This document discusses drug discovery from natural products. It begins by defining drug discovery as the process of discovering new candidate medicines. Natural products are highly valuable for drug discovery due to their chemical diversity and other favorable molecular properties. The document then outlines the various steps involved in drug discovery from natural products, including collecting source organisms, preparing initial extracts, preliminary biological screening, purification methods, structure elucidation, compound development, and examples of natural product-derived drugs currently in medical use.

1. DRUG DISCOVERY FROM NATURAL PRODUCTS
Presented by:
Abdul Rauf
Khaqan Saleem

2. INTRODUCTION
• What is Drug discovery?
• The process by which new candidate medicines are discovered is
called drug discovery.

3. ROLE OF NATURAL PRODUCTS IN DRUG DISCOVERY
Natural products have characteristics of:
• High chemical diversity
• Biochemical specificity
• Other molecular properties that make them favorable for drug discovery

4. a) Collection of source organisms:
• There are at least five recognized approaches for the collection of
source organisms for the laboratory investigation of their biological
components.
• 1. Random Screening
• 2. selection of specific taxonomic groups
• 3. Chemo taxonomic approach
• 4. information managed approach
• 5. Selection by ethno medical approach

5. TYPES OF SOURCES OF NATURAL PRODUCTS FOR DRUG DISCOVERY:

6. Sources of natural products for drug discovery:
• Animal source ( e.g.: insulin, thyroxin)
• Plant source(e.g.: vincristine, pilocarpine)
• Microbial source ( e.g. : penicillin, streptomycin)
• Marine source (Avarol, which is reported to be an immunodeficiency virus
inhibitor)

7. b) Preparation of Initial extract:
• Extraction means the separation of medically active portions of plant
or animal tissues from the inactive or inert components by using
selective solvents.
• Extract can be defined as:
• “ Preparations of crude drug which contain all the constituents which
are soluble in the solvent.”

8. Methods of extraction:
• Infusion
• Decoction
• Maceration
• Percolation
• Reflux extraction

9. INFUSIONS
Fresh infusions are prepared by macerating the crude drug for a short
period of time with cold or boiling water. These are dilute solutions of
the readily soluble constituents of crude drugs.
• Types of Infusion :
 Fresh Infusion : e.g. Infusion of orange
 Concentrated Infusion : e.g. Concentrated infusion of Quassia

10. DECOCTION
• Decoction is a method of extraction by boiling herbal or plant
material to dissolve the chemicals of the material, which may
include stems, roots, bark and rhizomes.
• In this process the crude drug is boiled in a specified volume of
water 1:4 for a defined time. Volume is reduced to 1/4th the
original. It is then cooled and filtered.
• It is used for water soluble and heat stable constituents
E.g. tea and coffee

11. MACERATION
• In this process solid ingredients are placed in a
stoppered container with the whole of the solvent
and allowed to stand for a period of at least 3 days
(3 - 7 days) with frequent agitation, until soluble
matter is dissolved.
• The mixture is then strained (through sieves /
nets), the marc pressed and the combined liquids
clarified by filtration or decantation, after standing.

12. PERCOLATION
 It is continuous downward displacement of the solvent through the
bed of crude drug material to get extract.

13. REFLUX EXTRACTION
• Reflux extraction is a solid–liquid extraction process at a constant
temperature with repeatable solvent evaporation and condensation
for a particular period of time without the loss of solvent.

14. Modern methods of extraction
o Soxhletextraction
o Pressurized liquid extraction
o Supercriticalfluidextraction
o Ultrasound assisted extraction
o Microwaveassistedextraction
o Pulsed electric field extraction
o Enzyme-AssistedExtraction

15. 1. Soxhlet extraction
• This extraction is used where compound has limited solubility in a solvent and
impurity is insoluble in solvent.
• Solid material containing some desired components is placed inside a thimble
made from thick filter paper. Which is loaded in the main chamber of soxhlet
chamber. The extractor is then placed in a flask containing extraction solvent.
Soxhlet is then equipped with condenser
• The solvent is heated to reflux. Solvent vapours travel up the distillation arm
condensed by condenser and travels back into the chamber holding the thimble
of solid.
• Chamber filled with warm solvent and dissolve some of desired components. The
chamber is automatically emptied down the distillation chamber by siphon side
arm.

17. 2. Pressurized liquid extraction(PLE)
• Pressurized liquid extraction is a sample preparation technique
that combines elevated temperature and pressure with liquid
solvents to achieve fast and efficient extraction of the components
from the solid matrix.
• PLE applies high pressure in extraction. High pressure keeps
solvents in a liquid state above their boiling point resulting in a
high solubility and high diffusion rate of lipid solutes in the
solvent, and a high penetration of the solvent in the matrix.

18. • PLE dramatically decreased the consumption of extraction time
and solvent and had better repeatability compared to other
methods.

19. 3. Ultrasonication-AssistedExtraction:
• The procedure involves the use of ultrasound waves, which have
frequencies higher than 20 kHz, have great effects on extraction yield and
kinetics.
• UAE involves ultrasonic effects of acoustic cavitations. Under ultrasonic
action solid and liquid particles are vibrated and accelerated and, because
ofthat solute quicklydiffusesout from solidphaseto solvent.

20. • Merits:
 Increaseofextractionyieldandfaster kinetics.
 Reducethe operatingtemperatureallowingthe extractionof thermolabile
compounds.
• Demerit :
 Theactiveconstituentsofmedicinalplantsthroughformationof free radicals
andconsequentlyundesirablechangesinthe drug molecules.

21. 4. Enzyme assisted extraction (EAE):
• The structure of the cell membrane and cell wall, micelles formed by
macromolecules such polysaccharides and protein, and the
coagulation and denaturation of proteins at high temperatures during
extraction are the main barriers to the extraction of natural products.
• The extraction efficiency will be enhanced by EAE due to the
hydrolytic action of the enzymes on the components of the cell wall
and membrane and the macromolecules inside the cell which
facilitate the release of the natural product.

22. 5. Pulsed electric field (PEF) extraction:
 PEF is the application of short time pulsed with high voltage into the
food product placed between two electrodes, thus promoting the
modification of membrane permeability and the increase of the
extraction yield. Recently, PEF treatment has been applied in order to
recover pigments from beetroot.
 Pulsed electric field extraction significantly increases the extraction
yield and decreased the extraction time because it can increase mass
transfer during extraction by destroying membrane structures.

23. 6. Microwave assisted extraction (MAE):
 Microwaves generate heat by interacting with polar compounds such
as water and some organic components in the plant matrix following
the ionic conduction and dipole rotation mechanisms.


24. • Different laboratories tend to adopt different procedures for initial
extraction of the source organisms.
• PREPARATION OF INITIAL EXTRACT OF A PLANT
• Terrestrial plants undergo extractions initially with a polar solvent
such as methanol, ethanol. Then, subject this extraction to a deflating
partition with a non-polar solvent such as chloroform or
dichloromethane and a polar aqueous solvent.

25. • Marine and Aquatic organisms are commonly extracted fresh into
methanol or a mixture of methanol and dichloromethane.

26. c) Preliminary biological screening:
• Biological screening is used to detect the presence of active
constituents in the initial extracts.
• Screening of natural products and their extracts is complicated due to
the presence of fluorescent or insoluble compounds.
• Advances in detection technologies and biological screening assays
have overcome these problems.

27. Methods of biological screening:
• 1. High through-put screening:
• In this process, large numbers of crude extracts from organisms can be
simultaneously evaluated in a cell- or non cell-based format, usually
utilizing multiwell micro titer plates.
• Using robotics, data processing/control software, liquid handling
devices, and sensitive detectors, high-throughput screening allows a
researcher to quickly conduct millions of chemical, genetic, or
pharmacological tests.

30. • 2. Cell-based in vitro bioassays
• Cell-based in vitro bioassays allow a considerable degree of biological
relevance, and manipulation may take place so that a selected cell
line may involve a genetically altered organism or incorporate a
reporter gene.
• Cell based bioassays are an important tool in the assessment of
potency, stability and safety of biopharmaceuticals, biosimilars, small
molecules, medical devices and cosmetic products.
• Such assays use spectrophotometric or turbidimetric method for
detection of activity.

31. 3. Noncellular (cell-free) assays
• In noncellular (cell-free) assays, natural product extracts and their
purified constituents are investigated for their effects on enzyme
activity or receptor binding.
• Subcellular fractions can be isolated by ultracentrifugation to provide
molecular machinery that can be used in reactions in the absence of
many of the other cellular components.

32. Techniques of biological screening:
• 1. Random screening
• 2. Virtual screening
• 3. Phytochemical screening

33. Random Screening:
• This approach strives to find, among a great number of
molecules(several hundreds or thousands), one that could be active
in a given indication.
• In this case therapeutic objective is fixed in advance.
• Great number of molecules is tested but on a limited number of
experimental models.

34. Major Discoveries by this process:
• Taxol
• Streptomycin
• Artemisinin
• Statins
• Morphine

35. Virtual Screening
• Virtual screening (VS) is a computational technique used in drug
discovery to search libraries of small molecules in order to identify
those structures which are most likely to bind to a drug target,
typically a protein receptor or enzyme.

37. Phytochemical screening:
• It refers to the extraction, screening and identification of the
medicinally active substances found in medicinal plants.
• The phytochemical research approach is considered effective in
discovering bioactive profile of plants of therapeutic importance.
• Phytocompounds such as Alkaloids, Tannins, Saponins, Phenols,
Flavonoids and Terpenoids are observed to be present in the
investigated plants in variable proportions.

38. d) Methods of compound purification:
• Purification is the physical or chemical process of removing contaminants
from a compound.
• The purification method mainly relies on chromatography, and the final
product is obtained through crystallization.
• The physical processes include:
• Sublimation
• Distillation
• Crystallization
• Chromatography

39. • Sublimation is the transition from the solid phase to the gas phase
without passing through an intermediate liquid phase.

40. • Distillation is the process of separating the components or
substances from a liquid mixture by using selective boiling and
condensation.

41. Fractional Distillation
• Fractional distillation is the separation of a mixture into its
component parts, or fractions.
• Chemical compounds are separated by heating them to a
temperature at which one or more fractions of the mixture will
vaporize.

42. • Crystallization is a purification technique to separate solids from a
solution.

43. • Chromatography is technique for separating the components, or
solutes, of a mixture on the basis of the relative amounts of each
solute distributed between a moving fluid stream, called the mobile
phase, and a stationary phase.

44. e. Structure elucidation and identification
• Structure elucidation in drug development is the process of
determining the chemical structure of a compound from the
identification and characterization of novel impurities, degradants
and metabolites across raw materials, API, formulated products or
biological matrices.
• Structure elucidation for organic compounds will often involve the
use of Nuclear Magnetic Resonance (NMR) spectroscopy.

45. • Other identification techniques include:
• Mass spectrometry
• Infrared spectroscopy
• X-ray crystallography

46. Dereplication:
• “Dereplication” is a process of determining whether an observed
biological effect of an extract or specimen is caused by a known
substance.
• OR
• Dereplication is the use of chemical techniques to eliminate extracts
that contain active constituents that have already been isolated and
characterized.

47. • This is applied during natural product drug discovery programs in an
attempt to avoid the reisolation of compounds of previously
determined structure.
• It is essentially chemical screening of the extracts using chemical
procedures such as chromatography, NMR, UV and visible
spectroscopy and mass spectrometry, and comparing the results to a
data base to identify active compounds that have already been
investigated.

48. Metabolomics
• “Metabolomics” is a recently developed approach in which the entire
or “global” profile of secondary metabolites in a system (cell, tissue,
or organism) is catalogued under a given set of conditions.

49. f. Compound development
• BACKGROUND:
• A major challenge in the overall natural product drug discovery
process is to obtain larger amounts of a biologically active compound
of interest for additional laboratory investigation and potential
preclinical development.

50. • STRATOGIES
• 1. Recollection of the species of origin:
• Recollection means to obtain the original source of compound for
further development.
• To maximize the likelihood that the recollected sample will contain
the bioactive compound of previous interest, the plant recollection
should be carried out in the same location as the initial collection, on
the same plant part , and during the same time of the year.

51. • 2. Collection of secondary metabolites:
• Secondary metabolites are organic compounds produced by bacteria,
fungi, or plants which are not directly involved in the normal growth,
development, or reproduction of the organism.
These metabolites can be used further for compound development.
• Other strategies include:
• 3. Cultivation
• 4. Mass Fermentation

52. • METHODS FOR COMPOUND DEVELOPMENT

53. Combinatorial biosynthesis
• Combinatorial biosynthesis can be defined as the application of genetic
engineering to modify biosynthetic pathways to natural products in order
to produce new and altered structures using
nature's biosynthetic machinery.
• This methodology involves the engineering of biosynthetic gene clusters in
microorganisms.
• Example: modification of bacterial polyketide synthases has led to
production of some 200 new polyketides that do not occur naturally.

54. Lead optimization
• Lead optimization aims at enhancing the most promising compounds
to improve effectiveness, diminish toxicity, or increase absorption.

55. Preliminary Structure-activity relationship (SAR) studies
• The structure–activity relationship (SAR) is the relationship between
the chemical structure of a compound and its biological activity.

56. •

57. e. Examples of natural product derived drugs
• PLANT SOURCE:
• Up to 50% the approved drugs during the last 30 years are from
either directly or indirectly from natural products.

58. Digitalis purpurea, the common foxglove
• Digitoxin, a trisaccharide derivative of digitoxigenin, is the only
compound to be used clinically to treat congestive heart failure and
cardiac arrhythmias.

59. Morphine, which is used as an analgesic, is isolated from the opium
poppy.

61. • Vincristine is an anticancer agent isolated from Vinca rosea.

62. ANIMAL SOURCE:

63. Insulin:
Insulin is a peptide hormone produced by beta cells of the pancreatic
islets; it is considered to be the main anabolic hormone of the body.
It helps control blood glucose levels by signaling the liver and muscle
and fat cells to take in glucose from the blood.

64. MICROBIAL SOURCE

65. Penicillin
• Penicillins are a group of antibiotics originally obtained from
Penicillium moulds, principally P. chrysogenum and P. rubens.
• It is often used to treat infections caused by Streptococcus and other
gram-positive bacteria.

66. Streptomycin
• Streptomycin is obtained from Streptomyces griseus.
• Streptomycin is an antibiotic medication used to treat a number of
bacterial infections.

67. MARINE SOURCE

68. Cephalosporin:
Cephalosporin C was isolated from the fungus Acremonium
chrysogenium (Cephalosporin acremonium).
• It was the lead for a wide range of active compounds, a number of
which are used as drugs.