Protein Binding

Protein Binding

Learning Objectives

At the end of this lecture, the student will be able to:

• Identify the drug binding components

• List the vascular and extravascular components to which the drug can bind

• Differentiate between plasma protein binding and tissue binding

• Discuss the factors affecting protein binding

• Analyse the influence of protein binding on the pharmacokinetics of the drug

• Significance of drug displacement interactions

• Discuss the kinetics of protein binding

• Analyse the significance of drug displacement interactions

• Discuss the kinetics of protein binding

Protein Binding of Drugs

• A drug in the body can interact with several tissue components of which the two major categories are blood and extravascular tissue

• The interacting molecules are generally the macromolecules such as proteins, DNA

Importance of protein binding:

• The bound drug is both pharmacokinetically as well as pharmacodynamically inert because a protein bound drug is neither metabolized nor pharmacologically active.

• A bound drug is also restricted since it remains confined to a particular tissue and because of its enormous size cannot undergo transport and thus its half-life is increased

Types of Binding

• Reversible

• Irreversible

Binding of drug to globulin

α1 globulin Bind to a number of steroidal drug cortisone , prednisolone $ thyroxin  , cynocobalamine

α2 globulin (ceruloplasmin ) bind to Vit. A D E K

γ- globulin Bind to antigen

β1-globulin (transferrin ) bind to ferrous ion

β2-globulin bind to carotenoid

Binding of drug to blood cells

Haemoglobin Bind to phenytoin, pentobarbital, phenothiazine

Carbonic anhydrase Acetazolamide, chlorthalidone

Cell membrane Imepramine, chlorpramazine bind to RBCs cell membrane

Tissue binding of drug

Majority of drug bind to extravascular tissue

The order of binding - liver > kidney > lung > muscle

Liver – epoxide of number of halogenated hydrocarbon, paracetamol

Lung – basic drug imipramine, chlorpromazine, antihistamines,

Kidney – metallothionin bind to heavy metal, lead, Hg, Cd,

Skin – chloroquine & phenothiazine

Eye - chloroquine & phenothiazine

Hairs- arsenicals, chloroquine bind to hair shaft.

Bone – tetracycline

Fats – thiopental, pesticide- DDT

Comparison between Plasma Protein Drug Binding and Tissue Drug Binding

Plasma protein- drug binding

Tissue – drug binding

Binding involves weak bonds and thus reversible

Binding generally involves strong and covalent bonds and thus irreversible

Drugs that bind to plasma proteins have small apparent volume of distribution

Drugs that bind to extravascular tissues have large apparent volume of distribution

Half-life of plasma protein bound drug is relatively short

Half-life of extravascular tissue bound drug is relatively long

Does not result in toxicity

Tissue toxicity is common

Displacement from binding sites is possible by other drugs

Displacement by other drugs generally does not occur

Competition between drugs for binding to plasma protein can occur

Tissue-drug binding is generally non- competitive

Factors affecting drug protein binding

1.  Factor relating to the drug

a) Physicochemical characteristic of drug

b) Concentration of drug in the body

c) Affinity of drug for a particular component

2. Factor relating to the protein and other binding component

a) Physicochemical characteristic of the protein or binding component

b) Concentration of protein or binding component

c) Num. of binding site on the binding site

3. Drug interaction

4. Patient related factor

Drug related factors

Concentration of Drug in the Body

• The extent of drug - protein binding can change with both change in drug and protein concentration

• The conc. of drug that binding HSA does not have much of an influence as the therapeutic concentration of any drug is insufficient to saturate it

Ex: Therapeutic concentration of lidocaine can saturate AAG with which it binding as the conc. of AAG is much less in comparison to that of HSA in blood

Drug Protein / Tissue Affinity

• Lidocaine have greater affinity for AAG than HSA

• Digoxin have greater affinity for protein of cardiac muscle than skeleton muscles or plasma

Protein or tissue related factor

Physicochemical property of protein / binding component

– Lipoprotein or adipose tissue tend to bind lipophilic drug by dissolving them to lipid core

– The physiological pH determine the presence of anionic or cationic group on the albumin molecule to bind a variety of drug

Concentration of protein / binding component

• Mostly all drug bind to albumin as it presents a higher concentration than other protein

Number of binding sites on the protein

Albumin has a large number of binding site as compare to other protein and is a high capacity binding component

• Several drug capable to binding at more than one binding site

Examples: flucoxacillin, flurbiprofen, ketoprofen, tamoxifen and dicoumarol bind to both primary and secondary site of albumin

• Indomethacin binds to three different sites

• AAG is a protein with limited binding capacity b/c of it low - conc. and molecular size.

• The AAG has only one binding site for lidocaine , in presence of HSA two binding site have been reported due to direct interaction b/w them

Significance of protein binding of drug

• Absorption

• Systemic solubility of drug

• Distribution

• Tissue binding, apparent volume of distribution and drug storage

• Elimination

• Displacement interaction and toxicity

• Diagnosis

• Therapy and drug targeting


• The binding of absorbed drug to plasma proteins decrease free drug conc

• Thus sink condition and conc. gradient are established acting as the driving force for further absorption.

Systemic solubility of drug

• Water insoluble drugs, neutral endogenous macromolecules, like heparin, steroids, and oil soluble vitamin are circulated and distributed to tissue by binding especially to lipoprotein

• LP act as a vehicle for the circulation of such hydrophobic drug compounds


• The plasma protein-drug binding favours uniform distribution of drug throughout the body by its buffer function

• A protein bound drug in particular does not cross the BBB, placental barrier and the glomerulus

From the above equation, it is clear that greater the unbound or free concentration of drug in plasma, larger its Vd


• The drug - protein complex cannot penetrate into the liverthe chief metabolizing organ

• The larger molecular size also prevents it from getting filtered through the glomerulus

• Only the unbound or free drug is capable of being eliminated

• Drugs which are more than 95% bound are eliminated slowly, i.e. they have long elimination half-live


Percentage binding

Elimination half- life







• Exception: Penicillin is extensively bound but has short elimination half live

• Reason: Rapid equilibrium is achieved between the free and bound drug

Displacement interaction and toxicity


Drug A

Drug B

% drug before displacement













% drug after displacement













% increase in free drug




Displacement interaction and toxicity

1. Displacement of bilirubin from albumin by NSAID’S

2. Displacement of digoxin from its tissue (cardiac muscle) binding site by quinidine

Digoxin has a high volume of distribution since it is extensively bound to extravascular tissue

Quinidine can displace digoxin from its binding site, resulting in high unbound drug concentration in tissues.

Volume of distribution

A. Drugs with large volume of distribution like digoxin

• Even a substantial increase in the degree of displacement of drug in plasma may not effect large increase in free drug concentration


i. Only a small fraction of such drug is present in plasma, most of it is localized in extravascular tissues

ii. Following displacement, the free drug redistributes in extravascular tissues

B. Drugs with small volume of distribution

• Example- Warfarin, displacement can result in large increase in free drug concentration in plasma


• The chlorine atom of chlroroquine when replaced with radiolabelled I-131 can be used to visualize melanomas of the eye since chloroquine has a tendency to interact with the melanin of eyes

• The thyroid gland has a great affinity for iodine containing compounds; hence any disorder of the same can be detected by tagging such a compound with a radioisotope of iodine

Therapy and Drug Targeting

• The binding of drugs to lipoproteins can be used for site-specific delivery of drugs

• This is particularly useful in cancer therapies since certain tumor cells have greater affinity for LDL than normal tissues.

• Thus binding of a suitable antineoplastic to it can be used as a therapeutic tool

• Oestradiol binds selectively and strongly to prostrate and thus prostate cancer can be treated by attaching nitrogen mustard to oestradiol for targeting of prostate glands

Kinetics of protein drug binding



• Drug binding components – blood components and extravascular componets

• Blood – plasma proteins, cell components

• Tissue binding : liver > kidney > lung > muscle

• Plasma binding- reviersible, competitive, no toxicity

• Tissue binding – irreversible, non-competitive, toxicity

• Factors affecting protein binding Drug related, protein related, interaction, disease related

• Protein binding has effect on absorption, distribution, metabolism and drug excretion

• Displacement of drugs with large Vd  no significant effect of displacement

• Displacement of drugs with small Vd  significant increase of free plasma drug concentration

• Protein binding has applications in diagnosis and drug therapy

• Measurement of protein binding – Direct plot, Double reciprocal plot, Scatchard plot, Hitchcock plot

• Protein binding has effect on absorption, distribution, metabolism and drug excretion

• Displacement of drugs with large Vd à no significant effect of displacement

• Displacement of drugs with small Vd à significant increase of free plasma drug concentration

• Protein binding has applications in diagnosis and drug therapy

• Measurement of protein binding – Direct plot, Double reciprocal plot, Scatchard plot, Hitchcock plot

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