Binding to Plasma Proteins

Having entered the blood, drugs may bind to the protein molecules that are present in abundance, resulting in the formation of drug-protein complexes. Protein binding involves primarily albumin and, to a lesser extent, p-globu-lins and acidic glycoproteins. Other plasma proteins (e.g., transcortin, transferrin, thyroxin-binding globulin) serve specialized functions in connection with specific substances. The degree of binding is governed by the concentration of the reactants and the affinity of a drug for a given protein. Albumin concentration in plasma amounts to 4.6g/100 mL or O.6 mM, and thus provides a very high binding capacity (two sites per molecule). As a rule, drugs exhibit much lower affinity (Kd approx. 10-5 -10-3 M) for plasma proteins than for their specific binding sites (receptors). In the range of therapeutically relevant concentrations, protein binding of most drugs increases linearly with concentration (exceptions: salicylate and certain sulfonamides).

The albumin molecule has different binding sites for anionic and cationic li-gands, but van der Waals' forces also contribute (p. 58). The extent of binding correlates with drug hydrophobicity (repulsion of drug by water).

Binding to plasma proteins is instantaneous and reversible, i.e., any change in the concentration of unbound drug is immediately followed by a corresponding change in the concentration of bound drug. Protein binding is of great importance, because it is the concentration of free drug that determines the intensity of the effect. At an identical total plasma concentration (say, 100 ng/mL) the effective concentration will be 90 ng/mL for a drug 10% bound to protein, but 1 ng/mL for a drug 99% bound to protein. The reduction in concentration of free drug resulting from protein binding affects not only the intensity of the effect but also biotransformation (e.g., in the liver) and elimination in the kidney, because only free drug will enter hepatic sites of metabolism or undergo glomerular filtration. When concentrations of free drug fall, drug is resupplied from binding sites on plasma proteins. Binding to plasma protein is equivalent to a depot in prolonging the duration of the effect by retarding elimination, whereas the intensity of the effect is reduced. If two substances have affinity for the same binding site on the albumin molecule, they may compete for that site. One drug may displace another from its binding site and thereby elevate the free (effective) concentration of the displaced drug (a form of drug interaction). Elevation of the free concentration of the displaced drug means increased effectiveness and accelerated elimination.

A decrease in the concentration of albumin (liver disease, nephrotic syndrome, poor general condition) leads to altered pharmacokinetics of drugs that are highly bound to albumin.

Plasma protein-bound drugs that are substrates for transport carriers can be cleared from blood at great velocity, e.g., p-aminohippurate by the renal tubule and sulfobromophthalein by the liver. Clearance rates of these substances can be used to determine renal or hepatic blood flow.

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Drug is -strongly proteins

Drug is -not bound

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Drug Plasma Protein Binding

Renal elimination ^— Renal elimination

Plasma concentration Plasma concentration

Renal elimination ^— Renal elimination

Plasma concentration Plasma concentration

Bound drug

Bound drug



A. Importance of protein binding for intensity and duration of drug effect

The Liver as an Excretory Organ

As the chief organ of drug biotransformation, the liver is richly supplied with blood, of which 1100 mL is received each minute from the intestines through the portal vein and 350 mL through the hepatic artery, comprising nearly 1/3 of cardiac output. The blood content of hepatic vessels and sinusoids amounts to 500 mL. Due to the widening of the portal lumen, intrahepatic blood flow decelerates (A). Moreover, the endothelial lining of hepatic sinusoids (p. 24) contains pores large enough to permit rapid exit of plasma proteins. Thus, blood and hepatic parenchyma are able to maintain intimate contact and intensive exchange of substances, which is further facilitated by microvilli covering the hepatocyte surfaces abutting Disse's spaces.

The hepatocyte secretes biliary fluid into the bile canaliculi (dark green), tubular intercellular clefts that are sealed off from the blood spaces by tight junctions. Secretory activity in the hepatocytes results in movement of fluid towards the canalicular space (A). The hepatocyte has an abundance of enzymes carrying out metabolic functions. These are localized in part in mitochondria, in part on the membranes of the rough (rER) or smooth (sER) endoplas-mic reticulum.

Enzymes of the sER play a most important role in drug biotransformation. At this site, molecular oxygen is used in oxidative reactions. Because these enzymes can catalyze either hydroxylation or oxidative cleavage of -N-C- or -O-C-bonds, they are referred to as "mixed-function" oxidases or hydroxylases. The essential component of this enzyme system is cytochrome P450, which in its oxidized state binds drug substrates (R-H). The Fe'"-P450-RH binary complex is first reduced by NADPH, then forms the ternary complex, 02-Fe"-P450-RH, which accepts a second electron and finally disintegrates into Fe'"-P450, one equivalent of H20, and hydroxylated drug (R-OH).

Compared with hydrophilic drugs not undergoing transport, lipophilic drugs are more rapidly taken up from the blood into hepatocytes and more readily gain access to mixed-function oxidases embedded in sER membranes. For instance, a drug having lipophilicity by virtue of an aromatic substituent (phenyl ring) (B) can be hydroxylated and, thus, become more hydrophilic (Phase I reaction, p. 34). Besides oxidases, sER also contains reductases and glucuronyl transferases. The latter conjugate glucuronic acid with hydroxyl, carboxyl, amine, and amide groups (p. 38); hence, also phenolic products of phase I metabolism (Phase II conjugation). Phase I and Phase II metabolites can be transported back into the blood — probably via a gradient-dependent carrier — or actively secreted into bile.

Prolonged exposure to certain substrates, such as phenobarbital, carbama-zepine, rifampicin results in a proliferation of sER membranes (cf. C and D). This enzyme induction, a load-dependent hypertrophy, affects equally all enzymes localized on sER membranes. Enzyme induction leads to accelerated biotransformation, not only of the inducing agent but also of other drugs (a form of drug interaction). With continued exposure, induction develops in a few days, resulting in an increase in reaction velocity, maximally 2-3 fold, that disappears after removal of the inducing agent.

Drug Excretion Process

Biotransformation of Drugs

Many drugs undergo chemical modification in the body (biotransformation). Most frequently, this process entails a loss of biological activity and an increase in hydrophilicity (water solubility), thereby promoting elimination via the renal route (p. 40). Since rapid drug elimination improves accuracy in titrating the therapeutic concentration, drugs are often designed with built-in weak links. Ester bonds are such links, being subject to hydrolysis by the ubiquitous esterases. Hydrolytic cleavages, along with oxidations, reductions, alkylations, and dealkylations, constitute Phase I reactions of drug metabolism. These reactions subsume all metabolic processes apt to alter drug molecules chemically and take place chiefly in the liver. In Phase II (synthetic) reactions, conjugation products of either the drug itself or its Phase I metabolites are formed, for instance, with glucuronic or sulfuric acid (p. 38).

The special case of the endogenous transmitter acetylcholine illustrates well the high velocity of ester hydrolysis. Acetylcholine is broken down at its sites of release and action by acetylcholinesterase (pp. 100, 102) so rapidly as to negate its therapeutic use. Hydrolysis of other esters catalyzed by various esterases is slower, though relatively fast in comparison with other biotransformations. The local anesthetic, procaine, is a case in point; it exerts its action at the site of application while being largely devoid of undesirable effects at other locations because it is inactivated by hydrolysis during absorption from its site of application.

Ester hydrolysis does not invariably lead to inactive metabolites, as exemplified by acetylsalicylic acid. The cleavage product, salicylic acid, retains pharmacological activity. In certain cases, drugs are administered in the form of esters in order to facilitate absorption (enalapril ^ enalaprilate; testosterone undecanoate ^ testosterone) or to reduce irritation of the gastrointestinal mucosa (erythromycin succinate ^ erythromycin). In these cases, the ester itself is not active, but the cleavage product is. Thus, an inactive precursor or prodrug is applied, formation of the active molecule occurring only after hydrolysis in the blood.

Some drugs possessing amide bonds, such as prilocaine, and of course, peptides, can be hydrolyzed by pepti-dases and inactivated in this manner. Peptidases are also of pharmacological interest because they are responsible for the formation of highly reactive cleavage products (fibrin, p. 146) and potent mediators (angiotensin II, p. 124; bradykinin, enkephalin, p. 210) from biologically inactive peptides.

Peptidases exhibit some substrate selectivity and can be selectively inhibited, as exemplified by the formation of angiotensin II, whose actions inter alia include vasoconstriction. Angiotensin II is formed from angiotensin I by cleavage of the C-terminal dipeptide histidylleu-cine. Hydrolysis is catalyzed by "angio-tensin-converting enzyme" (ACE). Peptide analogues such as captopril (p. 124) block this enzyme. Angiotensin II is degraded by angiotensinase A, which clips off the N-terminal asparagine residue. The product, angiotensin III, lacks vasoconstrictor activity.

Esterases Ester Peptidases Amides Anilides


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