2. Pre-formulation studies
• Evaluation of drug prior to formulation development → pre-
formulation studies.
Why pre-formulation studies required?
• Drug discovery and development → complex, lengthy process
• Failure of a molecule during development stage
• Reasons →poor pharmacokinetics, lack of efficacy, and/or toxicity
3. Significance of pre-formulation studies
• Understanding the physical and chemical properties of a drug helps to
formulate the crude drug into a suitable dosage form/ various dosage
regimens etc.
• Lead to simple and elegant formulations/ successful commercial products
Provides the scientific basis for formulation development
Classification of pre-formulation studies
(i) Fundamental properties →specific to drug, chemical structure
(ii) Derived properties→ related to development of specific dosage form
4. Preliminary pre-formulation study and molecular optimization
Active pharmacological compound invention
Preliminary studies on suspected problem area
Development into its optimum molecular state
Stability and solubility→ molecular modification
E.g. salts, pro-drug, solvates, polymorphs, or even new
analogs
5. Salt approach
• Solubility problem is resolved by converting into salt form
• Compound having ionisable group are converted into salt
• Ephedrine →Ephedrine hydrochloride, ↑solubility/ dissolution/
bioavailability
Pro-Drug approach
• Prodrugs are synthetic derivatives (esters, amides) → transformation
in vivo to liberate the active drug molecule.
• Erythromycin is water soluble, has a bitter taste, and is rapidly
hydrolyzed in gastric acid
• Erythromycin estolate, the lipophilic ester prodrug , absorbed 4 times
more efficiently, but hydrolyzed only 24% in serum.
6. Initial / preliminary solubility investigation
Total solubility (cs)
• Total solubility (Cs) is solubility of ionized + unionized form of the drug
• Method→ Equilibrium solubility method
• Procedure: Excess solute + solvent → agitate at constant temperature until
equilibrium, chemical analysis of drug
Intrinsic solubility (Co)
• Intrinsic Solubility (Co) is solubility of only unionized form of the compound.
• The solubility obtained in acid (0.1 N Hcl) for a weak acid
• The solubility obtained in alkali (0.1 N NaOH) for a weak base
• Unit →the number of moles of solute per litre that dissolves into solution at 25oc.
7. Effect of pH on solubility of drug
• Weak acid solubility increases with increase in pH
• Weak base solubility increases with decrease in pH
• Amphoteric drug solubility increases with increase/ decrease in pH
• Neutral drug no change in the solubility with change in pH
The solubility should ideally be measured at two temperatures
Max. density of water is at 4 o C – to check mini. aqueous solubility.
37 o C to support biopharmaceutical evaluation.
8. Solubility in various pH
Every new drug must be determined as a function of pH ,why?
• Over the physiological pH range of 1-8.
• For relatively insoluble drugs , rate of dissolution depends on its
solubility which can influence its biological absorption.
• Stability during processing
Henderson -Hassel Bach equation to find ratio of ionised/unionised drug
• pH = pKa + log [(un-ionized drug) / (ionized drug)]→basic drug
• pH = pKa + log [(ionized drug) / (un-ionized drug)]→acidic drug
9. Stability studies
(i)Solid state stability, (ii)Solution phase stability → ICH guidelines
Solid state stability
Physical stability
Polymorphism, hygroscopicity
Factors influencing physical stability
Solubility, pKa, melting point, crystal structure , equilibrium moisture content
Chemical stability
Degradation, hydrolysis, oxidation, photo-degradation….
Factors influencing chemical stability → heat, light, moisture, oxygen
10. Effect of temperature on stability
• Elevated temperatures like 400C, 500 and 600 etc. / ambient humidity
• Are compared with a control for any physical /chemical change.
• The data obtained may be extrapolated by Arrhenius treatment
• The degradation rate at a lower temperature is determined
ICH guidelines
Long term→ 25± 2°C/60% ± 5% RH or 30 ± 2°C/65RH ± 5% RH for 12 months
Intermediate→ 30 ± 2°C/65% ± 5% RH for 6 months
Accelerated → 40 ± 2°C/75% ± 5% RH for 6 months
11. Stability studies under high humidity conditions
The presence of high humidity conditions
some drugs tend to hydrolyse
Some tend to react with other excipients
Some drugs tend to oxidize.
• During stability study the drug is exposed to different relative humidity
conditions
• Controlled humidity conditions may be obtained by using laboratory
desiccators.
Application
To find the material is to be protected/ stored in a controlled low humidity
environment
Formulation planning
Eg. An aqueous based granulation system is to be avoided or not.
12. Photolytic stability
• Many drug will fade / darken on exposure to light.
• The extent of degradation depends on the exposed surface area
• Can be overcome→ protection of the drug using an amber coloured
container / a photo stable dye in the system/dark place/physical
barriers
• For confirmatory studies,
• Samples are exposed to light providing an overall illumination of
• Not less than 1.2 million lux hours and an integrated near ultraviolet
energy of not less than 200 watt hours/square meter .
• Samples may be exposed side-by-side with a validated chemical
actinometric system.
13. Stability to oxidation
• Testing its stability in an atmosphere of high oxygen tension usually at
40% oxygen atmosphere.
• Results are compared against a standard stored under the inert or
ambient conditions.
Application of test for Sensitivity to oxygen
• To decide atmospheric conditions for product packing
• To Include an anti-oxidant in the formulation
Stability to pH
• The pH stability profile data helps in predicting solid state stability of a drug in
the presence of acidic and basic impurities.
14. SOLUTION PHASE STABILITY
• Why solution phase stability is required ?
Stability of the drug from pH ranging from 1 to 8 is to be investigated
To assure that the substance does not degrade when exposed to GI
fluids.
It is important for the selection of granulation solvent and the drying
conditions.
15. Developing a Profile of the Active Pharmaceutical Ingredient
When preliminary pre-formulation study assures the solubility/ stability of compound
The best (optimum) molecular form of a drug has been selected
Physicochemical characterization begins
Following parameters are to be studied further
Fundamental Properties
• Physical description/Organoleptic properties
• Analytical methods, Purity
• Partition coefficient, ionization constant
• Crystal properties and polymorphism
• Dissolution, Permeation across the membrane
Derived Properties
Compatibility Studies : Stability in the presence of excipients
16. PHYSICAL DESCRIPTION/ORGANOLEPTIC PROPERTIES
Colour, physical appearance, taste and odour
The colour
• attributes of a drug substance will be indicative of stability problems,
• May indicate the necessity of improving the appearance by including a dye
in the body or the coating of the final product.
Taste
• May indicate palatability problems
• Excipients and flavours may also be used to obtain desired taste.
Odour
• Is indicative of degradation products ( Aspirin hydrolysis imparts odour of
acetic acid).
• Also may indicate the need of flavours and excipients in the formulations
for required odour.
17. ANALYTICAL METHODS: UV SPECTROSCOPY
• The first requirement , the development of a simple analytical method for
quantitative estimation in subsequent steps.
• Most of drugs have aromatic rings and/or double bonds as part of their
structure and absorb light in UV range.
• UV spectroscopy being a fairly accurate and simple estimation technique
• The absorption Co-efficient (E) of the drug can be determined by the formula:
E = AF / X A=Absorbance, F = dilution factor, X = weight of drug (mg)
• It is now possible to determine concentration of drug in any solution by
measuring absorbance
C = AF/E mg/ ml
18. Purity
• The determination of the purity and the data will help further studies to
be carried out safely.
• All the impurities present in the compound should be within the range
specified.
• Impurities may affect the stability of the compound and also the
appearance of the compound.
• Some impurities may make the compounds toxic.
• The techniques used in the purity studies are TLC, HPTLC, paper
chromatography, HPLC, XRD and GC.
19. Dissolution
• Dissolution precedes absorption
• The dissolution behaviour should be investigated
• To make necessary changes to enhance the dissolution rate /to select
an optimum form for its development.
20. Dissolution rate and Noyes-Nernst equation
dW/dt = dissolution rate
A = surface area of the dissolving solid
D=diffusion coefficient
V= volume of the medium,
h = thickness of the diffusion layer
Cs =concentration of solute in diffusion layer,
C =solute concentration in the bulk medium
22. The intrinsic dissolution rate (IDR)
• The dissolution rate of a pure active substance, where the conditions
of surface area, temperature, agitation, and medium pH and ionic
strength are all constant.
• Using a specific device for this purpose, where the compressed drug
is exposed in a dissolution medium over a constant surface area, and
its value is expressed in mg cm-2 s-1.
• IDR data helps to obtain the chemical purity and equivalence of drugs
from different sources.
23. Parameters Affecting Absorption
The absorption of drugs consists of 2
processes;
The dissolution of the drug molecules
Its transport across the GI/biological
membrane into the systemic
circulation.
For the relatively soluble
compounds, the rate determining
step is the permeation across the
membrane
For the relatively insoluble
compounds, the rate determining
step is the rate of dissolution.
The three main parameters that influence the rate of absorption are
partition coefficient, ionization constant and the permeation across the
membrane.
24. Partition coefficient
• Partition coefficient→ the ratio of the concentration of the drug in
the oil phase to that in the aqueous phase.
• n-octanol is used as the oil phase / water as the aqueous phase
• Other organic solvents→ chloroform, ether, amyl acetate etc.
Method
• Known amount of drug + two phases → equilibrating, separation of
the phases, analysis of drug
26. Ionization constant
• Most drugs are weak acids or bases ,
• They exist as ionized or unionized species based on the pH of the
medium.
• Unionized species are easily absorbed as they are lipid soluble.
The pH partition theory includes three principles
1. The pH as the site of absorption,
2. Ionization constant
3. Lipid solubility of the drug
27. The relative concentration of the ionized and the unionized form
Henderson Hassel Bach equation
pH = pKa + log [unionized form] / [Ionized form] FOR BASES
pH = pKa + log [ionized form]/ [unionized form] FOR ACIDS
Ratio can be determined
• Potentiometric pH-Titration,
• pH-Spectrophotometry Method,
• pH-Solubility Analysis.
28. Permeation across the membrane
• in vitro experiments using biological membranes.
• The method used is the everted rat gut method.
• This technique determines the degree & rate of passage of drug
through the membrane sac by passive & active transport.
29. Crystal properties and polymorphism
• Drug can exist in more than one crystalline from with different space
lattice arrangements → Polymorphism
• Pre-formulation activity → Study on Crystal structure, polymorphism,
and solvate form
Changes in crystal characteristics can influence
bioavailability
chemical and physical stability
in dosage form process functions like flow and compaction behaviours
in tablet dosage form
Techniques are used to determine crystal properties
• Hot stage microscopy, thermal analysis, infrared spectroscopy, and X-
ray diffraction.
30. Derived Properties
• Derived pre-formulation properties are carried out
• to learn about the issues related to development of a particular
dosage form
• like solid oral, liquid oral or parenteral.
31. Derived properties for solid dosage form
• Particle size, shape and surface area
• Powder Flow Properties
• Density
• Hygroscopicity
• Compactibilty and compressibility
• Wettability (for Suspension also)
32. Excipient selection
• Chemical structure of the API
• Its physicochemical characteristics
• The type of delivery system required
• The proposed manufacturing process.
33. Compatibility studies
• Compatibility studies are usually the last activity
• Studying the interactions of drug substance with other excipients
Need of compatibility study
• for careful selection of the excipients
• To facilitate administration
• To promote the consistent release and Bioavailability of the drug
• To protect it from degradation.
34. Methods used to study DEC
• DSC- Differential Scanning Calorimetry
• DTA- Differential Thermal Analysis,
• FT-IR Spectroscopy,
• DRS-Diffuse Reflectance Spectroscopy,
• TLC-Thin Layer Chromatography,
• HPLC-High Pressure Liquid Chromatography,
• XRD
35. Characteristics of Powder XRD and its interpretations
• Results of the PXRD procedure are commonly presented as
• Peak positions at 2θ and X-ray counts (intensity)
• in the form of a table or a x-y plot,
• known as diffractogram.
36. Characteristics of Powder XRD and its interpretations
• It is reliable and powerful tool for crystalline sample identification.
• It is a non-destructive technique.
• Each crystalline substance has a unique X-ray diffraction pattern.
37. Crystal structure
• Crystal structure is a description
of the ordered arrangement
of atoms, ions or molecules in a
crystalline material.
• Crystal lattice is the periodic and
systematic arrangement of
atoms.
• The smallest portion of a crystal
lattice is called Unit Cell.
38. Crystal structure and X-RAY diffraction pattern
• The crystallographic planes are
geometric planes linking
atoms/ions/molecules in crystal
lattice.
• Given any plane in a lattice, there is
an infinite set of parallel lattice
planes (family of planes) that are
equally spaced (d value) from each
other.
• More the number of planes in given
family greater is the intensity of
peaks
39. X-Ray diffraction pattern
• Higher the intensity greater the
crystallinity
• Their widths are rather dependent on
particle size (overly small particles will
result in line broadening) i.e. broad
nature of peak indicates higher degree
of amorphousness or less crystalline
in comparison with other samples
• The number of observed peaks is
related to the symmetry of the unit
cell (higher symmetry generally means
fewer peaks).
40. DIFFERENTIAL SCANNING CALORIMETRY (DSC)
• DSC is a technique in which the
difference in energy inputs into a
substance and a reference
material is measured as a
function of temperature while
the substance and reference
material are subjected to a
controlled temperature program.
• The graph of heat flow in mJ/s on
the Y-axis plotted versus
temperature at a fixed rate of
change of temperature in °C on
the X-axis shows the output of
the DSC.
41. Application and Interpretation of DSC curves
• To check the Purity ,
• For polymorph identification,
• To check decomposition
44. FTIR spectra
• The Functional Group Region (4000-1500 cm-1)
• The Fingerprint Region (1500-400 cm-1)
45. Characteristics of IR spectra and its interpretation
• Used as a supplementary technique to investigate the DEC and to confirm
the results obtained by the thermal analysis.
• The appearances of new absorption band, broadening of band, and
alteration in intensity are the main characteristics to evidence DEC.
• IR is most useful in providing information about the presence or absence of
specific functional groups.
• Different bonds have diff. frequencies Eg; C-C, C=C, C ≡ C, C-O, O-H, N-H
• IR does not provide detailed information or proof of molecular formula or
structure.
• Hydrogen bonding (Physical interaction) and shift in IR peaks: The
number and strength of hydrogen bonds differs with chemical
environment. The force constant varies and the wavenumber differs at
which these molecules absorb infrared light. The number and strength of
intermolecular interactions varies greatly within the sample, causing the
bands in these samples to be particularly broad.
