INTRODUCTION TO BIOPHARMACEUTICAL SCIENCE

1. Pharmaceuticals, biologics and biopharmaceuticals

INTRODUCTION TO PHARMACEUTICAL PRODUCTS

 chemical synthesis plants finished product pharmaceutical facilities

HISTORY OF THE PHARMACEUTICAL INDUSTRY

THE AGE OF BIOPHARMACEUTICALS

recombinant DNA technology (genetic engineering) monoclonal antibody technology (hybridoma technology)

 It overcomes the problem of source availability It overcomes problems of product safety It provides an alternative to direct extraction from inappropriate/dangerous source material It facilitates the generation of engineered therapeutic proteins displaying some clinical advantage over the native protein product

BIOPHARMACEUTICALS: CURRENT STATUS AND FUTURE PROSPECTS

 most biopharmaceuticals approved to date are intended for human use At least 500 potential biopharmaceuticals are currently being evaluated in clinical trials the first generic biopharmaceuticals are already entering the market

TRADITIONAL PHARMACEUTICALS OF BIOLOGICAL ORIGIN

Pharmaceuticals of animal origin

The sex hormones

The androgens

Oestrogens
stimulation of the growth and maintenance of the female reproductive system (their principal effect); influencing bone metabolism; as is evidenced from the high degree of bone decalcification (osteoporosis) occurring in post-menopausal women; influencing lipid metabolism.

Progesterone and progestogens

Corticosteroids

Catecholamines

adrenaline
emergency management of anaphylaxis; emergency cardiopulmonary resuscitation; addition to some local anaesthetics (its vasoconstrictor properties help to prolong local action of the anaesthetic).

Prostaglandins

Pharmaceutical substances of plant origin

Pharmaceutical substances of microbial origin

generalized penicillin structure

tetracycline

aminoglycoside antibiotics

2. MOLECULAR PROPERTIES OF DRUGS pharmacodynamics pharmacokinetics

drug dose-response profile

drug dose-response diagram

Main molecular properties of drugs

Partition coefficient Dissociation constant (ionization state) Solubility Chemical stability

ADME
Drug administration: How is the drug to be formulated?

Drug absorption: can the drug pass through the barrier membranes in the gastro-intestinal tract? Can it pass through the skin barriers?

 Drug metabolism: metabolism increases the water solubility of drugs by enzymatically introducing polar functional groups so that they can be excreted

Drug excretion: the kidney excretes watersoluble metabolites.

 Drug action: the shape of the drug, its chemistry and its compatibility with the target receptor/enzyme determines the extent of the response

PARTITIONING
Affinity for water: HYDROPHILIC (water loving) or LIPOPHOBIC (lipid hating) Affinity for oil: LIPOPHILIC (lipid loving) or HYDROPHOBIC (water hating)

the partition coefficient

 The standard solvent for partition measurements of medicinal compounds is octan-1-ol

Positive and negative effects on log P

Fragmental constants: quantification of substituent effects on partitioning

Aromatic compounds Benzene: 2.13 Toluene (methylbenzene): 2.79 Ethylbenzene: 3.45 Aliphatic compounds

Exceptions and provisos

THE ACIDITY AND BASICITY OF ORGANIC COMPOUNDS

The ionized/charged forms of drugs (salts) tend to dissolve in water and they will not cross lipid membranes The unionized/uncharged forms (free acids or bases) tend to dissolve in organic solvents and will cross lipid membranes

 For a drug to be administered, it needs to dissolve in an aqueous medium if it is to be given orally or by injection (it needs to be ionized/charged).

For a drug to be absorbed through a lipid membrane, it needs to partition from the aqueous to the lipid medium (it needs to be unionized/uncharged and lipophilic).

 For a drug to be transported around the body, it needs to dissolve in the aqueous plasma (it needs to be ionized/charged).

Aspirin, pKa = 3.5

Definitions
ACIDS ARE PROTON DONORS

 BASES ARE PROTON ACCEPTORS

 THE HIGHER THE pKa VALUE,

THE FURTHER THE EQUILIBRIUM LIES TO THE LEFT i.e. THE SPECIES PREFERS TO BE UNDISSOCIATED RATHER THAN DISSOCIATED

BENZYLAMINE HYDROCHLORIDE

DIPHENYLAMINE HYDROCHLORIDE

Benzoic acid has a pKa 4.2. What percent is dissociated in a solution of pH 4.2 ?

 Acids and conjugate acids can have any numerical value of pKa, but only values less than 14 will be of any significance in determining the solubility of acids in water and only values greater than zero will be of significance in determining the solubility of conjugate acids.

 ONLY THE UNIONISED FORM OF A DRUG CAN PARTITION ACROSS BIOLOGICAL MEMBRANES (providing the unionized form is lipophilic) [required for drug absorption into the body] THE IONISED FORM TENDS TO BE MORE WATER SOLUBLE [required for drug administration and distribution in plasma]

RCOOH, pKa = 4.0

a basic drug, pKa = 7.0

The effect of the Partition Coefficient on Drug Distribution

For an acid substance

Papp: apparent Partition coefficient

pH at roughly equal distribution

RCOOH, pKa = 4.0; P = 200

Papp = 198 in the stomach

FACTORS AFFECTING ACID AND BASE STRENGTH

The ability of a compound to exist as an acid depends on the ability of the anion formed by dissociation to stabilise the negative charge !

Carboxylic acids

Effect of R on pKa?

inductive effect of alkyl groups

mesomeric effects

Phenols

Sulphonamides

Sulphonic acids

Molecular Structures of Bases

no drug is basic without nitrogen in its structure

Aromatic Amines

Amides

Amidines and Guanidines

Diacidic bases (two nitrogens without charge sharing)

-Amino Acids

zwitterion

Heterocycles

 - deficient heterocycles

' -excessive' heterocycles

ANALYTICAL MEASUREMENT IN MEDICINE AND DRUG DEVELOPMENT

The Control Of Errors In Analysis

 A batch of paracetamol tablets are stated to contain 500 mg of paracetamol per tablet. Four students carry out a spectrophotometric analysis of an extract from the tablets and get the following percentages of stated content for the repeat analysis of paracetamol in the tablets:

Accuracy And Precision In Analytical Measurement

Dilutions And Weights And Measures

A sodium chloride infusion is diluted as follows. 5 ml to 100 ml followed by 5 ml to 250 ml. The sample is then analysed and is found to contain 1.03 mg/100 ml of sodium chloride. Calculate:

1. The concentration of sodium chloride in the original solution in % w/v. 2. The amount of sodium chloride in mg present in 5 ml of the original solution.

APPLICATIONS OF SPECTROPHOTOMETRY IN BIOPHARMACEUTICAL SCIENCE

Spectrophotometry is a group of analytical techniques that are used to provide information about the nature (i.e. the identity) and composition (i.e. amount) of substances in a sample.

Ultraviolet/visible absorption spectrophotometry

the measurement of the amount of light absorbed by the drug substance in solution in the ultraviolet and visible regions of the spectrum

 What Is Electromagnetic Radiation?

a form of energy whose behavior is described by the properties of both waves and particles

Red: 780 nm, Orange: 620 nm, Yellow: 585 nm, Green: 570 nm, Blue: 490 nm, Indigo: 440 nm, Violet: 420 nm

blue is minus yellow green is minus magenta red is minus cyan yellow is minus blue magenta is minus green cyan is minus red

Quantitative UV-Vis Spectrophotometry

When making a quantitative spectrophotometric measurement of a solution of a substance at a particular wavelength, two light intensities are measured, viz. the intensity passing into the solution (Io) and the (reduced) intensity (It) transmitted from the solution after absorption of some of the light has occurred.

The Beer Lambert Law

A = abc where A: the absorbance, a: the ABSORPTIVITY (a proportionality constant), b: the pathlength c: the concentration of the solution

 A=bc where c is in moles/l and b is in cm : the molar absorptivity

A = A(1%, 1cm) b c where c is in g/100 ml and b is in cm A(1%, 1 cm): the specific absorbance

 What is the absorbance at a particular wavelength if the % transmittance of a solution is 10% at that wavelength? What % of the light at a particular wavelength is absorbed by a solution having an absorbance of 0.5 at that wavelength?

What Happens When Light Is Absorbed In The UV-Vis Region?

UV/Vis Spectra for Molecules

Energy quantification of a molecule

with Eelec >Evib>Erot

 The absorption of UV/visible radiation occurs through the excitation of electrons within the molecular structure to a higher energy state; These transitions occur from the lowest vibrational state in the electronic ground state of the molecule to any one of a number of higher vibrational levels in the electronic excited state.

What Type of Molecules Absorb UV/Visible Radiation?

Extended Chromophores
A CHROMOPHORE is group that is composed of a series of double bonds (at least two) separated by single bonds

The Benzene Ring Chromophore

Auxochromes
Groups containing nitrogen and oxygen, which have non-bonding electron pairs, also interact with an extended chromophore such as the benzene ring. To create bathochromic/hyperchromic shifts auxochromes have to be attached directly to the system i.e. directly on one of the double bonds in the extended chromophore.

Instrumentation in the UV/Visible

Applications of UV/Visible Spectrophotometry in Pharmaceutical Analysis

Quantification Based on Comparison with a Standard Solution (i.e. solution of known concentration)

 Some Calculations Using Standard A (1% 1cm) Values of the Drug

 Assay of paracetamol tablets. Weigh and powder 20 tablets. Add a quantity of powder containing about 0.15 g of paracetamol to 50 mL of 0.1 m NaOH, dilute with water to 100 ml and shake for 15 min. Add sufficient water to make up to 200 ml. Filter the solution and dilute 10 ml of the filtrate to 100 ml with water. Add 10 ml of the resultant solution to 10 ml of 0.1 M NaOH and dilute to 100 ml with water. Measure the absorbance at 257 nm. The A(1%,1cm) value of paracetamol at 257 nm is 715. Experimental results The measured absorbance = 0.520. Weight of 20 tablets = 11.413 g The weight of tablet powder taken = 0.1657 g. The stated content of paracetamol is 500 mg per tablet. Calculate the % of stated content in the tablets.

Determine pKa of Drug Substances

 The absorbance of a fixed concentration of phenylephrine at 292 nm is found to be 1.224 in 0.1 M NaOH and 0.02 in 0.1 M HCl. Its absorbance in buffer at pH 8.5 is found to be 0.349. Calculate the pKa value of its acidic phenolic hydroxyl group.

Determine Partition Coefficients

A sample of a neutral drug (5 mg) is dissolved in 5 ml of water and then the water is shaken with 5 ml of n-octanol. A portion of n-octanol (2 ml) and is withdrawn and is diluted to 50 ml with methanol. The absorbance reading for the diluted sample is 0.657. If the A(1%,1cm) value for the drug is 181 calculate its n-octanol/water partition coefficient.

The Effect of pH on Partitioning

CHEMICAL STABILITY OF PHARMACEUTICALS

Decomposition products of the drug may be far more toxic to the patient than its parent drug Significant decomposition of the drug in its dosage form (e.g. tablets) will reduce the effective dose to the patient

 Up to 10% degradation may be acceptable provided the decomposition products are not toxic to the patient !

Isoniazid

hydrazine (< 0.05%)

 expiry date: the Medicines Act 1968

The limiting factor in the setting of the expiry date is the chemical stability or other component of the medicine

adverse conditions
Heat Light Moisture For substances affected by water, the effect of pH will also be investigated Air (oxygen)

HYDROLYSIS OF ESTERS
The general reaction for acid-catalysed or base-catalysed hydrolysis:

saponification

Lactides and lactones

Rates of Hydrolysis

HYDROLYSIS of AMIDES

AUTOXIDATION
Autoxidations are usually radical-induced chain reactions

Typical pharmaceutical examples

Adrenaline

Inhibition of Oxidation

5. Use an antioxidant
Oxygen scavengers

Ascorbic acid

 Chain terminators

Typical antioxidants: thymol, butylated hydroxyanisole, butylated hydroxytoluene, propyl gallate, octyl gallate and dodecyl gallate

THE SHAPES OF ORGANIC MOLECULES

drug molecules interact with optically active, asymmetric biological macromolecules such as proteins, polynucleotides, or glycolipids acting as receptors, stereochemical specificity

Stereochemistry
Isomers: identical molecular formulae but different in the nature or sequence of bonding of their atoms or in arrangement of their atoms in space

 constitutional isomers: the order of atomic connections that defines a molecule

Stereoisomers: different in the three dimensional arrangement of atoms in space

Configuration
The configuration of a molecule of defined constitution is the arrangement of its atoms in space without regard to arrangements that differ only after rotation about single bonds.

 Geometric isomers are configurational isomers that result from the presence of double bonds or rings that impose rigidity on the molecule thus preventing free rotation about certain bonds

 Configurational isomers that are mirror images of each other are termed enantiomers Chirality

Optical activity
Enantiomers have identical physical properties except in one respect, they rotate the plane of polarized light in opposite directions Molecules that rotate polarized light are said to be 'optically active indicated by (+) or (-) before the name of the substance

 asymmetric atoms: tetrahedrally bonded to four different atoms or groups (chiral centre)

Racemic: equal amounts of enantiomeric molecules are present together

Importance of Chirality in Drug Therapy

Exercises!

INTRODUCTION TO PHARMACOKINETICS AND PHARMACODYNAMICS

 Drugs may be defined as chemicals that alter physiological or biochemical processes in the body in a manner that makes them useful in the treatment, prevention, or cure of diseases.

 the onset, intensity, and duration of therapeutic effects for a particular disease condition dose, frequency of administration, route of administration

PHARMACODYNAMICS
Drug Effects at the Site of Action

Interaction of a Drug with Its Receptor

Postreceptor Events

 Agonists, Antagonists, and ConcentrationResponse Relationships

 A drug that mimics the endogenous receptor ligand to activate the receptor is referred to as an agonist

 A drug that binds to a receptor but does not activate it is referred to as an antagonist

PHARMACOKINETICS
Pharmacokinetics (PK) is the study of drug movement into, around, and out of the body. By extension it involves the study of drug absorption, distribution, and elimination (metabolism and excretion) (ADME).

Plasma Concentration of Drugs

The goal of pharmacokinetics

Identifying the drug and patient factors that determine the rate and extent of each process

 Identifying a way to quantify or summarize each process in ADME using a single parameter

Basic Pharmacokinetic Considerations

Effective and toxic plasma concentrations; The pharmacokinetic parameters which reflect the physiological processes in the body; (Bio)Availability; Volume of distribution; Half-Life.

Pharmacokinetic Representations of Absorption, Distribution and Elimination

One-Compartment Open Model: Intravenous Bolus Administration

DB: drug in body VD: apparent volume of distribution; k: elimination rate constant.

Elimination Rate Constant

D B = at time t and D B 0 = at t = 0

Two-Compartment Open Model

Volume of Distribution