as per GTU sem -7 students biosynthetic pathways - shikimik acid pathway

1. Prepared by
Pooja H. Khanpara
Asst. Professor
Apip, jamnagar
Biosynthetic Studies &
Basic Metabolic Pathways

2. What is Biosynthesis?
Biosynthesis is a process of forming larger organic
compounds from small subunits within a living organism.
Biosynthesis is mainly done by enzymes.
Biosynthesis is also known as anabolism since simple
compounds are joined together to form macromolecules by
enzymes.
As an example, photosynthesis occurs inside the chloroplast.
The light energy is converted into chemical energy during
photosynthesis.
 The larger molecule glucose is biosynthesized from water and
carbon dioxide by photosynthetic organisms.(ATP, Enzyme,
Cofactors)

4. What is the Difference Between Synthesis
and Biosynthesis?
Synthesis vs. Biosynthesis
Synthesis refers to the formation of
macromolecules from small
molecules artificially.
Biosynthesis refers to the formation
of larger organic compounds from
small molecules within a living
organism.
Process
Synthesis is artificial and chemical. Biosynthesis is biological and
catalyzed by enzymes.
Resulting Polymers
Synthesis can result in polymers
which are organic or non-organic.
Biosynthesis is biological and
catalyzed by enzymes.
Occurrence
Synthesis occurs outside living
organisms.
Biosynthesis occurs within a living
organism.

5. Biosynthesis of Primary Metabolites
 Living plants are solar-powered biochemical and biosynthetic
laboratory which manufactures both primary and secondary
metabolites from air, water, minerals and sunlight.
 The primary metabolites like sugars, amino acids & fatty acids that
are needed for general growth & physiological development of
plant which distributed in nature & also utilized as food by man.
 The secondary metabolites such as alkaloids, glycosides,
Flavonoids, volatile oils etc are biosynthetically derived from
primary metabolites.
 Biosynthetic reactions are replica of common organic reactions like
catalytic reactions, phosphorylation, hydride transfer, oxidation,
elimination, acylation, alkylation, reduction, condensation,
rearrangement etc.

6. Metabolism & Metabolic Pathways
Cell Metabolism: Process by which living cell process
nutrient molecule & living state.
Metabolic Pathway: A complete set of chemical
reactions that occur in living cells, allowing cells to grow
and reproduce, maintain their structures, and respond to
their environments.
Living cell require energy for biosynthesis, transport of
nutrient, motility and maintenance.
Energy is obtained from the catabolism of carbon
compounds (carbohydrate)
Carbohydrates are synthesized from CO2 and H2O in the
present of light by photosynthesis.

7. ~ produce energy to the cell
~ requires energy
glucose to glycogen

8. Major Metabolic Pathways
Cellular respiration:
Glycolysis
Anaerobic respiration
Kreb’s cycle / Citric acid cycle
Oxidative phosphorylation
Creation of energetic compounds from non-living matter:
Photosynthesis (plants, algae cynobacteria)
Chemosynthesis (some bacteria)
Other pathways occurring in (most or) all living cell:
Fatty acid oxidation (β-oxidation)
Gluconeogenesis
HMG-CoA reductase pathway (isoprene prenylation)
Pentose phosphate pathway (hexose monophosphate)
Porphyrin synthesis (or heme synthesis) pathway
Urea cycle

9. Metabolites
Metabolites are the intermediates & products of
metabolism.
The term metabolite is usually restricted to small
molecules.
A primary metabolite is directly involved in the
normal growth, development, and reproduction.
A secondary metabolite is not directly involved
in those processes, but usually has important
ecological function.

10. Importance of photosynthesis in
formation of primary metabolites
Photosynthesis is the process where plants convert sunlight into
energy, then store it as carbohydrates, sugars, such as glucose.
Photosynthesis may be the most important process in ecosystems,
both brings in energy needed within the ecosystem, and
produce oxygen (O2) needed for cellular respiration, and the
production of more ATP.
Photosynthesis has three basic steps:
1. Energy is captured from the sunlight.
2. Light energy is converted into chemical energy in the form of ATP
and NADPH.
3. Chemical energy is used to power the synthesis of organic
molecules (e.g. carbohydrates) from carbon dioxide (CO2).

11. Photosynthesis
H2O + light + ADP + P ---> O2 + ATP + e-
After the above steps occur in
photosystem II, the electron is finally sent
to photosystem I, where the following
happens.
e- + NADP+
+ H ---> NADPH
Now there are two high energy molecules,
fully charged and ready to be used. Plants
make more energy that it needs
immediately, so the NADPH and ATP are
used to make glucose as follows:
CO2 + ATP + NADPH ---> C6H12O6
This happens in Calvin cycle.

12. Calvin Cycle
The Calvin cycle is the last step in photosynthesis.
The purpose of the Calvin Cycle is to take the energy from
photosystem I and fix carbon. Carbon fixation means building organic
molecules by adding carbon onto a chain. The following formula
summarizes the Calvin cycle.
C5 + CO2 + ATP + NADPH → C6H12O6
Where C5 is a five carbon molecule, such as pyruvate, when is
recycled as glucose is synthesized.
The first step in the Calvin cycle is for the 3C5 to bind with 3CO2,
producing a six 3-carbon organic molecules (6C3).
Next, 6ATP and 6NADPH energizes the binding of a C3 to make a 6-
carbon molecule (C6), glucose.
The remaining 5C3 continues moving through the Calvin cycle,
being turned back into the starter C5 organic molecule.

13. Calvin Cycle

14. Glycolysis
(Embden-meyerhoff pathway)
Glycolysis represents an anabolic pathway common in both
aerobic and anaerobic organisms.
Sugars and polysaccharides are transformed into glucose or
one of its phosphorylated derivatives before being processed
any further. In the course of degradation, ATP is produced.
Pyruvate may be regarded as the preliminary final product of
the degradation. Pyruvate is fed into the citric acid cycle
via an intermediate product.
This pathway produces energy in the form of ATP. The
starting product glucose is completely oxydized to water and
carbon dioxide.

16. Citric Acid Cycle (Kreb’s cycle)
The citric acid cycle, is the common mode of oxidative
degradation in eukaryotes and prokaryotes.
It accounts for the major portion of carbohydrate, fatty acid and
amino acid oxidation and produces at the same time a number of
biosynthetic precursors.
The GTP generated during the succinate thiokinase (succinyl-
CoA synthetase) reaction is equivalent to a mole of ATP by virtue
of the presence of nucleoside diphosphokinase.
The 3 moles of NADH and 1 mole of FADH2 generated during
each round of the cycle feed into the oxidative
phosphorylation pathway.
Each mole of NADH leads to 3 moles of ATP and each
mole of FADH2 leads to 2 moles of ATP. Therefore, for each mole
of pyruvate which enters the TCA cycle, 12 moles of ATP can be
generated.

18. Carbohydrate Utilization
Storage carbohydrate such as the starch of plants or the
glycogen of animals is made available for energy production
by a process.
As a result of this, the energy-rich carbohydrate is eventually
oxidized to CO2and H2O.
During the process, the hydrogen atoms liberated are carried by
coenzymes into the cytochrome system, in which energy is
released in stages, with the possible formation of ATP and ADP
and inorganic phosphate.
Eventually the hydrogen combines with oxygen to form water.
The overall reaction of glucose in terms of ADP and ATP is
C6H12O6 + 6CO2 + 38 ADP + 38P (inorganic) → 6H2O + 6CO2 + 38
ATP

19. The primary and secondary metabolites
derived from carbon metabolism

21. difference between Primary and
secondary metabolites

22. Biosynthetic Pathway of Secondary
Metabolites
It involves 3 basic mechanism:
1. Shikimic acid Pathway
2. Acetate-mevalonate pathway
3. Acetate malonate pathway

23. Shikimic acid
Commonly known as its anionic form shikimate, is a
cyclohexene, a cyclitol and a cyclohexanecarboxylic
acid.
Its name comes from the Japanese flower shikimi the Japanese star
anise, Illiciumanisatum), from which it was first isolated in 1885 by
Johan Fredrik Eykman.
The elucidation of its structure was made nearly 50 years later.
Shikimic acid is also the glycoside part of some hydrolysable
tannins.

24. Shikimic Acid Pathway
The Shikimic acid pathway is a key intermediate from carbohydrate for
the biosynthesis of C6-C3units (phenyl propane derivative).
The Shikimic acid pathway converts simple carbohydrate precursors
derived from glycolysis and the pentose phosphate pathway to the
aromatic amino acids.
The shikimate pathway is a 7 step metabolic route used by bacteria,
fungi, Algae, parasites, and plants for the biosynthesis of aromatic amino acids
(phenylalanine, tyrosine, and tryptophan).
This pathway is not found in animals; therefore, phenylalanine and
tryptophan represent essential amino acids that must be obtained from the
animal's diet.
 Animals can synthesize tyrosine from phenylalanine, and therefore is not
an essential amino acid except for individuals unable to hydroxylate
phenylalanine to tyrosine).

26. Phosphoenolpyruvate and erythrose-4-phosphate react to form 2-keto3-
deoxy7phosphoglucoheptonic acid, in a reaction catalyzed by the enzyme
DAHP synthase.
2-keto3-deoxy7phosphoglucoheptonic acid is then transformed to 3-
dehydroquinate (DHQ), in a reaction catalyzed by DHQ synthase.
Although this reaction requires nicotinamide adenine dinucleotide
(NAD) as a cofactor, the enzymic mechanism regenerates it,
resulting in the net use of no NAD.

27. DHQ is dehydrated to 3-dehydroshikimic acid by the enzyme 3-dehydroquinate
dehydratase, which is reduced to shikimic acid by the enzyme shikimate dehydrogenase,
which uses nicotinamide adenine dinucleotide phosphate (NADPH) as a cofactor.
The next enzyme involved is shikimate kinase, an enzyme that catalyzes the
ATPdependent phosphorylation of shikimate to form shikimate 3-phosphate.
Shikimate 3-phosphate is then coupled with phosphoenol pyruvate to give 5-
enolpyruvylshikimate-3-phosphate via the enzyme 5-enolpyruvylshikimate-3-
phosphate (EPSP) synthase.
Then 5-enolpyruvylshikimate-3-phosphate is transformed into chorismate by
a chorismate synthase.

28. Prephenic acid is then synthesized by a Claisen rearrangement of chorismate
by Chorismate mutase.
Prephenate is oxidatively
decarboxylated with retention of
the hydroxyl group by
Prephenate dehydrogenase to
give phydroxyphenylpyruvate,
which is transaminated using
glutamate as the nitrogen
source to give tyrosine and α-
ketoglutarate.

29. Role of Shikimic Acid
Pathway:
Starting Point in The Biosynthesis of Some Phenolics Phenyl alanine and
tyrosine are the precursors used in the biosynthesis of phenylpropanoids.
The phenylpropanoids are then used to produce the flavonoids,
coumarins, tannins and lignin.
Gallic acid biosynthesis Gallic acid is formed from 3-dehydroshikimate by
the action of the enzyme shikimate dehydrogenase to produce 3,5-
didehydroshikimate.
The latter compound spontaneously rearranges to gallic acid.
Other compounds
Shikimic acid is a precursor for:
Indole, indole derivatives and aromatic amino acid tryptophan and
tryptophan derivatives such as the psychedelic compound
dimethyltryptamine. & many alkaloids and other aromatic metabolites.

32. Uses:
In the pharmaceutical industry, shikimic acid from the Chinese star
anise (Illicium verum) is used as a base material for production of
oseltamivir (Tamiflu).
Target for drugs
Shikimate can be used to synthesize (6S)-6-Fluoroshikimic acid, an
antibiotic which inhibits the aromatic biosynthetic pathway.
Glyphosphate, the active ingredient in the herbicide
Roundup, kills plants by interfering with the shikimate
pathway in plants. More specifically, glyphosate inhibits the
enzyme 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS).
"Roundup Ready" genetically modified crops overcome that
inhibition.

33. Atropine

36. 2nd
method
Take Powder drug, ext with 95% alcohol(soxhlet hot percolation)
Ethanolic ext. concentrate it
Add dil. HCl & filter it
extracted with pet. Ether (to remove impurities)
Aq. Sol. Make alkaline with NH3
Extracted with CHCl3 (3 times)
Combine exts. Evaporate chloroform under vaccum
Again ext. with dil. Oxalic Acid
Finally get crystal of Atropine & Hyoscyamine

47. 1. Thalleioquin Test: sol. of chinchona drug when treated with
Bromin & ammonia it produce emerald green color.
2.

53. Ext. with chloroform
further purification make acidic
(Codein)
make alkaline (NH3)
ppt(Morphine) sol. (Narceine)

73. Isolation of Reserpine
Roots are powdered & moisten with 10 % NaHCo3 & ext. with
benzene untill give positive reaction with HgI2
Conc. It & add ether & dil. HCl again conc. It & separate acid layer.
Again washed with ether. Make it alkaline with NH3
Then ext. with CHCl3
The CHCl3 ext. washed with 10% Na2CO3
Dry it & purify it by using methanol
Get pure crystal of Reserpine

77. Isolation of Ephedrine
Powder drug , moist with Na2CO3 & ext. with Benzene
Filter it
Take residue
ext. With Dil. HCl
Make alkaline by using K2CO3 & add CHCl3
Add Na2SO4 in CHCl3 solution & dried it
Take residue, treat with Oxalic Acid ,warm, filter & cool it
Get Ephedrine oxalate crystal

79. Chemical Test:
Ephedrine in H2O + dil. HCl treated with  CuSO4 + NaOH 
violate color  add ether  purple color  Aq. Layer
shows blue color

84. Podophyllum
Indian Podophyllum
Synonym – Indian May apple, Wild lemon, Duck’s Foot, Hog
Apple
Biological source –
It consists of the dried rhizome and root of Podophyllum hexandrum
or Podophyllum emodi
Family – Berberidaceae
Use: purgative, tmt of cancer (Ovarian cancer), antirhematic,
insecticidal activity