Ika Putri Nurhayati

Jurusan Farmasi FKUB

Code: sm53ij

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 mampu menjelaskan definisi dan jalur metabolit primer dan sekunder

 mampu menjelaskan jenis senyawa metabolit primer dan metabolit sekunder

 mampu menjelaskan fungsi metabolit primer dan metabolit sekunder pada

tanaman
 mampu menjelaskan perbedaan senyawa metabolit primer dan sekunder

 mampu menjelaskan metode skrining metabolit sekunder menggunakan pereaksi

warna, dan KLT

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 A study of the chemical composition
of plants
Phytochemistry
Explanation of the various plant processes
in which chemical phenomena are
concerned

- Qualitative detection of plant component

- Actual isolation of plant component
- Quatitative estimation of plant component

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 Before the availability of synthetic drugs, phytodrugs or
herbal drugs were the mainstay of treatment
 An analysis into the sources of new drugs from 1981 to
2007 reveals that almost half of the drugs approved since
1994 were based on natural products. During the years
2005–2007, 13 natural product related drugs were
approved.
 Cancer and infections are the two predominant therapeutic
areas for which the drug discovery program is based on
natural products, but many other therapeutic areas also get
covered, such as neuro-pharmacological, cardiovascular,
gastrointestinal, inflammation, metabolic, etc
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PRIMARY METABOLITES
• Compound involved in primary metabolism, in which demonstrate the
fundamental unity of all living matter
• In plants, primary metabolism is made up of photosynthesis, respiration,
etc., using CO2, H2O, and NH3 as starting materials, and forming products
such as glucose, amino acids, nucleic acids. These are similar among
different species.
• Essentially the same in all organism
SECONDARY METABOLITES
• In secondary metabolism, the biosynthetic steps, substrates and
products are characteristic of families and species. Species which are
taxonomically close display greater similarities (and metabolites); those
which are distant have greater differences.
• Not necessarily produced under all conditions
• Ex. Flavonoid, alkaloid, terpenoid
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 Primary
• Sources of metabolic energy and
energy transfer
• Cellular building block and structural
support, ex: lignin
• Source of genetic information ex:
nucleus, ribosom, ER
• Catalyst of metabolic reaction
Secondary
• Deterrence of predators and
pathogens, ex: nicotine, hyociamine,
menthol, cyanide glycoside
• Attraction and deterrence of
pollinators
• Allelopathic action, ex: Arctostaphylos
uva-ursi (bearberry) that inhibit the
growth of grasses (Poaceae family)
and herbs
• Attraction of Symbionts, ex: host plant
and rhizobia nitrogen-fixing bacteria
(leguminoceae family
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 In green plants,
carbohydrates are
Size of Carbohydrates :
synthesized by the process
of photosynthesis utilizing Monosaccharides made up of one sugar
carbondioxide and water. unit (glucose or fructose)
 In most of the plants,
sucrose is the major form Disaccharides made up of two sugar units
of carbohydrates to be (sucrose is a glucose and a fructose)
translocated across the
plant body.
Photosynthetically fixed Polysaccharides are polymers made up of
carbon can also get stored more than two sugar units
in the form of starch.
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Polysaccharides

Structural polysaccharides are used to support plants

Storage polysaccharides are used to store energy for

later use by the plant

The most common structural polysaccharide in

plants is cellulose. It makes up 40 to 60% of the cell
wall. It is also the most common polymer on earth

Cellulose is extremely strong due to its chemical

organization. It is made of a long chain of beta-
glucose molecules – 100 to 15,000 glucose molecules
Storage Polysaccharides
The most important storage polysaccharides
are amylose and amylopectin.

Amylose is a long chain of alpha-glucose, several

hundred to several thousand molecules long.

Amylopectin is more complex, often made up of

50,000 molecules.

These two polymers are both used in making starch

grains. Most starch grains are about 20% amylose
and 80% amylopectin, but this varies with the plant.
 Plant use fats for mainly for carbon storage, but also for
energy production
 lipids, which make up both the plasma membrane and the
membranes of all internal compartments and organelles;
 Most lipids have a fatty acid portion made from acetyl-CoA
and malonyl-CoA in a reaction whose repetition produces
longer molecules.

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 OILS
Oils are fats that are liquid at room temperature.

Oils occur in all parts of a plant, but are most common in seeds. Some
seeds have so much oil that it can be commercially harvested. The most
commonly used oils are cotton, sesame, safflower, sunflower, olive, coconut,
peanut, corn, castor bean, and soybean oils.

The most common seed oil fatty acids are oleic acid (one double bond),
linoleic acid (two double bonds), and linolenic acid (three double bonds).

Linoleic and linolenic are essential fatty acids – we can’t make them
ourselves.
 WAXES
Waxes are complex mixtures of fatty acids linked to long-chain alcohols.

Waxes comprise the outermost layer of leaves, fruits, and herbaceous stems and are
called EPICUTICULAR waxes.

Waxes embedded in the cuticle of the plant are cuticular waxes.

Cutin is another wax in the cuticle and it makes up most of the cuticle.

Suberin is a similar wax that is found in cork cells in bark and in plant roots. Both
help prevent water loss by the plant.

Structures of waxes vary depending on which plant produced them.

Waxes are usually harder and more water repellant than other fats.
 proteins, which make up both
structural units of the cell such
as microtubles and all the enzymes
enzymes of every biochemical
process Enzymes catalyze biochemical reactions.
Most proteins in living cells are enzymes.
 nucleic acids and nucleotides,
which code for all proteins, act Pure enzymes that maintain their activity
as metabolic energy molecules when removed from plants are
commercially important to us.
such as ATP
 Ribosomes, for example, consist
of both protein and RNA, the
combination of which allows the
production of all other proteins. 16
 Secondary metabolites defend plant againts a variety of herbivores
and pathogenic microbes. SM may serve other imprtant functions as
well, such as structural support (ex lignin) or pigmen (ex anthocyanin)
 Secondary metabolites or secondary compounds are compounds
that are not required for normal growth and development, and are
not made through metabolic pathways common to all plants.
 Plant secondary metabolites can be divided into three chemically
distinc groups  terpenes, phenolics, and nitrogen containing
compound
 The building blocks for secondary metabolites are derived from
primary metabolism
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 Compound Plant
Chemical Class
Example Examples
Monoterpenes Mint
Essential oils and volatiles
(e.g. menthol)
Sesquiterpenes
Terpenoids
Largest group with Diterpenes
structures identified,
~30,000 compounds Triterpenes Cucumber

Tetraterpenoids

Tannins Condensed tannins Oaks

Phenolics
More ubiquitous in terms Lignins
of presence in plants
Flavonoids

Nitrogen Containing Alkaloids Nicotine, Cocaine Tobacco

Nitrogen and Sulfur Cabbage and other

Glucosinolates Sinigrin
Containing Brassicaceae
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 Plant produce a large variety of secondary products that
contain a phenol group, classified as phenolic compound.
 Plant phenolic compound are chemically heterogenous
groups of nearly 10.000 individual compound.
 Phenolics plays a variety roles in the plant, such as defense
compound, mechanical supporty, attracting pollinators and
fruit dispersers, absorbing harmful ultraviolet radiation,
reducing the growth of nearby competing plants.

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 Two basic pathways in phenolic biosynthesis are shikimic acid pathway and
the malonic acid pathway. The malonic pathway play important role in
phenolic production in fungi and bacteris, but less significance in higher
plant.
 The shikimic acid pathways converts simple carbohydrate precursors derived
from glycolysis and the pentose phosphate pathways to the aromatic amino
acid. This pathways present in plant, fungi, and bacteria, but is not found in
animals.
 Mostly, phenolic compound in plants are derived from phenylalanine via the
elimination of an ammonia molecule o form cinnamic acid.
 Trans-cinnamic acid, p-coumaric acid, and their derivatives are simple
phenolic compounds called phenylpropanoids because they contain a benzene
ring and three-carbon side chain. Phenylpropanoid are important building
blocks of the more complex phenolic compound.
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 Simple phenylpropanoid and benzoic acid, such as caffeic acid and ferulic acid
occur in soil in appreciable amounts and have been shown to inhibit the
germination and growth of many plants.
 Lignin, polymer of phenylpropanoid, is found in the cell walls of various types
of supporting and conducting tissues providing mechanical support and
protective function in plants.
 Flavonoid are one of the largest classes of plant phenolics. The basic carbon
skeleton of flavonoid contain 15 carbons arranged in two aromatic rings
connected by a three-carbon bridge. Isoflavonoids are group of flavonoids in
which the position of a one aromatic rings (ring B) is shifted.
 A second catergory of plant phenolic polymers beside lignin, is tannin. There
are two categories of tannin condensed and hydrolyzable.
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 The basic structural elements of terpenese are sometimes called isoprene
units because terpenes can decompose at high temperature to give
isoprene
 Terpenese are classified by the number of five carbon unit they contains.
Ten carbon terpenes which contain two C5 unit called monoterpenes, 15
C (three C5 units) are sesquiterpenes, 20C (four C5 unti) are diterpenes,
30 C are triterpenes
 Terpenes are biosynthesized from primary metabolites in at least two
different ways, mevalonic pathways and methylerythriol phosphate
(MEP) pathway

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 The red, orange, and yellow carotenoids are tetraterpenes that function as
accessory pigment in photosynthesis.
 In conifers such as pine and fir, monoterpenes accumulate in resin ducts
found in the needles, twigs, and trunk. These compound are toxic to
numerous insect.
 Many plants contain mixtures of volatile monoterpenes and sesquiterpenes,
called essential oils, that lend characteristic odor to their foliage.
 Mono and sesquiterpenes are commonly found in gandular hair on the plant
surface, especially the terpenes are stored in a modified extracellular space
in the cell wall.
 Essential oil have wellknown insect repellant properties.

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 Plant secondary metabolites that contain nitrogen in their
structure are alkaloid, cyanogenic glycosides, glucosinolates,
etc. Most nitrogenous secondary metabolites are
biosynthesized from common amino acids.
 Nitrogen atom in alkaloid is usually part of a heterocyclic
ring, that contain both nitrogen and carbon atoms.
 Most alkaloid are alkaline. At pH values commonly found in
the cytosol (pH 7,2) or the vacuole (pH 5 to 6), the nitrogen is
protonated, hence alkaloid are positively charged and are
generally water soluble.
 Most alkaloid are now believed to function as defense againts
predators because of their general toxicity and deterence
capability.
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 Cyanogenic glycosides and glucosinolates are not themselves toxic but are
readily broken down to give off volatile poisons when plant is crushed.
 Cyanogenic glycosides release the well-known poisonous gas hydrogen cyanide
(HCN).
 Cyanogenic glycosides are not normally broken down in the intact plant
because the glycoside and the degradative enzymes are spatially separated, in
different cellular compartements or in the different tissues.
 Cyanogenic glycosides have protective function in certain plants. This deters
feeding by insect and other herbivores, such as snails and slugs.
 Glucosinolates found principally in the Brassicaceae and related plant families.
These compound responsible for the smell and taste of vegetables such as
cabbage, broccoli, and radishes.
 Glucosinolates break down to release volatile defensive substances.
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 Alkaloid are usually synthesized from one of a few common
amino acids, in particular lysine, tyrosinen, and tryptophan.
 Carbon skeleton of some alkaloid contains a component derived
from the terpene pathways
 Nearly all alkaloid are also toxic to humans when taken in
sufficient quantity. For example strychnine, atropine, and coniin
are classic alkaloid poisoning agents. At lower doses, many are
useful pharmacologically.
 Morphine, codeine, and scopolamine are just a few of the plant
alkaloid currently used in medicine. Other alkaloid, including
cocaine, nicotine, and caffeine, enjoy widespread nonmedical use
as stimulant or sedatives.

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PHYTOCHEMICAL SCREENING
Phytochemical screening  The plant extracts analyzed for the
presence of secondary metabolites like alkaloids, terpenes, and
flavonoids.

Following this, a simple separation technique like thin-layer

chromatography (TLC) is generally used to analyze the number and
type of components present in the mixture.

The tests are simple to perform, however, it is not suitable for the
efficient separation of metabolites, and has low selectivity and
sensitivity of detection.

This procedure enables the early recognition of known metabolites

in the extracts, and is thus economically viable.

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PURPOSES OF
PHYTOCHEMICAL
SCREENING
Identify phytochemical group
of new explored plants

Finding chemical constituents

in the plant that may lead to
their quantitative estimation

Locating the source of

pharmacologically active
compound

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METHODS Simple

Should give additional Rapid

information as to the
presence or absence of
specific members of the
group being evaluated
A method for use
in phytochemical
screening should
be

Quantitative in so far as Designed for a

having a knowledge of the minimum of
lower limit of detection is equipment
concerned, and if possible
Reasonably
selective for the
class of compounds
under study

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DETECTION OF ALKALOID

Alkaloids usually occur in plants as their water-soluble salts

Most alkaloids are precipitated from neutral or slightly acid solution by

 Mayer's reagent (potassio mercuric iodide solution)  Cream precipitates
 Wagner's reagent(solution of iodine in potassium iodide)  Reddish-brown
precipitates
 Hager's reagent (a saturated solution of picric acid)  yellow precipitates
 Dragendorff's reagent (solutionof potassium bismuth iodide)  Reddish-brown
precipitates

Care must be taken in the application of these alkaloidal tests, as the reagents also give
precipitates with proteins
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ALKALOID SCREENING USING TLC

All Rauwolfia alkaloids give with

Dragendorff reagent orange-brown
zones

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DETECTION OF FLAVONOID
generally dissolved in
Flavonoid aglycone polar solvent such as
polyphenols are slightly in the presence of oxygen methanol, ethanol, acetone,
acidic and will thus many flavonoid will degrade dimethyl
dissolved in alkali. sulfoxamide,dimethly
furomide, water etc

while polar aglycones such

presence of sugar tends to
render the flavonoid more
H2O soluble
as isoflavone flavonone and
slightly methoxylated
flavone and flavonones
tends to be more soluble in
solvent such as ether and
propanol

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DETECTION OF FLAVONOID
Alkaline reagent test: The Extracts were treated with few drops of sodium
hydroxide solution. Formation of intense yellow colour, which becomes colour
less, on addition of few drop of dilute acid indicates the presence of flavanoids.

Lead acetate Test: The Extracts were treated with few drops of Lead acetate
solution. Formation of yellow colour precipitate indicates the presence of
flavanoids.

Ferric chloride test: 1 ml of the extract was treated with 1 ml of ferric

chloride.Formation of brown colour precipitates indicates the presence of
flavonoids
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1. Alkaline reagent test 2. Lead acetate Test 3. Ferric chloride test

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FLAVONOID SCREENING

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FLAVONOID SCREENING

apigenin fluorescent spots

when observed at 254 nm
after derivatisation with NP-
PEG reagent.

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DETECTION OF SAPONIN

Foam test: 0.5 gm of extract was shaken with

2 ml of water. It foam produced persists for
ten minutes it indicates the presence of
saponins.

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DETECTION OF TANNIN
Gelatin Test: To the extract, 1%
gelatine solution containing sodium
chloride was added. Formation of
white precipitate indicates the
presence of Tannins.
Lead acetate Test: To the extract
few drops of Aqueous basic lead
acetate solution were added, reddish
brown bulky precipitate indicates the
presence of tannin.
Modified Prussian blue test: To 1 ml
of extract, add 1 ml .008M potassium
ferric cyanide and 1 ml of .oo2 M
ferric chloride in 0.01 M HCl.
Presence of blue colour indicates the
presence of tannins.
Ferric chloride Test: To the extract,
few drops of 1%natural ferric chloride
solution was added formation of
blackish blue colour indicates the
presence of tannins
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DETECTION OF ANTRAQUINONE

Borutrager’s Test: To 1 ml of the extract , add 1 ml of 10 % ferric

chloride and .5 ml of concentrate hydrochloric acid. Boil in a water bath
for few minutes. Filter it and the filtrate is treated with 1 ml of Diethyl
ether and concentrate ammonia. Appearance of pink or deep red
colour.

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Rhei radix analysis
All aglycones show fluorescence quenching
in W-254nm and uniformly yellow or
orange-brown fluorescence in UV-365 nm.
T1 rhein
T3 emodin

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 DETECTION OF CARDIAC GLYCOSIDES
Keller-kiliani test
To the alcoholic extract of drug equal volume of water and 0.5 ml of strong
lead acetate solution was added, shaked and filtered. Filtrate was extracted
with equal volume of chloroform. Chloroform extract was evaporated to
dryness and residue was dissolved in 3 ml of glacial acetic acid followed by
addition of few drops of FeCl3 solution. The resultant solution was transferred
to a test tube containing 2 ml of conc. H2SO4. Reddish brown layer is formed,
which turns bluish green after standing due to presence of digitoxose.

Legal test
To the alcoholic extract of drug equal volume of water and 0.5 ml of strong lead
acetate solution was added, shaked and filtered. Filtrate was extracted with
equal volume of chloroform and the chloroform extract was evaporated to
dryness. The residue was dissolved in 2 ml of pyridine and sodium nitropruside
2 ml was added followed by addition of NaOH solution to make alkaline.
Formation of pink colour in presence of glycosides or aglycon moiety
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 DETECTION OF TERPENOID AND STEROID
• Alcoholic extract of drug was evaporated to dryness
and extracted with CHCl3, add few drops of acetic
anhydride followed by conc. H2SO4 from side wall of
Libermann burchard test test tube to the CHCl3 extract. Formation of violet to
blue coloured ring at the junction of two liquid, indicate
the presence of steroid moiety

• Alcoholic extract of drug was evaporated to dryness

and extracted with CHCl3, add conc. H2SO4 from
sidewall of test tube to the CHCl3 extract. Formation of
Salkowaski test yellow coloured ring at the junction of two liquid, which
turns red after 2 min, indicate the presence of steroid
moiety.

• Alcoholic extract of drug was evaporated to dryness

and extracted with CHCl3, add saturated solution of
Antimony trichloride test SbCl3 in CHCl3 containing 20% acetic anhydride.
Formation of pink colour on heating indicates presence 51
of steroids and triterpenoids.
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 • is a separation technique based
on the different interactions of
compounds with two phases, a
Chromatography mobile phase and a stationary
phase, as the compounds travel
through a supporting medium.

mobile phase
• a solvent that flows through the supporting medium

stationary phase
• a layer or coating on the supporting medium that
interacts with the analytes
supporting medium
• a solid surface on which the stationary phase is
bound or coated
Principles of
Chromatography
Chromatogr
aphy is a
physical
process.

We can only
control
stationary and
mobile phase
and the mixtures
are the problem
we have to deal
with.

Chromatography
is a dynamic
process in which
the mobile phase
moves in definite
direction.
 What are the OBJECTIVES of
chromatography ?

To separate the components of

mixture
(preparative chromatography)

To identify the component of an

unknown
(analytical chromatography)
How does chromatography WORKS?

works by allowing the molecules

present in the mixture to distribute
themselves between a stationary and
a mobile phase.

molecules that spend most of their

time in the mobile phase are carried
along faster
Classification of Chromatographic
methods
According to mechanism of separation:
depends mainly on the nature of the stationary phase.
Chromatography can be classified into:

- Adsorpsion chromatography
- Partition chromatography
- Ion exchange chromatography
- Molecular/Size exclusion chromatography
- Zone electrophoresis
- Affinity chromatography
- Chiral chromatography
 Adsorption Chromatography

Separation is based on the interaction of the

molecule in a mobile phase with the surface of the
solid stationary phase (a liquid-solid interaction)
This is a distribution process
where the molecule interacts
reversibly
(binds and unbinds) with the
surface molecules of the
stationary phase
(usually a finely divided solid)
 Adsorption Chromatography

The molecule and the mobile phase compete for

adsorptive sites on the stationary phase.
1) Molecules (in the sample) that are most soluble in the
mobile phase, will pass through the system quickly;
2) Molecules (in the sample) that are most soluble in the
stationary phase will elute off the column slowly.

Generally, the adsorbant is applied

or bonded to a solid support (glass
plate, plastic sheet)
1) Polar adsorbants : silica, alumina
2) Non-polar adsorbants : charcoals,
paraffin
 Partition Chromatography

Separation is based on the interaction of the molecule in

a mobile phase with the liquid stationary phase (a liquid-
liquid or gas-liquid interaction)

Separation of solutes is based on their

differences in solubility between two
immiscible phases (solvents).
1) The solubility of the molecules is
based upon solute polarity and the
concept of “like dissolves like”
2) The molecules (solute) of interest
are ‘extracted’ into a solution based
on relative solubilities
 Partition Chromatography

normal phase

reverse phase
 Ion exchange Chromatography

Sample ions compete with the mobile phase ions for ionic
sites on the stationary phase (charged functional groups)

The sample ions that interact

weakly with the stationary phase
will be retained the least, whereas,
those that interact strongly with the
stationary phase will be retained
the longest and will elute off the
column later.

Cation-exchange and anion-

exchange methods are available
depending on the molecules to be
separated
Ion exchange Chromatography
 Molecular/Size exclusion
Chromatography

Separation is based on the size of the molecule

Small molecules will elute off the column slowly
Large molecules will elute off the column
quickly
Zone Electrophoresis Chromatography

Separation based on size

and charge
Smaller molecules will
migrate further, less tangled
Zone Electrophoresis Chromatography

Sample application

Bands of postivey Bands of negativey

charged compounds charged compounds
 Affinity Chromatography

Very selective
Specific binding site is used to concentrate analyte on column
Used a lot in biological applications

It uses the affinity of

proteins to specific ligands
such as enzymes.
The ligand is attached to
suitable polysaccharide
polymer such as cellulose
- agarose – dextran.
Affinity Chromatography
 Chiral Chromatography

In this type we can separate enantiomers – we used

chiral stationary phase that react with one
enantiomer more then the other so separation takes
place.
 CONCEPT OF POLARITY

The interaction of attractive forces (molecular

interactions) that exist between molecules is
the basis of polarity

Polarity is a relative term applied to solvents,

solutes and adsorbents.

The degree of polarity is dependent upon the

compound’s molecular geometry, electron
configuration and its hydrogen bonding
 CONCEPT OF POLARITY

Organic solvents are generally non-polar;

molecules are rigid and symmetrical;
termed hydrophobic because they are not
readily soluble in water (hexane, chloroform)

H H H H H
H – C – C – C – C – C – H (non-polar organic)
H H H H H
 CONCEPT OF POLARITY
Alcohols and acids are generally polar compounds;
molecules tend to be less symmetric and not as rigid;
more soluble in water as compared to the organic
solvents; termed hydrophilic because they are readily
soluble in water (acetonitrile, isopropanol, HCl, H)

H H H H H
H – C – C – C – C – C – OH (polar alcohol)
H H H H H

H H O
H – C – C – C (more polar acid)
H H OH
 According to mobile phase:
Chromatography can be classified into:
1- Liquid Chromatography (LC):
The mobile phase is liquid.
-Liquid-Solid Chromatography (LSC)
Separation by adsorption, the stationary phase is solid
-Liquid -Liquid Chromatography (LLC)
Separation occurs through partition, the stationary phase is
liquid

2- Gas Chromatography (GC)

The mobile phase is inert gas nitrogen or helium.
-Gas–Solid Chromatography (GSC)
the stationary phase is solid
-Gas-Liquid Chromatography (GLC)
the stationary phase is liquid
According to technique
(methods of holding the stationary phase):
Chromatography can be classified into:

• the stationary phase

Column
chromatography is contained in a tube
called the column

• stationary phase is
Planar
chromatography configured as a thin
two-dimensional sheet
PLANAR CHROMATOGRAPHY

In paper chromatography a sheet or a narrow

strip of paper serves as the stationary phase

In thin-layer chromatography a thin film of a

stationary phase of solid particles bound
together for mechanical strength with a binder,
such as calcium sulfate, is coated on a glass plate
or plastic or metal sheet.
PLANAR CHROMATOGRAPHY
 COLUMN CHROMATOGRAPHY

The primary division of chromatographic

techniques is based on the type of mobile phase
used in the system

Type of Type of Mobile

Chromatography Phase
Gas gas
chromatography
(GC)
Liquid liquid
chromatograph (LC)
According to purposeof use
Chromatography can be classified into:
1- Analytical Chromatography:

a. Qualitative Chromatography
b. Quantitative Chromatography

2- Preparative Chromatography
Qualitative Chromatography
1- Confirm the absence or probable presence of certain
constituent in the sample under investigation

2- Give an idea about the complexity of the mixture and the

least number of compounds present.

3- Check purity and identity of any compound.

4- Establish a (finger print ) pattern for extracts, volatile oils

or pharmaceutical preparations. These finger prints can be
then used to check the identity and purity in the future.

5- Monitor both column chromatography and organic chemical

reactions.
Quantities Chromatography:

The development of modern instruments

enable the use of chromatography to
determine the amount of any component
in a mixture as absolute amount or
relative to another component HPLC/ GC/
HPTLC can be used for there applications.
Preparatives Chromatography:

This was the first and is the main

application of chromatography. The
technique was developed primarily for
this purpose.
Chromatography is used to obtain
reasonable quantities of pure compounds
from mixtures.
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