7.2 Distribution, Plasma Protein Binding & Transporters (P-gp, OATs, OCTs, BSEP)

Why distribution is scored with transporters

Handbook Domain II.2 does not stop at “the chemical left the gut.” The internal dose that hits a target cell depends on how much is unbound, how fast blood delivers it, whether membranes let it through, and whether pumps concentrate or exclude it. Independent OpenExamPrep teaching in this section covers volume of distribution (Vd), perfusion- versus permeability-limited uptake, plasma protein binding, enterohepatic recirculation, the canalicular and renal transporters that generate exam-style drug-drug and drug-chemical interactions, and storage in adipose tissue and bone. It is not an ABT product and does not claim official approval, review, or partnership with ABT.

Volume of distribution

Vd = amount in the body / plasma concentration (choose the same concentration units and usually a specified time, such as Vss at steady state). Vd is a proportionality constant, not a beaker you could drain from the animal. A Vd near plasma volume (~0.04–0.07 L/kg in humans) means the chemical is largely confined to plasma, often because it is highly protein-bound and poorly permeable. A Vd of several liters per kilogram means extensive tissue partitioning or tissue binding—lipophilicity, intracellular pH trapping, or binding to proteins, DNA, or bone mineral. A large Vd lengthens half-life for a given clearance because more of the body burden sits outside plasma, the compartment from which liver and kidney remove chemical. Do not equate a large Vd with “safety” or with “toxicity”; it only describes where the mass went relative to the plasma number you measured.

Perfusion-limited versus permeability-limited uptake

Well-perfused organs (lung, liver, kidney, brain as a fraction of cardiac output, heart) can equilibrate quickly if the molecule can cross the capillary and cell membranes. For those chemicals, delivery of blood flow is rate-limiting: perfusion-limited (flow-limited) distribution. Poorly perfused tissues such as resting muscle and adipose still take up lipophilic chemicals, but they do so slowly and can become a reservoir.

If the molecule is polar, large, or charged, or if tight junctions and efflux pumps intervene (BBB, testes, placenta), membrane crossing is slower than blood delivery: permeability-limited distribution. Increasing cardiac output will not fill that tissue faster. BBB P-gp can make a lipophilic substrate behave as if the brain were permeability-limited even though cerebral blood flow is high. Fat can be perfusion-limited for highly lipophilic organochlorines once you account for the low blood flow per gram; the tissue still accumulates because the fat:blood partition is huge.

Plasma protein binding: free versus total

Albumin binds many organic acids and some neutrals. Alpha-1-acid glycoprotein (AAG) binds many organic bases and rises in inflammation, so the bound fraction of a base can change without any new chemical being added. Only the unbound (free) concentration is generally available to leave capillaries, bind receptors, be metabolized, or be filtered at the glomerulus (restrictive clearance). Total concentration = unbound / unbound fraction (fu). Laboratories often report total plasma levels because they are easier to measure. In a well-stirred hepatic model, intrinsic clearance of free drug and fu together determine hepatic extraction.

Displacement from albumin by a second ligand transiently raises fu. If elimination of free chemical is intact, clearance of the unbound species increases and unbound concentration often returns toward the previous baseline, while total concentration falls. That is why displacement rarely produces new clinical toxicity by itself. It does wreck naive interpretation: a drop in total concentration can look like “less exposure” when free exposure is unchanged. Displacement plus inhibited clearance, or a narrow-index chemical whose effect tracks a brief free spike, is the exception worth worrying about—not the default. In toxicokinetic studies, always ask whether the assay is free or total and whether AAG or albumin changed with inflammation or malnutrition.

Enterohepatic recirculation

Conjugates (often glucuronides) secreted into bile can be hydrolyzed by intestinal β-glucuronidases, including bacterial enzymes, releasing parent or a reabsorbable aglycone. Reuptake from the ileum returns chemical to the portal vein and liver. Plasma curves may show a secondary peak after a meal or after gallbladder emptying. Half-life lengthens; interrupting the cycle (cholestyramine, activated charcoal, some antibiotics that wipe out deconjugating flora) can drop body burden. MRP2-mediated biliary excretion of organic anions and glutathione conjugates feeds this loop. Species without a gallbladder (rat) still recirculate; they just lack a discrete bile bolus.

Canalicular ABC transporters: BSEP and MRP2

BSEP (bile salt export pump, ABCB11) is the ATP-dependent transporter on the canalicular (apical) hepatocyte membrane that moves bile salts into bile against a steep gradient. If BSEP is inhibited or genetically deficient (progressive familial intrahepatic cholestasis type 2), bile salts accumulate inside hepatocytes, producing cholestasis, pruritus, and hepatocellular injury that can look “bland” on aminotransferases at first. Cyclosporine is a well-documented BSEP inhibitor in membrane-vesicle and clinical experience; bosentan and troglitazone are additional experimental or historical examples. Teach the location (canaliculus), the energy source (ATP), and the substrate (bile salts)—not a memorized brand-name stem.

MRP2 (ABCC2) on the same canalicular membrane exports organic anions, including bilirubin glucuronides and many glutathione conjugates. Inherited MRP2 deficiency is Dubin-Johnson syndrome (conjugated hyperbilirubinemia with a dark liver). Inhibiting MRP2 can raise conjugated bilirubin without being a BSEP-bile-salt story. Do not swap BSEP and MRP2: bile salts versus organic-anion conjugates are different cargoes on different pumps, even though both use ATP at the canaliculus.

Basolateral renal transporters and P-gp DDIs

OAT1 and OAT3 (SLC22A6/A8) on the basolateral membrane of proximal tubule cells take up organic anions from blood (many beta-lactams, antivirals, methotrexate, some uremic toxins). Probenecid competes at OATs and can reduce tubular secretion—historically used to prolong penicillin, clinically relevant for methotrexate toxicity. OCT2 (SLC22A2) on that same basolateral membrane takes up organic cations, including metformin and cisplatin. OCT2-mediated uptake is one reason cisplatin concentrates in proximal tubule cells and injures them; OCT2 inhibitors can reduce that uptake in experimental systems.

P-gp (MDR1, ABCB1) is an ATP-dependent efflux pump on intestinal epithelium (apical, back into lumen), hepatocytes (canaliculus, into bile), renal tubule (apical, into urine), and brain endothelium (back into blood). Substrates include digoxin, many chemotherapeutics, and loperamide (normally kept out of CNS). Inhibitors (quinidine, verapamil, itraconazole, some macrolides, ritonavir) can raise systemic AUC and increase CNS exposure of P-gp substrates. Inducers (rifampin, St. John’s wort via PXR) can lower exposure. Gut P-gp and CYP3A overlap spatially, so many “CYP3A inhibitors” also hit P-gp; sort the chemistry before you blame only oxidation.

Storage in fat and bone

Organochlorines (DDT/DDE, many PCBs, dioxin-like compounds) partition into adipose triglyceride. Blood concentrations can rise during rapid weight loss as fat mass shrinks. Milk fat is an excretion path in lactation. Lead substitutes for calcium in bone hydroxyapatite; bone half-life is measured in years to decades. Chelation and pregnancy-associated bone turnover can remobilize lead into blood. Fluoride also seeks bone mineral. Cadmium bound to metallothionein accumulates in kidney cortex—a protein-binding storage story as much as a fat story. Reservoirs explain why plasma TK after a short exposure can look “cleared” while body burden is not.

Realistic scenario

A four-week oral dog study of a highly albumin-bound organic anion shows a mid-study drop in total plasma concentration after an anti-inflammatory drug is added, with no change in liver enzymes and no change in unbound concentration when you finally measure it. Calling a new clearance autoinduction from the total-concentration plot is premature: fu rose, total fell, free exposure was stable. In a second case, a transplant patient on cyclosporine develops rising bile acids and cholestatic enzymes; canalicular vesicle data show blocked ATP-dependent taurocholate transport. That is BSEP inhibition, not OCT2 cisplatin uptake and not LAT1. A third case: cisplatin plus an OCT2 substrate competition story in the kidney, while a P-gp inhibitor is added to a digoxin regimen—two different membranes, two different pumps.

Traps

• Treating Vd as an anatomic compartment you could dissect out.
• Equating a fall in total plasma concentration with a fall in free exposure after a displacer is added.
• Calling every cholestatic drug a CYP2E1 hepatotoxin without asking about BSEP.
• Placing OAT1 on the canaliculus or BSEP on the BBB.
• Ignoring bone and fat reservoirs because the terminal plasma half-life looked short.