disease.

33. THE FUTURE PROSPECTS OF IVIV EVALUATION
Mostly, drug development requires changes in formulations due to a lot of reasons , such as undesired
degradation causing problems in stability, Formulation development , availability of better materials, better
processing results etc. Having an established IVIV evaluation studies can help in avoiding bioequivalence
studies by using the dissolution profile from the changed formulation and subsequently predicting the in vivo
concentration time profile. This predicted in vivo concentration time profile could act as a surrogate of the in
vivo bioequivalence study. This has enormous cost-saving benefit in the form of reduced drug development
spending and speedy implementation of post-approval changes. The nature of post-approval changes could
range from minor (such as a change in non release-controlling excipient) to major (such as site change,
equipment change, or change in method of manufacture, etc).

34. CONCLUSION
The in vitro and in vivo evaluation studies are basically done of considering there wide
application in drug delivery system at various stages of development to setting dissolution
specifications for the new formulations and drug molecule. The most critical application of
in vitro and in vivo evaluation studies with respect to cost saving due to the avoidance of
expensive clinical trials . The in vitro and in vivo evaluation studies includes in vivo
relevance to in vitro dissolution specifications that can serve as surrogate for in vivo
bioavailability. It can also assist in quality control for certain scale-up and post-approval
changes. The FDA Guidance on IVIV evaluation provides general methods and guidelines
for the establishment of IVIVC. The number of studies reported in the area of establishing
IVIVC for several dosage forms are very scarce and further research is necessary in the
development of more meaningful dissolution and permeation methods.