FAU 1302 Kimia Medisinal Organik

Drug structure and solubility, Drugs from natural sources,

Biological membranes

Ari Satia Nugraha PhD., Apt

Pharmaceutical Chemistry Department
University of Jember
 Mankind has explored natural resources
(abiotic and biotic) since early times and
has used them for survival and to develop
their civilization.

Plants are one of the biotic components in the biosphere which have been used
for food, clothing, ritual, medicine, dye, construction, cosmetics and others.

Extracts : medicine, dye, cosmetics

Natural products
Can be classified into three components, namely primary metabolites,
cellular structures and secondary metabolites.

 Natural products (sec. met) have a higher value in the market than
primary metabolites. Over 100,000 secondary metabolites have been
extracted from plants.

Secondary metabolites can be structurally divided into alkaloids,

phenylpropanoids, polyketides, and terpenoids.
Basic biosynthetic pathways CO2 H2O

Polysaccharides O2 O2
Glycoside Monosaccharides
O
HO
OH
O
OH HO
OH
O
shikimic acid 2
pyruvic acid 1

peptides

HO O O HO O
O
aliphatic
HO OH OH
OH amino acids
mevalonic acid 3 acetic acid 4 HO O
alkaloids Prephenic acid 5
O O
O O
P P
O - O - O-
O O HO OH
3,3-dimethylallyl pyrophosphate 6 malonic acid 7 aromatic amino acids

terpenoids polyketides cinnamic acids

fatty acids flavonoids, other aromatics coumarins

fat
Natural products: Isolation and
characterisation
 The oldest attempt on natural product purification is distillation and
extraction process by decoction

In general, isolation starts from sample preparation (pulverization),

extractions-fractionation, and separation.

Natural product separation was commonly conducted using preparative

chromatographic methods

Molecular structure characterisation, spectroscopic and spectrometric

(MS, NMR, UV, IR)
Natural products: Research and exploration
The study of plants is not new with evidence of studies since ancient times,
including the interaction between human-plants, plants-plants, and abiotic
environment-plants
Human-plants:

Food  Phoenix spp (palm dates) was

domesticated back in the Late Middle
Paleolithic (300,000 to 30,000 years ago).

Medicine  Achilles santolina has been used as

anti-dysentery since 60,000 years ago.
1981-2002: 868 new compound entities revealed from different sources

2008-2013: 100 natural products and natural product derived new entities
were in clinical trials which were investigated for potential for oncology
treatments (38%), anti-infective disease (26%), cardio-vascular and
metabolic disease (19%), inflammatory and related disease (11%), and
neurogical treatments (6%)
Plant-plant:
Allelophatic plantsRelease allelopathic chemicals into the immediate
environment which have an influence upon the growth and development of the
surrounding agricultural and biological ecosystem.

Weed management, Oryza sativa L (rice crop)

release momilactone A and B, effective against the
most harmful weeds in rice fields (banyrad grass:
Echinochloa crus-galli and Echinochloa colonum)
with concentrations of > 1 and > 10 µM, respectively.

H H
O HO
H H H H
O O
momilactone A 34 momilactone B 35
Abiotic environment-plants:
These abiotic stresses included drought, temperature, UV-radiation and CO 2
level. Several plants produced flavonoids (i.e anthocyanins) as a protective
barrier against drought and UV-radiation. A range of cryoprotective
compounds such as sugar alcohols were synthesized as a reaction to cold
during winter.
A study on plants grown in higher levels of CO2 revealed changes in the
plant’s chemical composition whereby the synthesis of phenolics is increased
and there is condensation of tannins in leaves.
The impact of increasing the level of CO2 consistently increased the level of
phenolic compounds.
Drugs from natural resources
A. Scheme
1. Introduction
Sources:
• Plant
• Animal
• Marine
Common sources, plant and microorganism both land and marine
The selection may be based on ethnopharmacology or current interest
Example:
South American Indians chewed coca leaves to alleviate fatigue/tiredness 
cocaine.
2. Investigation
• General purpose  general compounds
• Specific purpose  bioactive compound targets for specific diseases
3. Bioactive isolated compounds
• Molecular structure determination
• Commercial interest  synthesis and analogues synthesis  QSAR

Synthesis vs natural production

Economic efficiency  cost of production comparison
Example:
Vincristine, vinblastine, etoposide, taxol

Catharanthus roseus
• Vincristine
• Firstly isolated in 1961
• Chemotherapeutic agent
• WHO essential medicine
Catharanthus roseus
• Vinblastine
• Firstly isolated in 1958
• Chemotherapeutic agent
• WHO essential medicine

Podophyllum peltatum
• Etoposide
• Chemotherapeutic agents
4. Bioassay
• Screening  to detect the presence of active compound
• Monitoring  to follow the path
Collection and screening
• Enviroment
• Time of collection
• Preparation and storage
Screening test
• Rapid, accurate and reproducible, high capacity, cheap
• Tarry and poorly water soluble extracts
• However, there is still possibile to miss active extracts
• Broad or specific screening test (cytotoxic, anti-microbial, etc)

Methods:
• Whole organism screening test
i.e. Brine shrimp lethality test (BSLT), crown gall tumour inhibition
• Cultured cell tests
• Isolated enzyme tests
• Isolated tissue tests
 λ 400 nm

Example, isolated enzyme tests  trypsin inhibition

Isolated tissue test, example
Monitoring tests
• Concentration of active compounds increases during fractionation
• Activities might be fall/disappear during fractionation
• Activity changes due to decomposition (nature of process or inherent
compound stability  oxidation, hydrolysis, polymerisation).
The removal of naturally occurring antioxidants, increases the enzyme
ctalyse the decomposition, light exposure
Vigorous fractionation condition can be reduced by, low temperature,
solvent acidity-alkalinity as near neutral. Solvent removed by either
freeze drying or distillation under low vacuum.
• Activity changes due to synergy

Synergy????? Not understood yet.

Thalidomide is more
effetcive in myloma
treatment in the presence
of dexamethasone

• Activity change due to fractionation loss

• Activity change due to fractionation loss
Locked up in the procedures. i.e Chromatographic technique: active
compounds strongly or permanently absorbed in a solid stationary
phase. Distillation: co-sublimation, co-distillation  leaving inactive
concentrate
5. Dereplication

• Technique to eliminate extracts that contain active constituents that have

already been isolated and characterised. Wasting time and resources
• Compared to database
• Case: NCI 40,000 plants were screened from 1987-1992. 15% active extracts
anti-HIV
• Example of dereplication procedure in anti-HIV drug discovery from plants
Known active polysacharide

Polyphenols/tannins were retained  known

anti-HIV active
6. Molecular structure analysis of isolated compounds
Must be pure
Small isolates  hard to get crystalline
Pure based on HPLC/GC
7. Active compound development
Naturally occurring active compounds into commercially viable drugs
B. Extraction procedures
• Separate desired chemicals from unwanted cellular debris and chemicals i.e
lipids, proteins, polysaccharides.
• The design: scale, nature of screening assays, chemical nature of the
compounds in the extracts
• Polarity
• Acidity
1. Commonly used methods of extractions
• Solvent-based methods
• Infusion
• Steam distillation
• Supercritical fluid extraction
• Taxol were extracted using CO2 SFE from Catharanthus roseus
2. Cleaning up procedures
• Constituents  fractionation difficulties, interfering bioassay.
• i.e. Tannisn, carotenes, chlorophyll  bioassay problem. Tannins 
cross link with protein, bioassay problem.
• Inorganic salts, i.e NaCl hard to be removed
• Removal good and bad depends on the activity
3. Fractionation methods
• Liquid-liquid partition

Acidic and basic compounds fractionations

• Mechanical methods
Automated liquid-liquid partition
Craig counter-current distribution

• Slating out
Introducing very soluble iorganic salts, i.e NaCl, NH4Cl into aqueous
fraction. This will reduce solubility of non-ionic compounds in water
and enhance their solubility inany less polar immiscible slovent.
4. Chromatographic methods
5. Precipitation
6. Distillation
7. Dialysis
8. Electrophoresis
Case story of taxol

Taxus brevifolia (Pacific yew)

• Taxol
• 1955  NCI screenings (synthesis and natural
products) Jonathan E Well
• 1958  plant screenings (1000 species a year)
• 1962  Pacific yew collection
• 1964  extract, active
• 1968  1200 kg bark  28 kg crude extract
extract  10 g pure taxols
• 1975  anti-cancer in vitro (3,000 kg bark)
• 1978  in vivo active (leukemic mice)
• 1980  clinical (9,000 kg bark)
• 1984-1986  clinical trial (27,000 kg bark)
• 1990   Bristol-Myers Squibb (BMS), taxol
• 2000  $1.2 billion peak wholesale
• 1962  650 extracts investigated in Res. Triangle institute (Dr. M.E. Wall under
NCI contract
• Extracts were positive against 9KB cell in vitro and L-1210 mouse in vivo
• Taxus brevifolia  12 Kg was 95 percent methanol extracted
• Biological assay against the Walker-256 solid tumour (5WM)

• Partition (water and Chloroform methanol (4:1))

• Organic layer  146 g solids showed good activity against 5WM
• Solid applied into CCCD
• Yield 0.004 percent
• Low yield, solubility problem, limited bark supply
• Three mature pacific yew tree to produce 1 g taxol
• Asymmetric centres  difficult to synthesis
• Insoluble in water  administration problem as most chemitherapeutic
use intravenous
• Problem counteration
• Polyethoxylated caster oil and absolute alcohol  solublise in 5%
dextrose or saline
• 1977  preclinical started
• 1979  mechanism of action (cell division)
• 1982 phase I trials
• 1985  phase II trials

• 1992  FDA approved  32 years after the discovery

• Currently produced using semisynthetic pathway from baccatin III and 10-
deacetylbaccatin III (isolated from Leaves other Taxus). No need for tress
destruction.