Biopharmaceutics lecture 2

TRANSPORT ACROSS BIOMEMBRANES

MECHANISMS OF DRUG TRANSPORT ACROSS BIOMEMBRANES

• Introduction• The apical cell membrane of the columnar

absorption cell behaves as a ‘lipoidal’ membrane, interspersed by sub-microscopic water-filled channels or pores.

• Water soluble substances of small molecular size (radius 0.4 nm) such as urea are absorbed by simple diffusion through the water-filled channels.


• Most drugs molecules are too large to pass through the aqueous channels. The apical cell membrane of the g.i.-blood barrier allows the passage of lipid-soluble drugs in preference to lipid-insoluble drugs.

• 2 Main Mechanisms– Paracellular pathway: between cells– Transcelular pathway


• Paracellular Pathway may be divided into:– Convective – “Solvent drag”– Diffusive component


• Transcellular pathway includes:– Simple passive diffusion– Carrier-mediated transport

• Active transport• Facilated transport

– Endocytosis

Refer to table 16.2 page 231 of Pharmaceutics: The Science of Dosage Form Design (2nd ed) by M. Aulton

From: Hunter J, Hirst BH. Intestinal secretion of drugs. The role of P-glycoprotein and related drug efflux systems in limiting oral drug absorption. Advanced Drug Delivery Reviews 25:129-157, 1997.


– Paracellular Pathway• Transport of materials in the aqueous pores

between cells rather than across them.• The epithelia of the small intestine has more

pores than others.• This pathway is important for the absorption of

ions such as calcium, sugars, amino acids and peptides at concentrations above the capacity of their carriers.

• Small hydrophillic and charged drugs also use this route.

• Transport molecules up to 200 Da mol. Wt.


Convective absorption• Compound is carried across the epithelium by the water

flux.• By this mechanism, very small molecules such as

water, urea and low molecular weight sugars and organic electrolytes are able to cross cell membranes through aqueous filled channels or pores.

• The effective radii of these channels are small (≈ 0.4 nm) such that the mechanism is of little significance in the absorption of large, water-insoluble drug molecules or ions.

• It is the mechanism involved in the renal excretion of drugs and the uptake of drugs into the liver.

Trancellular PathwayPassive Diffusion

• Involves the movement of drug molecules from region of relatively high to low concentration without expenditure of energy.

• Movement continues until equilibrium has been reached between both sides of the membrane and the equilibrium tend to be achieved faster with highly permeable (i.e. lipid soluble drugs) and when membrane has a large surface area (e.g. intestine vs stomach or duodenum).

• The apical cell membrane plays only a passive role in the passive diffusion transport process.

Trancellular PathwayPassive Diffusion

• The main factors determining the rate of drug transport are:

• Physicochemical properties of the drug i.e. particle size, solubility, partition coefficient, pH and pKa.

• The nature of the membrane, and

• The concentration gradient of drugs across the membrane.

Diagrammatic representation of g.i. absorption by passive diffusion

h

Partition PartitionDiffusion

Drug in solution

Drug in solution carried away by circulating blood

G.I FLUIDBLOOD

G.I. MEMBRANE

Fick’s Law of diffusion

• Where dQ/dt = rate of appearance of drug in the blodd at the site of absorption

• D = the effective diffusion coefficient of the drug in the gi membrane

• A = the surface area of g.i. membrane available for absorption by passive diffusion

• k1 = the apparent PC of drug between g.i. ‘membrane’ & the g.i. fluid.

h

CkCkDA

dt

dQ bg 21(

fluid g.i.in drug ofion concentrat

interface membranefluid/ g.i.at membrane theinside drug ofion concentrat the1 k

Fick’s Law of diffusion (Cont.)

• Cg is the concentration of drug in solution in the g.i. fluid at the site of absorption

• k2 is the apparent PC of drug between the g.i. membrane & the blood

• Cb is the concentration of drug in the blood at the site of absorption

• h is the thickness of the g.i. membrane.

Fick’s Law of diffusion (Cont.)

• The drug in blood vessel is rapidly cleared away and the blood thus serves as a “sink” for absorbed drug as a result of:

• Distribution in a large volume of blood i.e. systemic circulation

• Distribution into body tissues and other fluids of distribution

• Metabolism and excretion• Protein binding• Hence, a large concentration gradient is always

maintained across the g.i. membrane during absorption process and this conc. gradient becomes the sole driving force behind drug absorption by passive diffusion mechanism.

Specialized Transport Mechanisms

• Active transport• Facilitated transport

• Active transport• Substances are transported against their concentration

gradient (i.e. from low to high regions of concentration) across a cell membrane.

• It is an energy-consuming process and involves active participation of the apical cell membrane of the columnar absorption cell.

Specialized Transport Mechanisms Active transport

• Drug molecule or ion forms a complex with a “carrier” which, may be an enzyme or some other components of the cell membrane, to form a “drug-carrier” complex.

• This complex then moves across the membrane, liberates the drug on the other side and the carrier returns to the original state and surface to repeat the process.

• As for g.i absorption, transfer occurs only in the direction of g.i. lumen to the blood i.e. not normally against the conc. gradient, the carrier being generally a ‘one-way’ transport system.

Specialized Transport Mechanisms Active transport

• Several carrier-mediated transport systems exist in the small intestine and each is highly selective with respect to the structure of substances it transports.

• Drugs resembling such substances can be transported by the same carrier mechanism. E.g. Levodopa resembles tyrosine and phenylalanine and is absorbed by the same mechanism.

• Active transport proceeds at a rate directly proportional to the concentration of the absorbable species only at low concentration and the mechanism becomes saturated at high concentrations.

xenobiotic

transport

protein

out

in

out

in

Proposed Model for Carrier-Mediated TransportProposed Model for Carrier-Mediated Transport

Membrane Transporters and Their Substrates

TransporterTransporter SubstratesSubstratesAmino acid transportersAmino acid transporters baclofen, cyclosporin, L-dopa, baclofen, cyclosporin, L-dopa,

gabapentin, methyldopagabapentin, methyldopa

Peptide transportersPeptide transporters -lactam antibiotics, ACE -lactam antibiotics, ACE inhibitors, inhibitors, (hPEPT1, HPT1)(hPEPT1, HPT1) cephalexin, cyclosporin, cephalexin, cyclosporin, methyldopamethyldopa

Nucleoside transportersNucleoside transporters zidovudine, zalcitabine, zidovudine, zalcitabine, dipyridamoledipyridamole (CNT1, CNT2)(CNT1, CNT2)

Organic anion transportersOrganic anion transporters ceftriaxone, benzoic acid, ceftriaxone, benzoic acid, methotrexatemethotrexate (OATP1, OATP3, OATP8)(OATP1, OATP3, OATP8) pravastatin pravastatin

Organic cation transportersOrganic cation transporters thiamine, desipramine, thiamine, desipramine, quinidine, quinidine, (OCT1,OCT2)(OCT1,OCT2) midazolam, verapamil midazolam, verapamil

Bile acid transportersBile acid transporters chlorambucil, thyroxinechlorambucil, thyroxine (IBAT/ISBT)(IBAT/ISBT)

Illustration of Specialized Transport

Specialized Transport Mechanisms Facilitated Transport

• Differs from active transport in that it can not transport a substance against its concentration gradient

• Does not require energy input.

• Its driving force is the concentration gradient.

• Another transport facilitator is required in addition to the carrier molecule.

Facilitated Transport of Vit. B12

Carrier

B12

IF

B12-IFTransported Vit. B12

Transcellular PathwayEndocytosis

Definition

The process by the plasma membrane of the cell invaginates and the invagination becomes pinched off.

Small intracellular membrane-bound vesicles are formed and enclose a volume of material.

Material can then be transported into the cell

Transcellular Pathway Endocytosis

• After invagination the material is transferred to othe vesicles or lysosomes and digested.

• This is an energy dependent process.• May be divided into 4 main processes

– Pinocytosis or fluid-phase endocytosis– Receptor-mediated endocytosis– Phagocytosis– transcytosis

Transcellular Pathway Pinocytosis

• Substance does not have to be in aqueous solution to be absorbed.

• Like phagocytosis, it involves invagination of the material by the apical cell membrane of the columnar absorption cell lining the g.i.t. to form vacuoles containing the material.

• These vacuoles then cross the columnar absorption cells.

• It is the main mechanism for the absorption of macromolecules such as proteins and water-insoluble substances like vit. A, D, E and K.

Receptor-mediated endocytosis

• Process of ligand movement from the extracellular space to the inside of the cell by the interaction of the ligand with a specific cell-surface receptor.

• The receptor binds the ligand at its surface• Internalizes it by means of coated pits and

vesicles • The coated cell lose their coat once transported

to the cytoplasm of the cell.

Receptor-mediated Endocytosis

• The ligand is delivered to lysosomes (spherical or oval cell organelles surrounded by a single membrane)

• The lysosomes contain digestive enzymes which break down bacteria and large molecules such as protein, polysaccharides and nucleic acids.

Receptor-mediated endocytosisCell membrane

Free drug

Released drug

Phagocytosis

Phagocytosis is:

• The engulfment by the cell membrane of particles larger than 500 nm

• This is the process of absorption of polio and other vaccines from the GIT.

Transcytosis

• The process by which the material internalized by the membrane domain is transported through the cell and secreted on the opposite side.

Ion-pair transport

• In this mechanism, some ionized drug species interact with endogeneous organic ions of opposite charge to form absorbable neutral specie i.e. an ion-pair.

• The charges are “buried” in ion pair and the complex can now partition into the lipoidal cell membrane lining the g.i.t. and be absorbed by passive diffusion.

• A suitable mechanism for the absorption of quaternary ammonium compounds and tetracyclines which are ionized over the entire g.i. pH range.

• Ion pair ≡ Organic anions + Organic cations = Neutral molecules (crossing lipoidal membrane by passive diffusion.

Efflux of Drugs from the Intestine

• Countertransport efflux proteins expel specific drugs back to the lumen of the GIT after absorption.

• The main countertransport protein is P-glycoprotein.

• High levels of glycoprotein are expressed in the jejunum.

• The efflux reduces bioavailability• Drugs with wide structural diversity are affected

eg. cycolsporine.

Consequence of the Efflux Transporter P-glycoproteinConsequence of the Efflux Transporter P-glycoprotein

1) Limited drug absorption

enterocytepgp

Gut lumen

2) Enhanced drug elimination2) Enhanced drug eliminationProximal tubule cells

Tubule lumen

hepatocytes

bile3) Limited distribution

Endothelial cells

capillary

Brain or testessyncytiotrophoblast

Maternal blood

lymphocyte

Adapted from: Fromm MF. Trends in Pharmacol Sci 25:423, 2004

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Biopharmaceutics lecture 2