Assignment

Course - Biopharmaceutics and Pharmacokinetics 1

Course code - PHR 304

Submitted by: Submitted to:

Md. Azamu Shahiullah Prottoy Zara Sheikh ID – 11146018 Dept. of Pharmacy

Effect of protein-binding on apparent volume of distribution

The extent of drug distribution into tissues depends on the degree of plasma protein and tissue binding.

Extensive plasma protein binding will cause more drugs to stay in the central blood compartment.

Therefore drugs which bind strongly to plasma protein tend to have lower volumes of distribution. Of

these plasma proteins, albumin, which comprises 50 % of the total proteins, binds the widest range of

drugs. Acidic drugs commonly bind to albumin, while basic drugs often bind to alpha1-acid

glycoprotein and lipoproteins. Many endogenous substances, steroids, vitamins, and metal ions are

bound to globulins. The higher the protein binding, lower the tissue distribution.

Proteins with Potential Binding Sites for Various Drugs

In the bloodstream, drugs are transported partly in solution as free (unbound) drug and partly

reversibly bound to blood components (e.g., plasma proteins, blood cells). Of the many plasma

proteins that can interact with drugs, the most important are albumin, α1-acid glycoprotein, and

lipoproteins. Acidic drugs are usually bound more extensively to albumin; basic drugs are usually

bound more extensively to α1-acid glycoprotein, lipoproteins, or both. Only unbound drug is available

for passive diffusion to extravascular or tissue sites where the pharmacologic effects of the drug occur.

Therefore, the unbound drug concentration in systemic circulation typically determines drug

concentration at the active site and thus efficacy. At high drug concentrations, the amount of bound

drug approaches an upper limit determined by the number of available binding sites. Saturation of

binding sites is the basis of displacement interactions among drugs (see Drug–Receptor Interactions).

Drugs bind to many substances other than proteins. Binding usually occurs when a drug associates

with a macromolecule in an aqueous environment but may occur when a drug is partitioned into body

fat. Because fat is poorly perfused, equilibration time is long, especially if the drug is highly lipophilic.

Accumulation of drugs in tissues or body compartments can prolong drug action because the tissues

release the accumulated drug as plasma drug concentration decreases. For example, thiopental is

highly lipid soluble, rapidly enters the brain after a single IV injection, and has a marked and rapid

anesthetic effect; the effect ends within a few minutes as the drug is redistributed to more slowly

perfuse fatty tissues. Thiopental is then slowly released from fat storage, maintaining subanesthetic

plasma levels. These levels may become significant if doses of thiopental are repeated, causing large

amounts to be stored in fat. Thus, storage in fat initially shortens the drug's effect but then prolongs it.

Some drugs accumulate within cells because they bind with proteins, phospholipids, or nucleic acids.

For example, chloroquine concentrations in WBCs and liver cells can be thousands of times higher

than those in plasma. Drug in cells is in equilibrium with drug in plasma and moves into plasma as the

drug is eliminated from the body.

Increases half-life of the drug

The drug/protein complex does not permeate phospholipid bilayers, including capillary membranes,

glomerular membranes in the nephrons, and the blood brain barrier. Bound drugs are also less

available to the enzymes involved in first-pass metabolism. After much of the free drug has been

cleared by the metabolic and excretory processes, the reversible drug/protein complex serves as a

depot to replenish the concentration in vivo. For these reasons, drugs with high protein binding activity

values tend to have a greater half-life compared to those with lower values. The prolonged activity

resulting from these factors may be desirable, or may promote the emergence of undesirable side

effects.

Effect of protein binding on elimination of drugs

Protein binding decreases the renal excretion of drugs and enhances the biological half-life. Only the

free drug can get excreted through filtration. For example, tetracycline is excreted mainly by filtration.

These drugs bind to proteins and results in decreased renal excretion. Interaction of tetracycline (TC)

derivatives viz. oxytetracycline, doxycycline, demeclocycline and chlorotetracycline with bovine

serum albumin (BSA) and concomitant changes in protein conformation were studied using

fluorescence quenching and circular dichroism measurements. Fluorescence data revealed the presence

of one to three binding sites on BSA for different TC derivatives. This binding study with the marker

ligands, warfarin and bilirubin, elucidated site-I as a primary binding site for TCs on albumin.

Scatchard analysis revealed the binding affinity (K(a)) and capacity (n) for these derivatives vary in

the range from 0.8 to 3.2 x 10(6) l/mole and 1.3-3.4, respectively. Significant reduction (60-45%) in

secondary structure (alpha-helical content) of BSA was noticed upon interaction with different TC

derivatives in presence of Cu (II) ions. High affinity binding of TCs with BSA signifies drug stability.

However, excessive binding at higher TC concentrations in combination with Cu (II) induces

conformational change in protein structure, which may exert detrimental effect on cellular protein.

The binding of drugs in extravascular organs decreases their concentration in blood and thereby slows

their elimination by the kidney, liver, or lungs. However, the binding of drugs to plasma proteins may

either hasten or retard the elimination of drugs, depending on the mechanism of elimination. For

example, the binding of drugs to plasma proteins lowers their unbound concentrations in blood and

thereby decreases their rate of elimination by glomerular filtration and by inefficient transport systems

in the kidney. Moreover, binding to plasma proteins would decrease the rate of elimination of lipid-

soluble drugs that diffuse rapidly from the glomerular filtrate back into blood, even though they are

rapidly transported by active transport systems. In addition, the binding of drugs to plasma proteins

would also decrease the rate of drug metabolism by relatively inactive enzyme systems in liver,

especially when the K,, of the enzyme is high. However, when drugs are metabolized rapidly by liver

enzymes and polar drugs are rapidly eliminated by transport systems in the kidney; their clearance by

an organ may approach the blood flow rate through the organ. Under these conditions, the binding to

plasma proteins and to blood cells may enhance the rate of elimination of drugs. Thus, the efficiency

of the mechanisms by which a drug is eliminated determines whether the binding of drugs to plasma

proteins should be considered as storage depots or as transport mechanisms. To illustrate the diverse

effects of binding to plasma proteins on drug elimination, the apparent rate constants of elimination

(rate/concentration) at various concentrations of two hypothetical drugs were calculated. One of the

drugs is completely cleared from plasma as it passes through the liver, and the other is eliminated so

slowly that only 1.0% of the unbound drug is extracted as it passes through the liver. In making of

these calculations, some assumptions were taken such as- 1. The distribution phases have been

completed; 2. The unbound forms of the drugs distribute with body water (50 liters); 3. The kinetic

volumes of distribution are the same as the equilibrium volumes of distribution, that is, the rate

constants for the transfer of drugs from the body compartment to the blood are large compared with

the apparent rate constants of elimination; the blood flow rate through the liver is 1.6 litedmin. When

the total concentrations of the drugs in plasma are infinitely greater than the concentrations of the

bound forms, the apparent volume of distribution of both drugs would be 50 liters, and thus the rate

constant of elimination for the rapidly eliminated drug would be 100 times that for the slowly

eliminated drug. However, as the plasma levels of the drugs decrease, the ratio of the total amounts of

the drugs in the body to their total concentrations in the plasma decreases, whereas the ratio of the total

amounts of the drugs in the body to their unbound concentrations in the plasma increases. Because the

apparent rate constant of elimination equals clearance/v, the apparent rate constant for the rapidly

eliminated drug increases as the plasma level of the drug declines, whereas the rate constant for the

slowly eliminated drug decreases.

Effect of protein binding on patients with kidney

diseases

Albumin is primarily responsible for the binding of acidic drugs whereas basic drugs appear to bind

preferentially to AAG. The low concentration of albumin and AAG in patients with liver disease

indicates the inability of the liver to synthesize these drug-binding proteins. Impaired binding of acidic

and basic drugs is well documented in patients with liver disease. The albumin concentrations in the

transplant patients were low despite stable biochemical liver tests. Because albumin is primarily

responsible for binding of acidic drugs, liver transplant patients would not be able to bind acidic drugs

as efficiently as the normal subjects. However, the AAG concentrations were elevated in all the

transplant patients studied. AAG is an acute-phase reactant and is primarily synthesized in the liver.

AAG concentrations are known to increase in plasma.

Effect of protein binding on patients with hepatic

disease

Protein binding affects distribution. The impaired liver is unable to synthesize plasma proteins

(albumin) adequately. Liver impairment causes accumulation of substances (bilirubin) that displace

drugs from protein-binding sites. A statistics on this case shows that liver disease patients with normal

conc. Of albumin and bilirubin have unbound percentages of frusemide similar to those in healthy

subjects. Defective binding of frusemide is linked to hypoalbuminaemia or hyperbilirubinaemia.

Albumin is an important determinant of the plasma protein binding of frusemide when bilirubin is

normal. Bilirubin binds to albumin strongly and thus reduces the binding reserve of albumin. A

correlation was found between the unbound percentage of frusemide and bilirubin concentration.

Patients with liver disease may have normal degree of frusemide if albumin concentrations are normal.

A bilirubin concentration less than 40mg/l has no effect on the binding of frusemide.

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