Bioavailability

Session objectives

By the end of this session, students will be able to:

• Describe the concept and characteristics of bioavailability

• Appreciate the importance of bioavailability studies

• Describe the different methods for the assessment of bioavailability

• Discuss on the various considerations in bioavailability

• Justify the role of bioavailability studies in drug approval process

• Identification of right plan for conducting bioequivalence studies

• Analyze the data produced from the bioequivalence studies

Bioavailability

Definition

Rate and Extent of absorption of unchanged drug from its dosage form is known as bioavailability

Bioavailable Dose

The dose available to patient.

Systemic Availability

Amount of drug that reaches systemic circulation (i.e. extent of bioavailability).

Bioavailable fraction (F)

The fraction of administered dose that enters systemic circulation.

F = Bioavailable Dose / Administered Dose

Why Bioavailability Studies??

Bioavailability studies performed for both approved active drug ingredients and not yet approved for marketing by FDA

As standards like identity, strength, quality and purity should be met

Un-marketed drugs which do not have full NDA approval by FDA, in vitro / in vivo bioequivalence studies must be performed

In vivo bioavailability studies are also performed for new formulations of active ingredients that have full NDA approval

These studies are useful in determining safety and efficacy of drug products.

Objectives of Bioavailability studies

Development of suitable dosage form for new drug entity

Development of new formulations of existing drugs

Determination of influence of excipients, patient related factors and possible interactions with other drugs on efficiency of absorption.

Control of quality of drug product during early stages of marketing.

Considerations in bioavailability studies

Bioavailability absolute v/s relative

Single dose vs. multiple dose

Human volunteer – healthy subject vs. patients

Types of Bioavailability

Single dose vs. multiple dose

Multiple dose study has several advantages like:

1. More accurately reflects the manner in which the drug should be used

2. Drugs levels are higher due to cumulative effect which makes its determination possible even by less sensitive analytical methods

3. Better evaluation of the performance of controlled release formulation is possible

4. Small inter-subject variability is observed which allows use of fewer subjects

5. Nonlinearity in pharmacokinetics, if present, can be easily detected.

Single dose

• Very common & easy

• Less exposure to drug & less tedious

• Difficult to predict steady state

Multiple dose

• Difficult to control

• More exposure to drug & tedious

• Time consuming

Human volunteer – healthy subject vs. patients

Female volunteers are used only ideally, the bioavailability study should be carried out in patients for whom the drug is intended to be used.

Advantages

Patient is benefited from the study

Reflects better therapeutic efficacy of drug

Drug absorption pattern in disease state can be evaluated

Avoids the ethical quandary of administering drug to healthy subjects when oral contraceptives are to be tested

No. of subject- extent of inter- subject variability – minimum required to obtain reliable data

They must be informed about the importance of study, conditions to be followed and possible hazards if any. Medical examination should be performed

Drug washout period for minimum of ten biological half-lives must be allowed for between two studies in same subject.

Measurement of Bioavailability

Pharmacokinetic methods (Indirect)

Plasma level time studies

Urinary excretion studies

Pharmacodynamics methods (Direct)

Acute pharmacological response

Therapeutic response

Plasma level time studies

• Plasma level time studies or the plasma concentration – time curve or blood level curve.

• A direct relationship exists concentration of drug at the site of action & concentration of drug in the plasma.

• Serial blood samples are taken after drug administration & analyzed for drug concentration.

• A typical blood level curve obtained after oral administration of drug.

• Three parameters of plasma level-time studies considered important for determining bioavailability are:

Cmax - gives an indication whether drug is sufficiently absorbed systematically to provide therapeutic response.

tmax - gives indication of rate of absorption.

AUC - gives indication of extent of absorption.

• Extent of Bioavailability determined by

F = [AUC]_oral D_iv / [AUC]_iv D_oral

Fr = [AUC]_test   D_std / [AUC]_std    D_test

F= Absolute bioavailability

Fr= Relative bioavailability

With multiple dose study, extent of bioavailability is given by:-

Urinary excretion studies

• Based on principle that the urinary excretion of unchanged drug is directly proportional to plasma concentration of drug

• Used for drugs extensively excreted unchanged in the urine e.g. thiazide diuretics and drugs that have urine as site of action (nitrofurantoin)

• Three major parameters examined in urinary excretion data obtained with single dose study are:

1) (dX_u/dt)_max - Maximum urinary excretion rate. It is analogous to C_max derived from plasma level time studies since rate of appearance of drug in urine is proportional to its concentration in systemic circulation.

2) (t_u)_max - Time for maximum excretion rate. It is analogous to t_max of plasma level time data.

3) X_u - Cumulative amount of drug excreted in urine. It is related to AUC of plasma level time data.

• Extent of bioavailabilityis calculated from equation below:-

F = (X_u∞)_oral D_iv / (X_u∞)_iv D_oral

Fr = (X_u∞)_test  D_std / (X_u∞)_std   D_test

 

F = (X_u, _ss)_test  D_std τ_test / (X_u,_ss )_std D_test  τ _std

Thus, the bioavailability can be calculated in the steady state

Advantages of Urinary Excretion Method

Useful when there is lack of sufficiently sensitive analytical techniques to measure concentration of drug in plasma

Noninvasive method therefore better subject compliance

Convenience of collecting urine samples in comparison to drawing of blood periodically

If any case the urine drug concentration is low, assaying of larger sample volume is relatively more

Direct measurement of bioavailability, both absolute & relative is possible without the necessity of fitting the data to the mathematical model

Acute Pharmacologic Response

• Used when bioavailability measurements by pharmacokinetic method is difficult, inaccurate or non-reproducible

• Method requires measurement of Reponses for at least 3 biological half-lives of drug

Disadvantages

Pharmacologic response tend to be more variable and accurate correlation between measured response & drug available from formulation is difficult

The observed response may be due to an active metabolite whose concentration is not proportional

Therapeutic Response

Based on observing clinical response to a drug formulation given to patients suffering from disease for which it is intended to be used

A major drawback of this method – Quantization of observed response is too improper to allow for reasonable assessment of relative bioavailability between two dosage forms of same drug

Concept of Equivalence

Equivalences

It is a term that compares drug products w.r.t. a specific characteristic or function or to a defined set of standards

Several types of equivalences

1. Chemical equivalence: Two or more drug products contain the same labeled chemical substance

2. Pharmaceutic equivalence: Two or more drug products are identical in strength, quality, purity, content uniformity and disintegration & dissolution characteristics

3. Bioequivalence:- It denotes that drug substance in two or more identical dosage forms, reaches the systemic circulation at the same relative rate and relative extent

4. Therapeutic equivalence:- It indicates that two or more drug products that contain same therapeutically active ingredient, elicit identical pharmacologic effect

Bioequivalence and its Importance

• A drug product that differs from reference material in its rate of absorption, but not in its extent of absorption may be considered bioavailable.

• Bioequivalence requirement may be imposed by FDA on the basis of following:

1. Various drug products do not give comparable therapeutic effect as seen from well controlled clinical trials.

2. Availability of multiple brand of same compound each with claims for perfect bioavailability.

3. It is important for drug with narrow therapeutic index.

• For bioequivalence studies, both test and reference drug formulations contain the pharmaceutical equivalent drug in same dose strength, in similar dosage forms & both are given by same route of administration

• Study is performed in normal, healthy male volunteers who have given informed consent to be in the study

• Study might exclude any volunteers who are smokers, have known allergies to drug, are overweight or have taken any medications within a specified period (1 week) prior to the study

Study Design

1. Completely randomized design

2. Randomized block design

3. Repeated measures, cross-over & carry over design

4. Latin squire design

Completely randomized design

• All treatments are randomly allocated among all experimental subjects

• Method – label all subjects with the same number of digit

Ex: If 20 subjects are available then number them from 1-20.

• Randomly select the non-repeating numbers for the first treatment then repeated for all other treatments

Advantages

1. Easy to construct

2. Can accommodate many number of subjects & treatments

3. Easy & simple to analyze

Disadvantages

1. It is best suited for situation where few treatments are present

2. All subjects must be as homogenous as possible

Randomized block design

• Subjects are first sorted into homogenous group called as blocks then treatments are assigned at random within blocks

Method

• Subjects having similar background characteristics are formed as blocks, then they are randomized within each block

• Randomization for different blocks are done independent of each other

Advantages

 • It can accommodate any number of treatments or replications

• Different treatment need not to have equal sample size

• Easy to construct & analyze

• If an entire block needs to be dropped from the analysis for some reason, analysis is not complicated

• More precise results are obtained

• Variability in experimental units can be introduced to widen validity

Disadvantage

• Missing observation within a block require more complex analysis

• The degree of freedom of experimental error are not as large as with a completely randomized design

Repeated measures, cross-over & carry-over designs

• It is also a randomized block design in which the same subject serves as a block

• The same subject is utilized for each of the treatments under study, since repeated measures on each subject are done so it is called as repeated measure design

• The study involves several treatments or a single treatment evaluated at different point of time

• The administration of two or more treatments one after the other in a specified or random order to the same group of patients are called as crossover or change-over design

• The drawback of crossover studies is the potential for distortion due to carryover that is residual effects from preceding treatments

• To prevent carryover effects a wash out period of 10 elimination half-life should be maintained

• Complete randomization is used to randomize the order of treatments for each subject.

• Randomization for different for subjects are independent of each other

Repeated measures, cross-over & carry-over designs

BE

• Good precision for comparing treatments because experimental errors are excluded

• If the effect of a treatment overtime need to be known then observing the same subject at different intervals of time , then other way

• It is economic on the subjects

• There may be an order effect, which is connected with the position in the treatment order

• There may be a carryover effect, which is connected with the preceding treatments

Latin square designs

• In other methods the subjects remains on treatment from start to the end of experiment so called as continuous trials

• Each subject receive each treatment during the course of the experiment

• It is a 2 factor design (subjects & treatments) with one observation in each cell

• In this design 3 or more treatments are to be compared & carry over effects are balanced

• Rows represent subjects, and columns represent treatments

• r rows & r columns such that each of the r square cell contains one & only one of the r letters representing the treatments

• Each letter appears only once in the row & column Latin square design is considered as standard if the first row & first column consist of the r letter in alphabetical order

• Randomised, balanced, crossover latin square design are commonly used for bioequivalance studies

Advantage

Minimizes the inter subject variability in plasma drug levels

Minimizes the carry over effects which could occur (intra subject variability)

Minimizes the variation due to time effect

Focus on the formulation variables which are key to the bioavailability

Disadvantages

Small degree of freedom for experimental errors when only a few treatments are used

The randomization required is somewhat more complex than the other design

Takes a long time since an appropriate wash out period b/w 2 administration

When the number of formulation to be tested is more, the study becomes more difficult & subject dropout rates are also high

Bio equivalence protocol

1. Title

- Principal investigator

- Project number & date

2. Study objective

3. Study design

a) Design

b) Drug products

- test products

- Reference products

c) Dosage regimen

d) Sample collection schedule

e) Housing

f) fasting/ meals

g) Analytical method

4. Study population

a) Subjects

b) Subject selection

- Medical history

– Physical examination

- Laboratory tests

c) Inclusion /exclusion criteria

-Inclusion criteria

-Exclusion criteria

d) Restriction/ prohibition

5. Clinical procedures

- Dosage &drug administration

- Biological sampling schedule

- Activity of the subjects

6. Ethical considerations

a) Basic principle

b) Institutional review board

c) Informed consent

d) Indication for the subject’s withdrawal

e) Adverse reaction or emergency situation

7. Facilities

8. Data analysis

a) Analytical validation

b) Statically treatment of data

9. Drug accountability

10. Appendix

Statistical Interpretation of Bioequivalence data

• After data has been collected, statistical methods must be applied to determine the level of significance of any observed difference in the rate or extent of absorption in order to establish bioequivalence between two or more drug product.

• Typically, an analysis of variance (ANOVA) method is applied to determine statistical difference (p). if p <  0.05 then the product is not considered statically significant

• If relative bioavailability of test formulation is in range of 80 to 120% of reference standard, it is considered bioequivalent.

• The difference between bioavailability of test formulation should not be greater than +/- 20% of average reference standard.

Summary

• Bioavailability is the fraction of administered dose that enters systemic circulation in an unchanged form

• Bioavailability studies performed for both approved active drug ingredients and not yet approved for marketing by FDA

• Measurement of bioavailability involves both direct and indirect method

• Bioavailability is the determination of rate and extent of absorption of unchanged drug from its dosage form

• Bioavailability studies are performed for both the drug and excipients as a mandatory requirement of FDA to ascertain the safety and efficacy of drug products

• The main objective of bioavailability studies is development of suitable dosage forms for new drug entities as well as existing drugs

• The various considerations in bioavailability studies are:

-   Absolute and relative bioavailability

-   Single dose vs. multiple dose

-   Human volunteer – healthy subject vs. patients

• Bioavailability can be estimated by Pharmacokinetic (Blood level studies and Urinary excretions studies) and     Pharmacodynamics methods (Acute pharmacological response and therapeutic response

• The key pharmacokinetic parameters estimated by plasma level studies (indirect method) are tmax, Cmax and AUC

• Urinary excretion studies are based on the principle that the urinary excretion of unchanged drug is directly proportional to plasma concentration of drug

• Pharmacologic response tend to be more variable and accurate correlation between measured response & drug available from formulation is difficult, hence indirect is preferred

• Bioequivalence is a term that compares drug products w.r.t. a specific characteristic or function or to a defined set of standards.

• Different approaches for bioequivalence studies are randomized block approach, completely randomized approach, repeated measures, cross-over & carry-over design and latin square design

• Randomised and crossover latin square design are commonly used designs for bioequivalence studies

• Analysis of variance (ANOVA) is generally is applied to determine statistical difference (p) for bioequivalence data