Biopharmaceutics

 

Syllabus:

2.1 Passage of drugs across biological barriers: Passive diffusion, active transport, facilitated diffusion and pinocytosis.

2.2 Factors influencing absorption: physico-chemical, physiological and pharmaceutical.

2.3 Drug distribution in the body. Plasma protein binding.

 

TRANSPORT OF DRUG ACROSS BIOLOGICAL BARRIERS

For systemic absorption, a drug must pass from the absorption site through one or more layers of cells to gain access into the general circulation. For absorption into the cells, a drug must traverse the cell membrane.

STRUCTURE OF CELL MEMBRANE

Cell membrane surrounds the entire cells and acts as a boundary between cell and interstitial fluid. Cell membrane acts as a selective barrier to the passage of molecules. Water, some small molecules, and lipid-soluble molecules pass through such membrane; whereas highly charged molecules and large molecules, such as proteins and protein-bound drugs, do not.

Structure

Cell membranes are generally thin, approximately 70 to 100  in thickness. They are primarily composed of phospholipids in the form of bilayer. Some carbohydrates and proteins are interdispersed within this lipid bilayer.

Lipid bilayer or Unit membrane theory ( Proposed by Davson & Danielli; 1952)

According to this theory the cell membrane is composed of two layers of phospholipids between two surface layers of proteins. The hydrophilic “head” groups of the phospholipids facing the protein layers and the hydrophobic “tail” groups of the phospholipids aligned towards the interior.

* This theory can explain:

               the observations that lipid-soluble drugs tend to penetrate cell membranes more easily than polar                molecules.

* This theory cannot explain:

               the diffusion of water, small molecules such as urea, and certain charged ions through this lipid-               bilayer.

Fluid mosaic model (Proposed by Singer & Nicolson 1972)

According to this model the cell membrane consists of globular proteins embedded in a dynamic fluid, lipid-bilayer matrix.

Integral proteins are embedded in the lipid bilayer; The integral proteins provide a pathway for selective transfer of certain polar molecules and charged ion through the lipid membrane.

Peripheral proteins are associated with the inner and outer surfaces of the membrane. In the inner surface the peripheral proteins are attached to the fatty acid chain and at outer surface they are attached to the integral proteins or to oligosacchrides.

The carbohydrates consist of monosaccharides attached together in chains that are attached to proteins (forming glycoproteins) or to lipids (forming glycolipids).

Carbohydrates are always on the exterior side and peripheral proteins are always on the cytoplasmic or inner surface.

The principal mechanisms of transport of drug molecules across the cell membrane are :

1.            Passive diffusion

2.            Carrier mediated transport

                              (a) Active transport

                              (b) Facilitated transport

3.            Vesicular transport

                              (a) Pinocytosis

                              (b) Phagocytosis

4.            Pore transport

5.            Ion pair formation

1. PASSIVE TRANSPORT

Passive diffusion is the process by which molecules spontaneously diffuse from a region of higher concentration to a region of lower concentration. This process is passive because no external energy is expended.

Characteristics of passive transport

1.      Drug molecules moves from a region of relatively high concentration to one of lower concentration.

2.      The rate of transfer is proportional to the concentration gradient between the compartments involved in the transfer.

3.      The transfer process achieves equilibrium when the concentration of the transferable species is equal on both sides of the membrane.

4.      Drugs which are capable of existing in both charged and a non-charged form approach an equilibrium state primarily by transfer of the non-charged species across the membrane.

5.      Greater the membrane/water partition coefficient of drug faster the absorption [since the membrane is lipoidal in nature, a lipophilic drug diffuses at a faster rate by solubilising in the lipid layer of the membrane]

Mathematical expression

               Passive diffusion is best expressed by Fick’s first law of diffusion which can be expressed mathematically:

where,   dQ/dt     = rate of drug diffusion (mass/time)

               D            = diffusion coefficient of the drug through the membrane (area/time)

               A            = surface area of the membrane through which drug diffusion is taking place (area)

               K_m/w       = Partition coefficient of the drug between the lipoidal membrane and the GI-fluids                      (no units).

Several factors influence the passive diffusion of the drug:

1.      The degree of lipid solubility of the drug (Km/w)

Highly lipid soluble drug has large value of Km/w and hence has higher rate of transport.

2.      the surface area of the membrane (A)

Duodenal area shows most rapid drug absorption than that of other places of intestine because duodenal  area has villi and microvilli, which provide a large surface area. This villi are less abundant in other area of the GIT.

3.      thickness of the membrane (h)

Drugs usually diffuses very rapidly through the capillary cell membrane except through the cell membranes present in the capillaries of the brain. In the brain, the capillaries are densely lined with glial cells, so a drug diffuses slowly into the brain.

2. CARRIER MEDIATED TRANSPORT

Some polar molecules cross the membrane more readily than can be predicted from their concentration gradient and partition coefficient values. This suggests the presence of some specialized transport mechanisms without which many essential water-soluble nutrients like monosaccharides, amino acids and vitamins will be poorly absorbed. The mechanism is thought to involve a component of the membrane called as the carrier that binds reversibly or noncovalently with the solute molecules to be transported. This carrier-solute complex traverses across the membrane to the other side where it dissociates and discharges the solute molecule. The carrier then returns to its original site to complete the cycle by accepting a fresh molecule of solute. The carrier nay be an enzyme or some other component of the membrane.

Characteristics of Carrier Mediated Transport:

1.      The transport is structure specific i.e. the carrier can bind with a specific chemical structure only. Since the system is structure-specific, drugs having structure similar to essential nutrients, called false-nutrients are absorbed by the same carrier system. e.g. 5-fluorouracil and 5-bromouracil serves as false nutrients.

2.      As the number of carrier systems are limited there will be competition between similar chemical structures for the carrier molecules.

3.       Since there are a finite number of carriers available, the system is capacity limited. If the total number of transferable molecules exceeds the number of carrier sites available for transfer, the system will become saturated. The system will then be working in full capacity and the transfer of drug may thus occur at a constant rate until the concentration of drug falls below that of the capacity limit of the system.

4.      For a drug absorbed by passive diffusion the rate of absorption increases linearly with the concentration but in case of carrier mediated process, the drug absorption increases linearly with concentration until the carriers become saturated after which it becomes curvilinear and approach a constant value at higher doses. Such a capacity limited process can be adequately described by mixed order kinetics also called as Michaelis-Menten saturation or non-linear kinetics.

The process is called mixed order because it is first order at subsaturation drug concentration but apparent zero order at and above saturation levels.

N.B. The bioavailability of a drug absorbed by such a system decrease with increasing dose – for example vitamins like B₁, B₂ and B₁₂. Hence administration of large dose of such vitamins is irrational.

5.      Carrier-mediated absorption generally occurs from specific sites of the intestinal tract which are rich in number of carriers. Such an area in which the carrier system is most dense is called as absorption window. Drugs absorbed through such absorption windows are poor candidates for controlled release formulations.

Active Transport

1.      The drug is transported from a region of lower concentration to a region of higher concentration, i.e. against the concentration gradient.

2.      Since the process is occurring against the concentration gradient hence, energy is required in the work done by the carrier.

3.      As the process requires expenditure of energy it can be inhibited by metabolic poisons that interfere with energy production like fluorides, cyanide and dinitrophenol and lack of oxygen.

4.      It is a capacity limited process. When all the carriers become saturated the drug is carried at a constant rate.

Endogeneous substances that are transported actively include

               Sodium (Na⁺), potassium (K⁺), calcium (Ca⁺⁺), iron (Fe⁺⁺) in ionic state;

               certain amino acids and

               vitamins like niacin, pyridoxine and ascorbic acid.

Drugs having structural similarity to such agents are absorbed actively, particularly the agents useful in cancer chemotherapy.

Examples:            Absorption of 5-fluorouracil and 5-bromouracil via pyrimidine transport system,

                              Absorption of methyldopa and levodopa via L-amino acid transport system

                              Absorption of angiotensin converting enzyme (ACE) inhibitor (e.g. enalapril)via the small                     peptide carrier system

Facilitated diffusion

Facilitated diffusion is also a carrier mediated transport system but it moves along a concentration gradient (i.e from higher to lower concentration) and hence it does not require any energy.

Characteristics:

·        It is a carrier mediated transport system.

·        The carriers are saturable and structurally selective for a drug and shows competition kinetics for drugs having similar structures.

·        It does not require any energy expenditure.

·        Facilitated diffusion of ions takes place through proteins, or assemblies of proteins, embedded in the plasma membrane. These transmembrane proteins form a water-filled channel through which the ion can pass down its concentration gradient. The transmembrane channels that permit facilitated diffusion can be opened or closed. They are said to be "gated". Some types of gated ion channels:

• ligand-gated
• mechanically-gated
• voltage-gated
• light-gated

Example:

·        Acetylcholine (ligand) binds to certain synaptic membrane and opens Na⁺ channels and initiate a nerve impulse.

·        Gamma amino butyric acid (GABA) binds to GABA_A receptors and the chloride channel opens. This inhibits the creation of a nerve impulse.

3. VESICULAR TRANSPORT

Vesicular transport is the process of engulfing particles or dissolved materials by the cell.

There are two types of vesicular transport – Pinocytosis and Phagocytosis.

Pinocytosis refers to the engulfment of small solutes or fluid.

Phagocytosis refers to the engulfment of larger particles or macromolecules, generally by macrophages.

Endocytosis and exocytosis are the processes of moving macromolecules into and out of a cell, respectively.

During pinocytosis or phagocytosis, the cell membrane invaginates to surround the material and then engulfs the material, incorporating into the cell (fig). subsequently the cell membrane containing the material forms a vesicle or vacuole within the cell.

e.g.

·        Vesicular transport is the proposed process for the absorption of orally administered Sabin polio vaccine and large proteins.

·        Transport of proteins, polypeptides like insulin from insulin producing cells of the pancreas into the extracellular space.

4. PORE TRANSPORT

Very small molecules (such as urea, water, and sugars) are able to rapidly cross cell membranes as if the membrane contains channels or pores. [although pores are not evident microscopically]. A certain type of protein called transport protein may form an open channel across the lipid membrane of the cell.

e.g.

·        Drug permeation through aqueous pores is used to explain the renal excretion of drugs and the uptake of drugs into the liver.

5. ION PAIR FORMATION

Strong electrolyte drugs are highly ionized or charged molecules, such as quaternary nitrogen compounds with extreme pKa values. Strong electrolyte drugs maintain their charge at all physiologic pH values and penetrate the membrane very poorly.

When ionized drugs is linked up with an oppositely charged ion, an ion pair is formed in which the overall charge of the pair is neutral. This neutral drug-complex diffuses more easily across the membrane.

e.g.

·        Propranolol, a basic drug, forms an ion pair with oleic acid.

·        Quinine forms an ion pair with hexylsalicylate.

2.2 FACTORS INFLUENCING ABSORPTION

physico-chemical, physiological and pharmaceutical

A. PHYSICOCHEMICAL FACTORS

(i)     Drug solubility and dissolution rate

(ii)   Particle size and effective surface area

(iii) Polymorphism and amorphism

(iv)  Pseudopolymorphism (hydrates / solvates)

(v)   Salt form of the drug

(vi)  Lipophilicity of the drug – (pH partition hypothesis)

(vii)pKa of the drug and pH  – (pH partition hypothesis)

(viii)Drug stability

B. PHYSIOLOGICAL FACTORS

These includes factors relating to the anatomic, physiologic and pathologic characteristics of the patient.

(i)     Age

(ii)   Gastric emptying time

(iii) Intestinal transit time

(iv)  Gastrointestinal pH

(v)   Disease states

(vi)  Blood flow through the GIT

(vii)Gastrointestinal contents: a) Other drugs   b) Food                c) Fluids                              d) Other normal GI contents

(viii) Pre-systemic metabolism by        a) Luminal enzymes         b) Gut wall enzymes

                              c) Bacterial enzymes        d) Hepatic enzymes

C. PHARMACEUTICAL FACTORS

(i)     Disintegration time (tablets / capsules)

(ii)   Dissolution time

(iii) Manufacturing variables

(iv)  Pharmaceutical ingredients (excipients / adjutants)

(v)   Nature and type of dosage form

(vi)  Product age and storage conditions

1. Drug solubility and dissolution rate

               Orally administered solid dosage form are first disintegrated or deaggregated, then the solid particles are dissolved; drugs in solution then permeate across biomembrane to be absorbed in the body.

Two critical processes in the absorption of orally administered drugs are:

               1. Rate of dissolution, and

               2. Rate of drug permeation through the biomembrane (i.e. gastrointestinal membrane)

·        For poorly water-soluble drugs rate of dissolution is the rate determining step hence the absorption is called to be dissolution rate limited. e.g. griseofulvin, spironolactone.

·        For highly water-soluble drugs dissolution is rapid so the rate determining step is permeation hence, the absorption is called to be permeation rate limited. e.g., cromlyn sodium, neomycin sulfate etc.

2. Particle size and effective surface area of the drug particles.

From Noyes-Whitney’s equation of dissolution:

where,   D            = diffusion coefficient or diffusivity of the drug molecule

               A            = surface area of the dissolving solid exposed to the dissolution medium

               K_O/W       = water/oil partition coefficient of the drug

               V            = volume of dissolution medium

               h             = thickness of the stagnant layer

               Cs – C_B = concentration gradient of the diffusing drug molecule.

From this equation it can be concluded that the greater the surface area, A, faster the distribution.

When the particle size of a certain mass of a drug is reduced the surface area is increased, hence, if particle size is reduced dissolution rate increases.

Two types of surface area can be defined:

1.      Absolute surface area: Which is the total area of solid surface of any particle, and

2.      Effective surface area: Which is the area of solid surface exposed to the dissolution medium.

e.g.         Micronization of poorly water soluble drugs like griseofulvin, chloramphenicol and several                salts of tetracycline results in superior dissolution rates.

However, size reduction has some limitation. In case of hydrophobic drugs like aspirin, phenacetin and phenobarbital micronization actually results in a decrease in effective surface area due to the following reasons.

(i)     The hydrophobic surface of the drugs absorb air onto their surface which inhibit their wettability, such powders float on the dissolution medium.

(ii)   The particle reaggregate to form larger particles due to their high surface free energy.

(iii) Extreme particle size reduction may impart surface charges that may prevent wetting; moreover electrically induced agglomeration may prevent intimate contact of the drug with the dissolution medium.

3. Polymorphism and amorphism

               Depending on the internal structure, a solid can exist either in a crystalline or amorphous form.

·        When, a substance exists in more than one crystalline form, the different forms are designated as polymorphs and the phenomenon as polymorphism.

N.B. Various polymorphs can be prepared by crystallizing the drug from different solvents under diverse conditions. Depending on their relative stability, one of the several polymorphic forms will be physically more stable than the others. Such a stable polymorph represents the lowest energy state, has highest melting point and least aqueous solubility. The remaining polymorphs are called metastable forms which represents higher energy state, the metastable forms have a thermodynamic tendency to convert to the stable form. A metastable form cannot be called unstable because if it is kept dry, it will remain stable for years.

·        So the metastable forms have higher aqueous solubility and hence higher bioavailability than the stable polymorphs.

e.g. Chloramphenicol palmitate has three polymorphs A, B and C. The B -form shows best bioavailability and A form is virtually inactive biologically.

e.g. Polymorphic form-III of riboflavin is 20 times more water soluble than the form-I.

·        Due to aging of dosage forms containing metastable forms of the drug results in the formation of less soluble, stable polymorph.

e.g. more soluble crystalline form-III of cortisone acetate converts to less soluble form-V in an aqueous suspension resulting in caking of solid.

Amorphous form (i.e. having no internal structure)

               Such drugs represents the highest energy state and can be considered as supercooled liquids. They have greater aqueous solubility than their crystalline form.

e.g. the amorphous form of the novobiocin is 10 times more soluble than the crystalline form.

Thus the order for dissolution of different solid forms of drug is amorphous > metastable > stable.

4. Pseudopolymorphism (Hydrates / Solvates)

               During crystallization process the solvent molecules may be incorporated into the crystal lattice of the solid in stoichiometric proportion – these type of crystals are called solvates; and the trapped solvent molecules as solvent of crystallization.

               The solvates again can remain in different polymorphic states, called as pseudopolymorphs. The phenomenon is called as pesudopolymorphism.

               When the solvent with the drug is water, the solvate is known as hydrate.

Effect of absorption:

·        Generally, the anhydrous form of a drug has greater solubility than the hydrates. This is because the hydrates are already in equilibrium with water and therefore have less demand for water.

e.g. anhydrous form of theophyline and ampicillin have higher aqueous solubilities, dissolve at faster rate and show better bioavailability in comparison to their monohydrates and trihydrate forms respectively.

·        On the other hand nonaqueous solvates have greater aqueous solubility than the nonsolvates.

e.g. n-pentanol solvate of fludricortisone and succinyl sulfathiazole and the chloroform solvates of griseofulvin are more water soluble than their non-solvate forms.

5. Salt form of the drug

               Most drugs are either weak acids or weak bases. One of the easiest approach to enhance the solubility and dissolution rate of such drugs is to convert them into their salt forms.

·        Weak acid HA is more soluble in basic pH and weak base B is more soluble in acidic pH by the formation of salt.

·        Some time in-situ salt formation can be utilized, e.g. certain drugs like aspirin and penicillin are prepared as buffered alkaline tablets. When the tablets are put into water the pH of the microenvironment of the drug is increased which promotes the dissolution rate. So buffered aspirin tablets have two advantages

(i)     the gastric irritation and ulcerogenic tendency of the drug is greatly reduced and,

(ii)   in dry form the hydrolytic stability is better.

(iii) bioavailability is increased by increasing the dissolution.

·        Size of counter ion

               Smaller the size of the counter ion (of the salt form of a drug) greater the solubility of the salt. e.g. bioavailability of novobiocin from its sodium salt, calcium salt and free acid forms are in the following ratio:

               Novobiocin Na                  Novobiocin Ca                  Novobiocin free acid

                         50                                               20                                           1

·        Ionic strength of the counter ion

               When the counter ion is very large in size and/or has poor ionic strength (as in the case of ester form of the drugs), the solubility may be much lower than the free drug itself.

e.g. pamoates, stearates and palmitates of weak bases having poor aqueous solubility:

               prolong the duration of action                      – e.g. steroidal salts

               overcome bad taste                                         – e.g. chloramphenicol

               overcome GI-instability                                 – e.g. erythromycin estolate

decrease the side effects, local or systemic.

6. pKa of the drug and pH

·        Drug pKa and lipophilicity and GI pH (pH partition theory)

               The pH partition theory (Brodie et.al.) states that for drug compounds of molecular weight greater than 100, which are primarily transported across the biomembrane by passive diffusion. The process of absorption is governed by

1.      dissociation constant (Ka) of the drug

2.      lipid solubility of the unionized drug (K_o/w)

3.      the pH at the absorption site

The above statement of the hypothesis was based on the assumptions that:

1.      The GIT is simple lipoidal barrier to the transport of drug.

2.      Larger the fraction of unionized drug, faster the absorption.

3.      Greater the lipophilicity (K_o/w) of the unionized drug, better the absorption.

Handerson-Hasselbach equation

               The amount of drug that exists in unionized form is a function of dissociation constant (pKa) of the drug and pH of the fluid at the absorption site.

Handerson-Hasselbach equation

              

for weak acid:

Drugs	pKa	pH at the site of absorption
Very weak bases Theophyline Caffeine Oxazepam Diazepam	(pKa < 5.0) 0.7 0.8 1.7 3.7	Unionized at all pH values: absorbed along the entire length of GIT.
Moderately weak bases Reserpine Heroin Codeine Amitriptyline	(5 < pKa < 11) 6.6 7.8 8.2 9.4	Ionized at gastric pH, relatively unionized at intestinal pH better absorbed from intestine.
Stronger base Mecamylamine Guanethidine	(pKa > 11.0) 11.2 11.7	Ionized at all pH values: poorly absorbed form GIT.

               It is the pKa of the drug that determines the degree of ionization at a particular pH and that only the unionized drug, if sufficiently lipid soluble, is absorbed into the systemic circulation.

               Ideally, for optimum absorption, a drug should have sufficient aqueous solubility to dissolve in the fluids at the absorption site and lipid solubility (K_o/w) in the lipoidal biomembrane and into the systemic circulation. In other words, a perfect hydrophilic-lipophilic balance (HLB) should be there in the structure of the drug for optimum bioavailability.

B. PHYSIOLOGICAL FACTORS

I.  Physiology of GIT

·        The major components of the GIT are stomach, small intestine (duodenum, jejunum and ileum) and large intestine (colon) which differ from each other in terms of anatomy, function, secretions and pH.

·        The mean length of the entire GIT is 450 cm.

·        The entire inner surface of GIT from stomach to large intestine is lined by a thin layer of muco-polysaccharides (mucous membrane) which normally acts as a barrier to bacteria, cells or food particles.

1. Mouth	pH 6 – 8	small surface area	lipophilic, neutral and basic drugs are absorbed directly
2. Stomach	pH 1 – 3	not too large a surface area	lipophilic, neutral and acidic drugs absorbed but lesser than that from intestine
3. Small intestine	pH 5 – 7.5	very large surface area	major site for absorption of all types of drugs (lipophilic, neutral, acidic or basic)
4. Large intestine	pH7.9–8.0	small surface area	all types of drugs are absorbed but to a lesser extent
5. Rectum	pH 7.5–8.0	much smaller surface area	all types of drugs are absorbed, about half of the absorbed drug goes directly into the systemic circulation and the other half to the liver

Stomach

The stomach is a bag like structure having a smooth mucosa and thus small surface area. Its acidic pH, due to its secretion of HCl, favors absorption of weakly acidic drugs like aspirin.

Small intestine

Fig. Components of intestinal epithelium

               The folds in intestinal mucosa, called as fold of Kerckring result in 3 fold increase in surface area. The surface of this folds possess finger like projections as villi which increases the surface area by 30 times.

·        From the surface of villi protrude several microvilli resulting in 600 times increase in the surface area. All these combined to impart a large surface area of more than 200 sq.m.

·        The blood flow is 6 – 10 times more than stomach.

·        pH range is 5 to 7.5 which is more favorable for most drugs to remain unionized.

·        The peristaltic  movement of intestine is slow, transit time is long, and penetrability is high. All this factors make intestine the best site for absorption of most drugs.

 Large intestine

               Its length and mucosal surface area is very small (villi and microvilli are absent) compared to small intestine and thus absorption of drug from this region is very small.

               However, because of the long residence time (6 to 12 hrs), colonic transit may be important in the absorption of some poorly soluble drugs and sustained release dosage forms.

II. Patient related factors

1. Age

In infants gastric:              pH is high

                                             intestinal surface is small

                                             blood flow is less.

In elderly persons:            altered gastric emptying

                                             decreased intestinal surface area

                                             decreased GI blood flow

                                             achlorhydria

                                             bacterial overgrowth in small intestine.

In both of these age drug absorption is impaired.

2. Gastric emptying

               Passage of gastric content from stomach to small intestine is called gastric emptying.

·        Rapid gastric emptying is required where:

(i)     a rapid onset of action is required e.g. sedatives.

(ii)   dissolution of drug occurs in the intestine e.g. enteric coated dosage forms.

(iii) the drugs are not stable in gastric fluid e.g. penicillin-G and erythromycin.

(iv)  the drug is best absorbed from the distal part of the small intestine e.g. vitamin B₁₂.

·        Delay in gastric emptying is required where:

(i)     the food promotes drug dissolution and absorption e.g. griseofulvin

(ii)   disintegration and dissolution of dosage form is promoted by gastric fluid

(iii) the drugs are absorbed from the proximal part of the small intestine e.g. vitamin B₂ and vitamin C.

·        Gastric emptying is a first order rate process. Several parameters are used to quantify gastric emptying:

(i)     Gastric emptying rate is the rate at which the stomach content empty into the intestine.

(ii)   Gastric emptying time is the time required for the gastric content to empty completely into the small intestine.

(iii) Gastric emptying t_1/2 is the time taken for half the stomach contents to empty.

N.B.       In vivo gastric emptying can be studied by using radio-opaque contrast materials (e.g. BaSO₄) or                tagging the drug with a radio-isotope and scanning the stomach at regular intervals of time.

·        Factors influencing gastric emptying rate:-

1. Volume of meal:

               Larger the volume of meal longer the gastric emptying time.

2. Composition of meal

               The rate of gastric emptying for various food materials is in the following order:

                              carbohydrates > protein > fats

3. Physical state and viscosity of meal

               Liquid meals take less than an hour to empty solid meals take as long as 6 – 7 hours to empty.

               Viscous material empty at a slow rate in comparison to less viscous materials.

4. Temperature of the meal

               High or low temperature of the ingested fluid (compared to body temperature) reduce gastric                emptying rate.

5. Gastrointestinal pH

               Gastric emptying is retarded at low stomach pH and

                                               is promoted at higher or alkaline pH.

6. Electrolyte and osmotic pressure

Water, isotonic, and solutions of low salt concentration empty the stomach rapidly whereas higher electrolyte concentration decreases gastric emptying rate.

7. Body posture

Gastric emptying is favoured while standing and while lying on the right side; while lying on the left side or in supine position retards it.

8. Emotional state

Stress and anxiety promote gastric motility whereas depression retards it.

9. Exercise

Vigorous physical exercise retards gastric emptying.

10 Disease states

Diseases like gastroenteritis, gastric ulcer, pyloric stenosis, diabetes and hypothyroidism retard gastric emptying.

11. Drugs

Drugs that retard gastric emptying includes

(i) poorly soluble antacids e.g. aluminium hydroxide,

(ii) anticholinergics e.g. atropine, propantheline

(iii) narcotic analgesics e.g. morphine and

(iv) tricyclic antidepressants e.g. imipramine, amytriptyline.

Drug that stimulate gastric emptying are:

(i) metoclopramide

(ii) domperidone

(iii) cisapride

3. Effect of GI pH on drug absorption

GI fluid pH influence drug absorption in several ways:

1. Disintegration

The disintegration of some dosage forms is pH sensitive. With enteric coated formulations, the coat dissolves only in the intestinal pH, followed by disintegration of the tablet.

2. Dissolution

A large number of drugs are either weakly acidic or weakly basic whose solubility is greatly affected  by pH. A pH that favours the formation of salt of the drug enhances the dissolution rate. e.g. Weakly acidic drugs dissolve rapidly in the alkaline pH of the intestine whereas basic drugs dissolves in the acidic pH of the stomach.

3. Absorption

Depending upon the pKa of the drug and the pH of the GI fluid some amount of the drug remain in ionized state and some in unionized state. The unionized form will be absorbed through GIT quickly than the ionized form.

4. Stability

GI pH influences the chemical stability of drugs. e.g. The acidic stomach pH is known to affect degradation of Penicillin-G and erythromycin.

4. Effect of GI content

               A number of GI contents can influence drug absorption.

1. Food-drug interaction

Presence of food may either delay, reduce, increase or may not affect drug absorption.

Delayed	Decreased	Increased	Unaffected
Aspirin Paracetamol Diclofenac	Penicillins Erythromycin Ethanol Tetracyclines Levodopa, Iron	Griseofulvin Diazepam	Methyldopa Sulfasomidine

               As a general rule, drugs are better absorbed under fasting conditions and presence of food retards or prevents it.

               Food does not significantly influence absorption of a drug taken half an hour or more before meals and two hours or more after meals.

·        Delayed or decrease drug absorption by food can be due to one or more of the following reasons:

(a)    Delayed gastric emptying, affecting the drugs unstable in the stomach e.g. penicillin, erythromycin.

(b)   Preventing the transit of enteric tablets into the intestine which may be as long as 6 – 8 hrs.

(c)    Formation of poorly soluble, unabsorbable complex e.g. tetracycline-calcium complex.

(d)   Increased viscosity due to food thereby preventing drug dissolution and/or diffusion towards the absorption site.

·        Increased drug absorption following a meal can be due to the following reasons:

(a)    Increased time for dissolution of poorly soluble drug.

(b)   Enhanced solubility due to GI secretions like bile.

(c)    Prolonged residence time and absorption site contact of the drug e.g. water-soluble vitamins.

·        Types of meal

(i)     Meals high in fat aid solubilisation of poorly aqueous soluble drugs like griseofulvin.

(ii)   Food high in proteins increases oral availability of propranolol because

a) such a meal promotes blood flow to the GIT helping in drug absorption.

b) increases hepatic blood flow due to which the drug can bypass first-pass hepatic metabolism  (propranolol is a drug with high hepatic metabolism)     

5. Drug-drug interaction

Drug-drug interactions can be either physicochemical or physiological.

(a) Physicochemical drug-drug interactions can be due to –

Adsorption:         Antidiarrheal preparations containing adsorbents like attapulgite or kaolin-pectin retard / inhibit absorption of promazine and lincomycin when co-administered with them.

Complexation:    Antacids containing heavy metals such as aluminium, calcium, iron, magnesium or zinc retard absorption of tetracyclines due to the formation of unabsorbable complexes.

pH change:          Basic drugs dissolve in gastric pH. Co-administration of sodium bicarbonate with tetracycline results in evaluation of stomach pH and hence decreases dissolution rate or cause precipitation of drug.

(b) Physiologic drug-drug interaction can be due to following reasons:

Decreased GI transit:                Anticholinergic drugs such as propantheline retard GI motility and promote absorption of drugs like ranitidine and digoxin.

Increased gastric emptying:     Metoclopramide promotes GI motility and enhances absorption of tetracycline, pivampicillin and levodopa.

Altered GI metabolism:              Antibiotics inhibit bacterial metabolism of drugs e.g. erythromycin enhances efficacy of digoxin by this mechanism.

6. Presystemic metabolism / First pass effects

               The loss of drug through biotransformation by GIT and liver during the passage to systemic circulation in called First pass or presystemic metabolism.

The 4 primary systems which affect presystemic metabolism of drugs are:

1.      Lumenal enzymes

2.      Gut wall enzymes /mucosal enzymes

3.      Bacterial enzymes, and

4.      Hepatic enzymes

1. Lumenal enzymes

               These are enzymes present in the gut fluids and include enzymes from intestinal and pancreatic secretions.

·        Pancreatic enzymes contains hydrolases which hydrolyze ester drugs like chloramphenicol palmitate into active chloramphenicol.

·        Peptidases split amide ( –CONH) linkages and inactivate protein / polypeptide drugs. Thus one of the approaches is to deliver them to colon which lacks peptidases.

2. Gut-wall enzymes (also called mucosal enzymes)

               They are present in stomach, intestine and colon.

·        Stomach mucosa contains alcohol dehydrogenase (ADH) inactivates ethanol.

3. Bacterial enzymes

               The GI microorganisms are scantily present in stomach and small intestine and is rich in colon. Hence, most orally administered drugs remain unaffected by them.

·        The colonic microbes generally render a drug more active or toxic on biotransformation:

e.g. sulfasalazine (used in ulcerative colitis) is hydrolyzed to sulfapyridine and 5-amino salicylic acid by the microbial enzymes of the colon.

·        Digoxin, oral contraceptive drugs are absorbed in the upper intestine; exerted through bile as glucuronide conjugates. This conjugates of drugs are hydrolyzed by microbial enzymes. The free drugs are reabsorbed into the systemic circulation.

4. Hepatic enzymes

               e.g. isoprenaline, propranolol, alprenolol, pentoxyfylline, nitroglycerin, diltiazem, nifedipine, lidocaine, morphine etc.

C. PHARMACEUTICAL FACTORS

1. Disintegration time

               Disintegration time (DT) is of particular importance in case of solid dosage forms like tablets and capsules. After disintegration of a solid dosage form into granules, the granules must deaggregate into finer particles and then dissolution takes place. If DT is long the bioavailability will be less. Rapid disintegration is thus important in the therapeutic success of a solid dosage form.

               DT increases with increase in the amount of binder and hardness of a tablet.

               Disintegration can be aided by incorporating disintegrants in suitable amounts during formulation.

2. Manufacturing / process variables

Dissolution from a solid dosage form depends on:

(A) excipients and (B) manufacturing process.

(A) Excipients

               A drug is rarely administered in its original form. All dosage forms contains a number of suitable excipients (non-drug components of a formulation).

(a) Vehicle

               Vehicle or solvent system that carries a drug is the major component of liquid orals and parenterals. The three categories of vehicles generally used are:

(i) aqueous vehicles e.g. water, syrup etc.

(ii) nonaqueous but water miscible e.g. propylene glycol, glycerol, sorbitol.

(iii) nonaqueous and water immiscible vehicle e.g. vegetable oils.

               Bioavailability of a drug from vehicle depends, to a large extent, on its miscibility with biological fluids.

·        Aqueous and water miscible vehicles are rapidly miscible with body fluids (e.g. G.I.-fluid, tissue fluid, blood etc.) and drugs are rapidly absorbed from them.

·        Propylene glycol, glycerol etc. are used as co-solvent to increase the solubility of a drug in water. Sometimes solubilisers, such as Tween 80 are used to promote solubility of a drug in aqueous vehicle.

·        In case of water immiscible vehicles, the rate of drug absorption depends upon its partitioning from the oil phase to the aqueous body fluids, which could be a rate limiting step.

b) Diluents (Fillers)

               Diluents are commonly added to tablet (and capsules) formulations.

·        Hydrophilic powders used as diluent are starch, lactose, microcrystalline cellulose etc. These hydrophilic powders forms a coating over the hydrophobic drugs particles (e.g. spironolactone and triamterene) and rendering them hydrophilic.

·        Inorganic diluents like dicalcium phosphate (DCP) forms divalent calcium-tetracycline complex which is poorly soluble in water and thus unabsorbable.

c) Binders and granulating agents

               These materials are used to hold powders together to form granules or promote cohesive compacts for directly compressible materials and ensure that the tablet remains intact after compression.

·        Large amount of binders increase hardness and thus decrease disintegration / dissolution rates of tablets.

·        Non-aqueous binders like ethyl cellulose also retard dissolution.

d) Disintegrants

               These agents overcome the cohesive strength of tablet and break them up on contact with water.

·        Almost all the disintegrants are hydrophilic in nature.

·        A decrease in the amount of disintegrant can significantly lower the bioavailability.

e) Lubricants

               These agents are added to tablet formulations to aid flow granules, to reduce interparticular friction and to reduce sticking or adhesion of particles to dies and punches.

·        The commonly used lubricants are hydrophobic in nature (several metallic stearates and waxes). They reduce the wettability of particle surface, penetration of water into tablet.

·        The best alternative is to use soluble lubricants like sodium lauryl sulphate and carbowax which promotes drug dissolution.

f) Suspending agents /Viscosity building agents

               Agents like vegetable gums (acacia, tragacanth etc.), semisynthyetic gums (carboxy methyl cellulose, methyl cellulose) and synthetic gums which reduces the sedimentation rate of a suspension

·        The macromolecular gums often form unabsorbable complex with amphetamine.

·        An increase in viscosity by these agents acts as a mechanical barrier to the diffusion of drug from the dosage form into the bulk of GI fluids.

h) Surfactants

               Surfactants are widely used in formulations as wetting agents, solubilizers, emulsifiers, etc.

Surfactants increase the absorption of a drug by the following ways:

1.      Promotion of wetting (through increase in effective surface area) and dissolution of drugs e.g. Tween80 with phenacetin.

2.      Better membrane contact of the drug for absorption

3.      Enhanced membrane permeability of the drug .

Decreased absorption of drug in the presence of surfactants has been suggested to be due to :

1.      Formation of unabsorbable drug-micelle complex at surfactant concentrations above critical micelle concentration.

2.      Laxative action induced by a large surfactant concentration.

i) Complexing agents

Several examples where complexation has been used to enhance drug bioavailability are:

1. Enhanced dissolution through formation of a soluble complex

e.g.  ergotamine-caffeine complex

        hydroquinone-digoxin complex.

2. Enhanced lipophilicity for better membrane permeability e.g. caffeine-PABA complex (PABA = para amino benzoic acid) and

3. Enhanced membrane permeability e.g. enhanced GI absorption (normally not absorbed from the GIT) in presence of EDTA (ethylene diamine tetraacetic acid) which chelates Ca⁺⁺ and Mg⁺⁺ ions of the membrane.

Disadvantages of complexation:-

1.      complexation may produce poorly absorbable drugs complexes e.g. tetracycline with divalent and trivalent cations e.g.. tetracycline with divalent and trivalent cations like calcium (milk, antacids), iron (hematinics), magnesium (antacids) and aluminium (antacids).

2.      large molecular size of drug-protein cannot diffuse through the cell membrane.

j)  Colorants

               Even a very low concentration of water-soluble dye can have an inhibitory effect on dissolution rate of several

·        crystalline drugs. The dye molecules get adsorbed onto the crystal faces and inhibit drug dissolution – e.g. brilliant blue retards dissolution of sulphathiazole.

·        Dyes have also been found to inhibit micellar solubilizaion effect of bile acids which may impair the absorption of hydrophobic drugs like steroids.

(B) Manufacturing process

 (i) method of granulation and

(ii) Compression force

(iii) Intensity of packing of capsules

i) Method of granulation

               The wet granulation process is the most conventional technique of manufacturing tablet granules. The limitation of this method include –

(i)     formation of crystal bridge due to the presence of solvent,

(ii)   the liquid may act as medium or affecting chemical reactions such as hydrolysis, and

(iii) the drying step may harm the thermolabile drugs.

Wet granulation includes greater number of steps than dry granulation or direct compression which can adversely affect the dissolution.

ii) Compression force

               The compression force employed in tableting process influence density, porosity, hardness, disintegration time and dissolution of tablets.

               The curve obtained by plotting compression force versus rate of dissolution can take one of the 4 possible shapes shown in the figures:

A. Higher compression force ®    ­ density and hardness of tablet

¯ porosity, hence penetrability of the solvent into the tablet

¯ wettability by forming a firmer and more effective sealing layer by the lubricant

B. Higher compression force

®  causes deformation, crushing or fracture of drug particles into smaller ones or, convert a spherical granules into a disc shaped particle with large increase in effective surface area

® ­ in dissolution rate

C and D are combination of both the causes of A and B.

In short, the influence of compression force on the dissolution is difficult to predict.

(iii) Intensity of packing of capsule contents

               Packing density in case of capsule can either inhibit or promote dissolution.

·        Diffusion of GI fluids into the tightly filled capsules creates a high pressure within the capsule results in rapid bursting and dissolution of contents.

·        In some cases capsules with tight packing

® pore size of the compact mass is decreased

® poor penetrability of GI - fluid

® poor rate of drug release

NATURE AND TYPES OF DOSAGE FORM

Cause of events that occur following oral administration of various dosage forms:

As a general rule, the bioavailability of a drug from various dosage forms decreases in the following order:

Solution > Emulsions > Suspensions > Capsules > Tablets > Coated tablets > Enteric coated tablets > Sustained release tablets.

               Thus, absorption of a drug from solution is fastest with least potential for bioavailability problems whereas absorption from sustained release product is lowest with greatest bioavailability.

DRUG DISTRIBUTION IN THE BODY

Distribution Distribution is the reversible transfer of a drug between one compartment and other.

               Since the process is carried out by the circulation of blood, one compartment is always the blood or the plasma and the other represents extravascular fluids and other body tissues.

               Distribution of a drug is not uniform through out the body because different tissues receive the drug form plasma at different rates and to different extents. Differences in drug distribution among the tissues arise as a result of a number of factors as follows:

1. Tissue permeability of the drug

               (a) Physicochemical properties like molecular size, pKa and o/w partition coefficient.

               (b) Physiological barriers to diffusion of drugs.

2. Organ / tissue size and perfusion rate.

3. Binding of drugs to tissue components

               (a) Binding of drugs to blood components

               (b) Binding of drugs to extravascular tissue proteins.

4. Miscellaneous factors

               (a) Age                 (b) Pregnancy                    (c) Obesity

               (d) Diet                (e) Disease states                             (f) Drug interactions

TISSUE PERMEABILITY OF DRUGS

Two major rate-determining steps in the distribution of drugs are:

1. Rate of blood perfusion

2. Rate of tissue permeability

If the blood perfusion to the tissues are high then the tissue permeability will be the rate determining step in the process of distribution.

The tissue permeability of a drug depends upon the physicochemical properties of the drug as well as the physiological barriers.

(i)Physicochemical properties of the drugs

(a) Molecular size

                                                    MW < 500 daltons                                                  MW <50 daltons

                              Blood                                                                Extracellular                                                    Cells

                                                            Capillary                                 fluids                Cell membrane

                                                            membrane                                                         Water channels

Almost all drugs having molecular weight less than 500 to 600 daltons easily cross the capillary membrane to diffuse into the extracellular fluid (ECF).

Only small, water-soluble molecules and ions of size below 50 daltons enter the cell through water channels. Larger molecules are transported through specialized transport system existing on the cell membrane.

(b) Degree of ionisation

·        Blood and ECF pH normally remains constant at 7.4, they do not have much of an influence on drug diffusion unless altered in conditions such as systemic acidosis or alkalosis.

·        Most drugs are either weak acids or weak bases and their degree of ionization at plasma or ECF pH (i.e. 7.4) depends upon their pKa. All the drugs that ionize ar plasma pH (i.e. polar hydrophilic drugs) cannot penetrate the lipoidal cell membrane and tissue permeability is the rate determining step.

·        Only unionized drugs which are generally lipophilic, rapidly cross the cell membrane.

·        Species which has greater Ko/w (partition coefficient) penetrates well.

e.g. pentobarbital and  salicylic acid

               have same Ko/w but

               pentobarbital is more unionized at blood pH than salicylic acid and hence distributes rapidly.

e.g. thiopental, a nonpolar, lipophilic drug, largely unionized at plasma pH readily diffuses into the brain.

e.g. penicillins are polar, and ionized at plasma pH, hence does not cross the blood-brain-barrier.

(ii) Physiological barriers to distribution of drugs

Some of the important simple and specialized physiologic barriers are:

1.      Simple capillary endothelial barrier

2.      Simple cell membrane barrier

3.      Blood-Brain-Barrier (BBB)

4.      Cerebro Spinal Fluid Barrier (CSF Barrier)

5.      Placental Barrier

6.      Blood-Testis Barrier

1. The simple capillary endothelial barrier:

The membrane of capillary are unicellular in thickness; are practically no barrier for drugs having molecular weight under 600 daltons. Only drugs bound to blood components e.g. plasma protein, blood corpuscles are restricted due to large molecular size of the complex.

2. The simple cell membrane barrier

Once a drug diffuses from the capillary wall into the extracellular fluid, its further entry into cells of most tissues is limited by its permeability through the cell membrane that lines such cells.

The physicochemical properties that influence permeation of drug across such a barrier are summarized in the figure above.

3. Blood Brain Barrier

Unlike the capillaries found in other parts of the body, the capillaries in the brain are highly specialized and much less permeable to hydrophilic molecules.

The brain capillaries consist of endothelial cells which are joined to one another by continuous, tightly intercellular junctions comprising what is called as the blood-brain-barrier.

More over the glial cells and basement membrane forms a solid envelope around the brain capillaries. As a result, the intercellular passage is blocked and for a drug to gain access from the capillaries circulation into the brain, it has to pass through cells rather than between  them.

[N.B. However, there are specific sites where BBB does not exist, namely, in the chemo-receptor trigger zone, and the median hypothalamic eminence.

Drugs administered intransally may diffuse directly into the CNS because of the continuity between submucosal area of the nose and the submucosal  area of the nose and the subarachnoid space of the olfactory lobe.]

·        Since the BBB is a lipoidal barrier, it allows only the drugs having high Ko/w to diffuse passively.

·        Moderately lipid soluble and partially ionized molecules penetrate at a slow rate.

e.g. thiopental is 50 times more lipid soluble than pentobarbital and crosses BBB much more rapidly.

·        Polar, natural substances such as sugar, amino acids are tansported to brain actively.

e.g. For CNS disorders specialized drug molecules are administered. Parkinsonism is a disease caused by depletion of dopamine in the brain, but it cannot be treated by administration of dopamine as it does not cross the BBB. Hence, levodopa is given. It diffuses into the brain, metabolized there to produce dopamine.

4. Blood-cerebrospinal barrier

The cerebro-spinal fluid (CSF) is formed mainly by the choroidal plexus of the lateral, third and fourth ventricles and is similar in composition to the ECF of brain. The capillary endothelium that lines the choroid plexus have open junctions or gaps and drugs can flow freely into the extracellular space between the capillary wall and the choroidal cells. However, the choroidal cells are joined to each other by tight junctions forming the blood-CSF barrier which has permeability characteristics similar to that of the BBB.

·        Only highly lipid soluble drugs can cross the blood-CSF barrier with relative ease.

·        Moderately lipid soluble and partially ionized drugs permeate slowly. A drug that enters the CSF slowly cannot achieve a high concentration as the bulk flow of CSF continuously removes the drug.

5. Placental barrier

·        The maternal and fetal blood vessels are separated by a number of tissue layers made of fetal trophoblast basement membrane and the endothelium which constitute the placental barrier.

·        The human placental barrier has a mean thickness of 25 mm in early pregnancy that reduces to 2 mm at full term. Many drugs having molecular weight less than 1000 daltons and moderate to high lipid solubility e.g. ethanol, sulfonamides, barbiturates, gaseous anaesthetics, steroids, narcotic analgesic, anticonvulsants and some antibiotics, cross the barrier by simple diffusion quite rapidly. Hence, the placental barrier is not as effective as a barrier as that of BBB.

·        Nutirents essential for fetal growth are transported by carrier-mediated processes. Immunoglobulins transported by endocytosis.

·        Drugs are particularly dangerous to the fetus.

 6. Blood Testis Barrier

This barrier is located at the capillary endothelium level but at sertoli-sertoli cell junction. It is the tight junction between the neighbouring sertoli cells the act as blood-testis barrier. This barrier restricts the passage of drugs to spermatocytes.

ORGAN/TISSUE SIZE AND PERFUSION RATE

Perfusion rate is defined as the volume of blood that flows per unit time per unit volume of the tissue. It is expressed in ml (of blood)/min/ml (of the tissue).

Relative volume of different organs and tissues and their perfusion rates

(Assumption: Normally total body volume is 70 liters)

Organ/Tissue	% of Body volume	Perfusion rate  (ml blood/min/ml of tissue)
I. Highly perfused tissue                1. Lungs                2. Kidneys                3. Adrenals                4. Liver                5. Heart                6. Brain II. Moderately perfused tissue                7. Muscles                8. Skin III. Poorly perfused                9. Fat (adipose tissue)                10.Bone (skeleton)	0.7 0.4 0.03 2.3 0.5 2.0 42.0 15.0 10.0 16.0	10.2 4.5 1.2 0.8 0.6 0.5 0.034 0.033 0.03 0.02

k_t is the first-order distribution rate constant.

Now k_t is given by the following equation:L

where, K_t/b is the tissue/blood partition coefficient of a drug.

Extent of distribution:

The extent to which a drug is distributed in a particular tissue of organ depends upon the size of the tissue (i.e tissue volume) and the tissue/blood partition coefficient K_t/b .

Example:

Properties of thiopental:

               1.            Thiopental is a short acting anaesthetic. It is injected intravenously.

               2.            It is lipophilic drug and has affinity for both brain tissue and adipose tissue (fat). Blood-adipose                partition coefficient is higher than the blood-brain partition coefficient.

               3.            Brain is the site of action of thiopental.

Events

Cause

1.      When an i.v injection is given it shows rapid onset of action 2.      Short duration of action and rapid termination of action

1.      Thiopental diffuses rapidly into the brain. Adipose tissue being poorly perfused, takes longer to get distributed with thiopental 2.      As the concentration of thiopental in the adipose proceeds towards equilibrium, the drug rapidly diffuses out of the brain and localizes in the adipose tissue whose volume is more than 5 times that of brain and has greater affinity for the drug. The result is rapid termination of action of thiopental due to such tissue redistribution.

FACTORS AFFECTING DRUG DISTRIBUTION

Age

               Differences in distribution pattern of a drug in different age groups are mainly due to differences in –

(a) Total body water (both intracellular and extracellular) – is much greater in infants.

(b) Fat content – is also higher in infants and elderly.

(c) Skeletal muscles – are lesser in infants and in elderly.

(d) Organ composition – the blood brain barrier is poorly developed in infants, the myelin content is low and cerebral blood flow is high, hence greater penetration of drugs in the brain.

(e)  Plasma protein content – low albumin content in both infants and in elderly.

Pregnancy

               During pregnancy, the growth of uterus, placenta and fetus increases the volume available for distribution of drugs. The fetus represents a separate compartment in which a drug can distribute. The plasma and the extracellular fluid volume also increase but their is a fall in albumin content.

Obesity

               In obese persons, the high adipose tissue content can take up a large fraction of lipophilic drugs despite the fact that perfusion through it is low. The high fatty acid levels in obese persons alter the binding characteristics of acidic drugs.

Diet

               A diet high in fats will increase drugs such as NSAIDs to albumin.

Disease states

               A number of mechanisms  may be involved in the alteration of drug distribution characteristics in disease states:

a. Altered albumin and other drug-binding protein concentration

b. Altered or reduced perfusion to organs or tissues

c. Altered tissue pH.

Example: An interesting example of altered permeability of the physiologic barrier is that of blood brain barrier (BBB). In meningitis and encephalitis, the BBB becomes more permeable and thus polar antibiotics such as penicillin G and ampicillin which do not normally cross it, gain access to the brain.

Drug interactions

               Drug interactions that affect distribution are mainly due to differences in plasma protein or tissue binding of drugs.

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 tissues. The interacting molecules are generally the macromolecules such as proteins, DNS and adipose tissue.

               The phenomenon of complex formation with proteins is called protein binding of drugs.

Protein-drug binding: Binding of drugs to various tissue components and its influence on deposition and clinical response. Only the unbound drug moves reversibly between the compartments.

Bonds Involved In Protein Binding

Binding of drugs generally involves weak chemical bonds such as

1.      hydrogen bonds,

2.      hydrophobic bonds,

3.      ionic bonds, or

4.      vander Waal’s forces

and, therefore is a reversible process.

Irreversible drug binding, though rare, arises as a result of covalent binding and is often a reason for the carcinogenicity or tissue toxicity of the drug; for example chloroform (CHCl₃) and the metabolite of paracetamol binds irreversible with liver and thus results in hepatoxicity.

Binding of drugs falls into two classes:

1.  Binding of drugs to blood components like -

               (a) Plasma proteins

               (b) Blood cells

2.  Binding of drugs to extravascular tissue proteins, fats, bones, etc.

Of all types of binding, the plasma protein-drug binding is the most significant and most widely studied.

Binding of Drugs to Blood Components

Plasma Protein Binding

               The extent or order of binding of drugs to various plasma proteins is:

                              albumin > a₁Acid Glycoprotein > lipoproteins  >  globulins

Blood proteins to which drugs binds

Protein	Molecular weight	Concentration (g %)	Drugs that bind
Human serum albumin a1 - Acid glycoprotein Lipoproteins a1- Globulin a2- Globulin Hemoglobin	65,000 44,000 200,000 to 3,400,000 59,000 134,000 64,000	3.5 - 5.0 0.04 - 0.1 variable 0.003-0.007 0.015-0.060 11-16	large variety of all types of drugs basic drugs such as imipramine, lidocaine, quinidine, etc. basic, lipophilic drugs like chlorpromazine steroids like corticosterone, and thyroxine and cyanocobolamine (Vit. B12) vitamins A, D, E and K and cupric ions Phenytoin, pentobarbital and phenothiazines

Binding of drugs to blood cells

               More than 40% of the blood comprises of blood cells of which 95% is RBC. Thus significant RBC binding of drug is possible. The RBC comprises of 3 components:

1. Haemoglobin:                Phenytoin, pentobarbital and phenothiazines bind to haemoglobin.

2. Carbonic anhydrase :   Acetazolamide and chlorthalidone (carbonic anhydrase inhibitors)

3. Cell membranes :          Imipramine and chlorpromazine are reported to bind to RBC membrane.

Tissue binding of drugs

               Drug can bind to various tissues. Tissue-drug binding is important from two point of views:

(i)     It increase the volume of distribution (by reducing the concentration of free drug in the plasma) and

(ii)   drug bound to tissue acts as a reservoir and hence biological half life increases.

For majority of drugs that bind to extravascular tissues, the order of binding is : liver > kidney > lung > muscle.

1.      Liver: Oxidation products of carbon tetrachloride and paracetamol bind irreversibly with liver tissues resulting in hepatotoxicity.

2.      Kidneys: Metallothionin, a protein present in the kidneys, binds to heavy metals such as lead, mercury and cadmium.

3.      Lungs : Basic drugs like imipramine, chlorpromazine and antihistamines accumulate in the lungs.

4.      Skin : Chloroquine ad phenothiazines accumulate in the skin by interacting with melanin pigment.

5.      Hairs : Arsenicals are deposited in hair shafts.

6.      Bones : Tetracycline bind to bones and teeth. [N.B. Administration of tetracycline to infants or children during odontogenesis results in permanent brown-yellow coloration of teeth .]

7.      Adipose tissues : Lipophilic drugs such as thiopental and pesticide like DDT accumulate in adipose tissues (fat tissues).

8.      Nucleic acid : DNA interacts with drugs like chloroquine and quinacrine resulting in distortion its double helical structure.

Factors affecting protein binding

               Drug-protein binding is influenced by a number of important factors, including the following:

1. The drug

               Physicochemical properties of the drug

               Total concentration of the drug in the body

2. The protein

               Quantity of protein available for drug-protein binding.

               quality or physicochemical nature of the protein synthesized.

3. The affinity between drug and protein

4. Drug interactions

               Competition for the drug by other substances at a protein-binding site.

               Alteration  of the protein by a substance  that modifies the affinity of the drug for the protein

5. The pathophysiologic condition of the patient

               e.g. drug-protein binding may be reduced in hepatic diseases.

Kinetics of protein binding

Assumptions:       The drug-protein binding is reversible.

                              On the protein molecule one binding site is present

Under this condition the protein binding of drug may be described as follows:

               From the law of mass action

                               eqn (i)

where Ka is the association constant. Drugs strongly bound to protein have a very large Ka.

[  ] this symbol denotes molar concentration

To study the binding behavior of drugs, a ratio  ‘r’  is defined as follows:

hence,                             eqn. (ii)

Substituting  [PD] = Ka [P] [D] from eqn (i)  into eqn (ii) we get:

                                 eq. (iii)

Eqn. (iii) describes the situation where 1 mole of drug binds to one mole of protein in a 1 : 1 complex.

If drug molecules can bind independently to ‘n’ number of identical sites per protein molecule then the following equation may be used:

Significance of protein binding

1. Absorption

               From the absorption site the drug is absorbed to the blood. This absorption process will stop when free drug concentration at both sides become equal. If the drug is bound significantly to plasma protein then free drug in the plasma becomes less and hence the absorption process goes on. Thus much more amount of drug is absorbed.

2. Systemic solubility of drugs

               Water insoluble drugs, neutral endogenous macromolecules (such as heparin, steroids and oil soluble vitamins) are circulated and distributed to tissues by binding to lipoproteins.

3. Distribution

               Some drug may bind to a specific tissue and may produce toxic reaction to the tissue. Plasma protein binding restricts the entry of the drug into a tissue, thus saves the tissue. A protein bound drug does not cross the blood brain barrier, the placental barrier and the glomerulus.

4. Tissue binding, apparent volume of distribution and drug storage

               A drug that is extensively bound to blood components remains confined to blood and very little amount of drug will be available for distribution in the tissues. In this case the apparent volume of distribution (V_d) will be decreased.

               If the drug is bound to some tissue then the concentration of drug in the blood compartment will be less hence the V_d will be high.

               In both the cases the drug-protein complex will act as drug reservoir.

5. Elimination

               Only the unbound or free drug can be eliminated because the drug-protein complex cannot penetrate into the metabolising organ (e.g. liver). The large molecular size of the complex prevents it from filtration through glomerulus. Thus drugs which are more than 95% bound to protein eliminates slowly and the elimination half life will be prolonged.

6. Displacement interaction and toxicity

               If two drugs A and B, both have the same binding sites to plasma protein then one drug will displace the other. Thus the free drug concentration of both the drug in the plasma will rise and may precipitate toxic reaction. e.g. warfarin and phenylbutazone.

7. Diagnosis

               Thyroid gland (tissue) has great affinity for iodine. So any disorder of thyroid gland can be detected by administering compounds with radioactive iodine (I¹³¹)

8.  Therapy and drug targeting

               The binding of drugs to lipoproteins can be used for site specific delivery of hydrophilic moieties. e.g. in cancer therapy tumour cells have great affinity for LDL (low density lipoprotein) than normal tissues. Hence binding of suitable neoplastic agent to LDL can be used as a therapeutic tool.

Q1. What are the differences between passive and active transport?

Q2. Write short notes on: (i) Facilitated diffusion,  (ii) Phagocytosis and pinocytosis.

Q3. Discuss about the physicochemical factors on which absorption of a drug from the GIT depends.

Q4. Discuss about the physiological factors on which absorption of a drug from the GIT depends.

Q5. Discuss about the pharmaceutical factors on which absorption of a drug from the GIT depends.

Q6. What are the factors affecting drug distribution?

Q7. What is protein binding? What are the forces involved in protein binding? What are the factors affecting protein binding? What are the significance of protein binding?





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