Reserpine(Structure Elucidation, Extraction and Isolation)

Reserpine
(Structural elucidation, Extraction & Isolation)
Prepared By:
MOHAMMAD KHALID
(Assistant Professor)
Krishna Pharmacy College, Bijnor (UP)

content
Introduction
Constitution of reserpine
Structure of Reserpic acid
Structure of Yobyrine
Synthesis of Yobyrine
Structure of Reserpine
Synthesis of Reserpine
Classification
Extraction
Isolation:
Identification test
Mode of Action
16-Apr-20 Krishna Pharmacy College, Bijnor 2

introduction
Reserpine is the main constituent of Rauwolfia
species,perticularly Rauwolfia serpentina &
Rauwolfia vomitoria.
It is mainly used for the treatment of
hypertension, headache, tension, asthma &
dermatological disorders.

Constitution of reserpine
1. Molecular formula:- C33H40N2O9
2. Presence of five methoxy groups:-
By zeisel method i.e. when treated with HI yields five
molecules of methyl iodide indicating the presence of five
methoxy groups.
3. Nature of ‘N’ atom:-
a) Secondary ‘N’ – Formation of monoacetyl derivative with
acetic anhydride indicates secondary ‘N’ .
b) Tertiary ‘N’- Reserpine forms quaternary ammonium salts
with CH3I that indicating one of ‘N’ is tertiary.

4. Hydrolysis:- Hydrolysis of reserpine yields mixture of
i) Methyl alcohol
ii) 3,4,5-trimethoxy benzoic acid
iii) Acid (A) of composition
C33
H40
N2
O9 H2 O
NaOH CH3
OH C22
H28
N2
O5
COOH
OMe
OMe
MeO
+ 2
Hydrolysis
+ +

As reserpine does not contain -COOH & -
OH groups, hence its hydrolysis product
reveal that reserpine is a di-ester.
The ester linkage in reserpine has been
further confirmed by its reduction with
5. Reduction:-
C33
H40
N2
O9
LiAlH4
C22
H30
N2
O4
CH2
OH
OMe
OMe
MeO
Reserpic Alcohol +

Structure of Reserpic acid
a) Molecular formula:-
b) Presence of one carboxyl group:-
By usual tests e.g. silver salt method
c) Presence one –OH group:-
Reserpic acid on oxidation yields a ketone that means it has
secondary alcoholic group.
d) Nature of two methoxy groups:-
By zeisels method.
e) Nature of two ‘N’ atom:-
It shown that it contains two ‘N’ atoms in heterocyclic
ring in the form of- i) Secondary ‘N’
ii) Tertiary amino group
C22
H28
N2
O5

f) Reduction of reserpic acid:-
On reduction with yields reserpic alcohol.
g) Oxidation of reserpic acid:-
On oxidation with KMnO4 it gives 4-methoxy N-
oxalyl anthranilic acid.
Thus one of the methoxyl group is present in meta
position to –NH group
NHCOCOOHMeO
COOH
KmnO4
C22
H28
N2
O5
4-methoxy N-oxalyl
Anthranilic acid
oxidation
Reserpic acid

h) Fusion with KOH :-
When reserpic acid is fused with potash it yields 5-
hydroxy isophthalic acid.
One of the acidic groups of isophthalic acid must be
present in m- position to each other this confirm by the
fact that reserpic acid when heated with acetic anhydride
yields a gamma lactone.
C22
H28
N2
O5
OH
COOH
HOOC
KOH
Fusion
5-hydroxy isophthalic acid
C22
H28
N2
O5
AC2
O
OOC
v-lactone

i) Dehydrogenation:-
When methyl reserpate is dehydrogenated
with selenium it yields a hydrocarbon of
molecular formula -
This hydrocarbon is also obtained by
dehydrogenation of Yohimbine with selenium
& was therefore named as Yobyrine.
C19
H16
N2
Methyl reserpate
Se
Dehydrogenation C19
H16
N2
DehydrogenationSe
Yobyrine
Yohimbine16-Apr-20 Krishna Pharmacy College, Bijnor 10

Structure of Yobyrine
1. Molecular formula:-
2. Zinc Distillation:-
3. Oxidation:-
When yobyrine is oxidized with permangnate it yields phthalic
acid
C19
H16
N2
C19
H16
N2
Zn dust
N
N
H
C2
H5
Distillation
+
Isoquinoline
3-ehtyl indole
C19
H16
N2
KmnO4
CH3
COOH
cr2
o3
COOH
COOH
Pthalic acid
o-Toulic acid
16-Apr-20
Krishna Pharmacy College, Bijnor
11

4. Condensation with aldehydes:-
Yobyrine gives condensation products
suggesting the presence of pyridine ring
with a methylene substitution
adjacent to the nitrogen.
On the basis of above facts following
structure has been postulated for yobyrine.
N
H
N
CH3
Yobyrine

Synthesis of Yobyrine
N
H
NH2
CH3
CH2ClOC
NH
N
H CO
CH2
CH3
POCl3
CH3
N
N
H
CH2
2-(3-indolyl)ethyl amine
O-tolyl acetyl chloride
+
1) -HCl
2)Reduction
2H-
dehydrogenation
N
H
N
CH3
Yobyrine

j) As Yobyrine is formed from reserpic acid it means
that reserpic acid may possesses the following
types of skeleton structures.
k) From the fact 5(g) it follows that one of the methoxyl
group is present in m-position to the NH group of indole
i.e. on C-11 i.e. Ome is at C-11.
N
N
H
SKELETON STRUCTURE

Reserpic acid when dehydrogenated yields 11-hydroxy-16-
methyl yobyrine,this may be only formed if –COOH group
is present on C-16
From the step (h) it follows that –COOH & -OH group are in
m-position to each other but –COOH group is present at C-
16 therefore –OH group must be at C-18.
C22
H28
N2
O5
N
N
H
CH2
OH
CH3 CH3
Se
dehydrogenation
11-hydroxy-16-methyl yobyrine

From purely biogenic reasons, the 2nd methoxyl
group has been assigned position C-17.
On the basis above mentioned facts the structure
of reserpic acid may be
N
N
H
OH
MeO
OMe
HOOC
Reserpic acid

Structure of Reserpine
As reserpine is di-ester of reserpic acid, it means that,
N
N
H
OH
MeO
OMe
HOOC
CH3
OH
COOH
OMe
OMe
MeO
N
H
N
O
MeO
H3
COOC
OMe
CO
OMe
OMe
OMe
Reserpic acid
+ +
Reserpine
-2 H2O

Synthesis of Reserpine
The structure of reserpine has been confirmed by
its synthesis given by Woodward et.al.
O
O
H2
C
C
C
CH
COOH
O
O
CH2
CH
COOH
CH
CH
NaBH4
OH
O COOH
C6
H5
COOH
OH
O COOH
O
Ac2
O
+
Diels Alder
reaction
Adduct
Reduction
of less
hindered
( =CO)
phenoxy benzoic
acid
p-Benzoquinone

Synthesis continue…..
O
O OCO
OH
O CO
O
O CO
O
CH3
ONa
CH3
OH
O CO
OMe
O
H2
SO4
O CO
OMeOH
O
Br
Cro3
Aluminium
isopropoxide
-
-H2O
NBS-

O CO
OMeO
Br
O
Zn CH3
COOH
O
OH
COOH
OMe
/
CH2
N2
Ac2
O
OSO4
-H2
O
i)
ii) Pyridine
iii)
OH
OH
O
OAc
MeO
COOCH3
HIO4
CH2
N2
OAc
CH2
CHO
COOCH3
MeO
COOCH3
i)
ii)
6-methoxy
tryptamine
N
H
NH2MeO

N
N
H
MeO C
H
CH2
OAc
OMe
H3
COOC
H3
COOC
NaBH4
CH3
OH
N
H
MeO
OAc
OMe
H3
COOC
N
O
POCl3
NaBH4
ii)
i)
i)
ii)

OAc
OMe
H3
COOC
N
N
H
MeO
OMe
N
N
H
OC O
OMe
i)Resolve
ii)NaOH
iii)Hcl
iv)DCC

Isomerisation
CH3
OH
OMe
OMe
HOOC
OMe
(CH3
)3
CCOOH
N
OMe
N
H
MeO
OC O
OMe
H3
COOC
N
N
H
MeO
OCO
OMe
OMe
OMe
Reserpic acid Lactone
Reserpine

classification
Kingdom : Plantae
Division : Magnoliophyta
Class : Magnoliopsida
Order : Gentianales
Family : Apocynaceae
Genus : Rauwolfia
Species : serpentina

Geographical Source:
Asia, America
Morphology:
Snake shaped, brown coloured, longitudinal
wrinkles tapering towards the end

extraction
Plant material
Macerated with alcohol
Alcholic extract
Reduce in volume and residue is treated in water
Filter
Ppt insoluble in water
Ppt contain reserpine16-Apr-20
Krishna Pharmacy College, Bijnor
26

ISOLATION: Ppt of crude drug
Add benzene with continuous stirring Filter, concentrate it
Filter
Filtrate Ppt remove
• Dry in vaccum
• Add 150 ml acetic acid in methanol and ammonium solution
• pH is adjusted to 8.5 and stand
• Crystalline substance appear
• Dry, add benzene
• Then it is chromatographed on 100 gm acid washed alumina
• Further fraction eluted with sequance of solvent from chloroform
• And crystallized from benzene

Chemistry:
Reserpine is an indole Alkaloid
Structure:
IUPAC Name:
(3β,16β,17α,18β,20α)-11,17-Dimethoxy-18-[(3,4,5-trimethoxybenzoyl)oxy]yohimban-
16-carboxylic acid methyl ester 3,4,5-trimethoxybenzoyl methyl reserpate
Melting point: 262-2700C.

Identification test
Chemical test:
Freshly fractured surface of the root when treated with
concentrated HNO3 shows red coloured medullary rays.
Reserpine gives a violet colour with vanillin in acetic
acid Rauwolfia serpentina.
Other test for alkaloid:
Dragondrof test
Mayer test
Hagers test
Wagners test
Tannic acid test

Mode of Action
Reserpine acts by blocking the vasicular monoamine
transporter VMAT, which normally transports free
norepinephrine, serotonin, and dopamine from the
cytoplasm of the presynaptic nerve into vesicles for
subsequent release into the synaptic cleft. The unprotected
neurotransmitters then metabolized by MAO and therefore
never reach the synapse.
By this mode of action this give mainly the
antihypertensive action and also use in the treatment of
dykynesia mania etc.

Therapeutic application:
Essential hypertension
Mild anxiety
Dyskinesia
Psychosis symptoms
Maintenance dose: 100 to 250 mg (once a day)
Side effects:
Nasal congestion
Nausia
Vomiting
Gastric intolerance
Gastric ulceration
diarrhoeia

References
1)“ Organic Organic Chemistry of Natural Products ”- by Gurudeep
Chatwal ,Vol-I.
2)“ Organic Chemistry ”by I.L.Finar , Vol-II .
3)“Pharmacognosy” by C.K.Kokate , A.P.Purohit , S.B.Gokhale
4) http://www.authorstream.com/Presentation/agarawaldinesh47-
1168554-reserpine/
5)http://www.authorstream.com/Presentation/abdulrazzaqM.PHAR
M-642751-reserpine-structural-elucidation/

16-Apr-20 Krishna Pharmacy College, Bijnor
33

Reserpine(Structure Elucidation, Extraction and Isolation)

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