1

5. Biosynthesis of Alkaloid Natural Products

5.1. Alkaloids are derived from amino acids
Nitrogen-containing compounds, with a slightly basic character, have been isolated from many different
organisms, mostly plants and microorganisms, and are biosynthesized from amino acids - these are called
alkaloids. There are probably over 10'000 known alkaloids, having very diverse structures. They can
nevertheless be classified into families, on the basis of structural similarities and the amino acids that are
used for their biosynthesis Some alkaloids are also produced using building blocks derived from other
secondary metabolic pathways, such as terpenoids, polyketides and peptides. Some of the important classes
of alkaloid are shown below:
e.g. Pyrrolidine, Pyrrolizidine and Tropane Alkaloids
Me

NH3
H 3N

MeN

HO

COO

Ornithine

OH

Hygrine

Me

Scopolamine

N
Retronecine

Ph

MeN

OH

Tropinone
O

e.g. Piperidine, Pyridine und Quinolizidine Alkaloids

OH

NH3
H3N

COO

N
H
Coniine

Lysine

Me

N
Me
Nicotine

N
Me
N-Methylpelletierine

Lupinine

N
N
O

Sparteine

Lycopodine

z.B. Isoquinoline Alkaloids

MeO

MeO
COO
NH3

MeO
MeO

Phenylalanine
Tyrosine

NMe

HO

OMe

Papaverine
Autumnaline

MeO

OMe
OH

HO

MeO

HO

NHAc
MeO

O
MeO

Colchicine

Dopamine

Morphine

HO

OMe

NH2

HO

NMe

z.B.Indole Alkaloids

COO

NH2

NH3
N
H

N
H

N
H
Tryptamine

Tryptophan
+ Terpene

MeO

N
Me

OH
COOMe
Vindoline

MeOOC
Geissoschizine

OH

N
H
MeOOC
Vinblastine

OH

N
N
H

MeO

N
Me
MeOOC

OH

OAc

COOMe

Catharanthine

OAc

2
5.2. Benzylisoquinoline Alkaloids
Of special interest within the family of isoquinoline alkaloids are those containing the 1benzyl(tetrahydro)isoquinoline skeleton, which are found in many different plants. Studies on the
biosynthesis of these compounds made progress as soon as radioactively labelled compounds (14C and 3H)
became available. Potential precursors could be fed to intact plants, and later the natural prodicts could be
isolated from the plants, and then analyzed chemically to detemine whether, and if so, where the radioactive
labels had been incorporated. In this way, it was shown that the benzylisoquioline alkaloids are constructed
from two molecules of tyrosine:
Hydroxylase

Decarboxylase
(PLP)
HO

HO

NH2
HO

COOH
O

NH2

HO
Tyrosin

Transaminase
(PLP)

NH2

NH

HO

COOH

CHO
HO

Decarboxylase
(TPP)
HO

HO

HO

Norcoclaurine

The formation of norcoclaurine is catalyzed by an enzyme, which in effect catalyzes a Pictet-SpenglerReaction. The reaction shown actually occurs spontaneously in aqueous solution, but then slowly gives
racemic product, whereas the enzymic reaction runs much faster and gives optically pure product:
HO
HO

NH2

HO

HO
NH

HO

HO

CHO

HO

HO

HO

NH

HO

NH

Norcoclaurine

HO

Next, the norcoclaurine is converted into (S)-reticuline :

HO
HO

SAM

SAM

MeO

Hydroxylase

NH

HO

SAM

N-Me
H

HO
Me-O

HO
Norcoclaurine

Reticuline

Reticuline is used for the biosynthesis of many other benzylisoquinoline alkaloids, amongst others, the socalled aporphine alkaloids, e.g.:

HO
HO
MeO

MeO

MeO

MeO
NMe

HO

HO
MeO

NMe
H

MeO
MeO
MeO

NMe
H

Glaucine

3
An important step here is the formation of a direct aryl-aryl bond. This occurs in an oxidative phenol
coupling reaction. Nature has evolved a series of hemoproteins of the cytochrome P450 family that catalyze
specific oxidative phenol coupling reactions (not hydroxylations, compare earlier). Such coupling reactions
are well known in synthetic chemistry, where they can be carried out with phenolic compounds, under basic
conditions, using K3Fe(CN)6 as oxidizing agent, e.g.:
MeO

MeO
NMe

HO

-2H+

2 FeII

MeO

NMe

MeO
NMe

K3Fe(CN)6

MeO

MeO

MeO
O

OH

O
MeO

MeO

MeO
NMe
H

2 FeIII

MeO

NMe

HO

NMe
H

MeO

MeO

HO
MeO

OH
ortho-para

NMe

HO

ortho-ortho

Such reactions tend to produce mixtures of products, because the free radical intermediates can often couple
in more than one way. The enzymes, however, catalyze only one pathway specifically. The mechanisms of
the enzymic reactions are not well understood, but require molecular oxygen as well as the hemoprotein
(P450). The oxidizing power of compound-I is used to drive the coupling reaction, e.g.:

OH2

electrons
+ O2

FeIII
S-Cys
P450 enzyme
(resting state)

H2O

HO

OH

HO

OH

FeIV

Fe

Fe

S-Cys

S-Cys

S-Cys

+ H2O

compound-I

Oxidative phenol coupling reactions are often found in alkaloid biosynthesis. Perhaps the best-known
example occurs during the biosynthesis of morphine.
Morphine is a highly-potent opiate analgesic drug and is the principal active agent in opium and the
prototypical opioid. It is also a natural endocrine product in humans and other animals. Like other opiates,
e.g., diacetylmorphine (heroin), morphine acts directly on the central nervous system (CNS) to relieve pain,
and at synapses of the nucleus accumbens in particular. Studies done on the efficacy of various opioids have
indicated that, in the management of severe pain, no other narcotic analgesic is more effective or superior to
morphine. Morphine is highly addictive when compared to other substances; tolerance, physical and
psychological dependences develop very rapidly. The word "morphine" is derived from Morpheus, one of
the Greek gods of dreams.
The opium poppy is Papaver somniferum.

(R)-Reticuline is an important intermediate in the biosynthesis of morphine, and is produced by

racemization of (S)-reticuline in a redox process, as shown below:
MeO

MeO
NMe

HO

Oxid.

HO

HO

Red.

MeO

HO
N-Me

N-Me
MeO

MeO

(S)-Reticuline

MeO

oxid.
PhenolCoupling

(R)-Reticuline

Salutaridine

OH

Salutaridine is found as a minor alkaloid constituent in the opium poppy:

MeO

MeO
Reduction

HO

Acetyl-CoA

HO

AcOH MeO
CoASH
O

N-Me

N-Me

N-Me

MeO

MeO
Salutaridinol

MeO

O
N-Me

MeO
Thebaine

OH

MeO

MeO

O
N-Me

N-Me

O
Neopinone

OH
MeO

HO

O
N-Me
HO

Codeine

O
N-Me
HO

Morphine

Codeinone

5
The biosynthesis of morphine in the opium poppy was one of the first alkaloid pathways to be elucidated
with the aid of 14C-labelled precursors. It was shown that [2-14C]-tyrosine is incorporated into morphine,
with the 14C label appearing at the positions indicated above. This was proven, by degrading the 14C-labelled
morphine in the following way:
1) MeI / K2CO3 /
MeOH
MeO
2) Ag2O, then
pyrolysis

HO

NMe2

Morphine

HO

MeO
O

1) Ac2O
2) CrO3

MeO

NMe2

EtO
MeO

1) H2O2
2) NaOH/H2O AcO

3) H3O

COOH

MeO

MeO
H2SO4
!

O
NaOH/
Me2SO4

HO

N-Me
HO

MeO

EtONa /
EtOH, !

MeO
heat/ H+
MeO

+ CO2

HOOC

Another interesting benzylisoquinoline alkaloid is colchicine. Colchicine was originally extracted from
plants of the genus Colchicum (Autumn crocus, Colchicum autumnale, also known as the "Meadow
saffron"). Originally used to treat rheumatic complaints and especially gout, it was also prescribed for its
cathartic and emetic effects. Its present medicinal use is mainly in the treatment of gout; it is also being
investigated for its potential use as an anti-cancer drug.
Colchicine inhibits microtubule polymerization by binding to tubulin, one of the main constituents of
microtubules. Tubulin is essential for mitosis, and therefore colchicine effectively functions as a "mitotic
poison" or spindle poison. Since one of the defining characteristics of cancer cells is a significantly
increased rate of mitosis, this means that cancer cells are significantly more vulnerable to colchicine
poisoning than are normal cells. However, the therapeutic value of colchicine against cancer is (as is typical
with chemotherapy agents) limited by its toxicity against normal cells.
In 2008, the Botanic Gardens Conservation International (representing botanic gardens in 120 countries)
stated that "400 medicinal plants are at risk of extinction, from over-collection
and deforestation, threatening the discovery of future cures for disease." These
included Yew trees (the bark is used for the cancer drug taxol (paclitaxel));
Hoodia (from Namibia, source of weight loss drugs); half of Magnolias (used as
Chinese medicine for 5,000 years to fight cancer, dementia and heart disease);
and Autumn crocus (for gout). The group also found that 5 billion people
benefit from traditional plant-based medicine for health care.
Early labelling experiments showed that tyrosine and phenylalanine are required for colchicine biosynthesis,
and that autumnaline is a key intermediate. However, the Phe provides a C6C3 unit rather than a C6C2
fragment:

6
Tyrosine

MeO

COOH

MeO

H
Colchicum

COOH

H 2N

NHAc

NMe

HO

NH2

HO

MeO

MeO

MeO

MeO

(S)-Autumnaline

OH

Phenylalanine

Colchicine

OMe

The seven membered tropolone ring was shown by labelling experiments to originate by ring expansion of
the tyrosine-derived aromatic ring, including the adjacent benzylic carbon atom.
Dopamine

NH2

HO

HO

HO

HO

HO

NH

HO

H
CHO
cf. above

Tyrosine
Phenylalanine

OH

OH
OH

HO

HO
N-Me

N-Me

MeO

MeO

MeO

MeO
Isoandrocymbine

MeO

MeO

OH

(S)-Autumnaline

Androcymbine has been isolated from Androcymbium melanthioides. The later steps have not been proven,
but may involve the following reactions:
Oxidation
MeO

HO

MeO

MeO

NH-Me
HO

NHMe
MeO
MeO

MeO
Androcymbine

N-Me
MeO

MeO

MeO
MeO

MeO

MeO

MeO

MeO
Colchicine

MeO

MeO
O

HCHO
OMe

Me

NH

NH-Me
H
Demethylation

MeO

Acetylation
OMe

7
Various types of alkaloids are encountered in the daffodil family, called the Amaryllidaceae alkaloids
(Amaryllidaceae is the botanical name of a family of flowering plants. The plants are herbaceous perennials
that grow from bulbs, often with showy flowers). The Amaryllidaceae family includes Amarylis, Narcissus
and Galanthus, and the alkaloid content of bulbs from most
members makes them toxic. However, galanthamine from
daffoldils and snowdrops is currently an important drug for
the treatment of the symptoms of Alzheimer's disease. The
natural sources of galanthamine are certain species of daffodil
and because these species are scarce and because the isolation
of galanthamine from daffodil is expensive (a 1996 figure
specifies 50,000 US $ per kilogram; the yield from daffodil is 0.1-0.2% dry weight) alternative synthetic
sources have been developed. Galanthamine acts as a competitive inhibitor of acetylcholinesterase, and
enhances cognitive functions by raising acetylcholine levels in brain areas lacking cholinergic neurons. It
does not cure the condition, but merely slows the rate of cognitive decline.
Phe and Tyr are again the starting materials used for the biosynthesis of the Amaryllidaceae alkaloids:
HO
L-Phe
HO

HO

CHO

SAM

HO

H2N

NH

HO

L-Tyr
OH

Norbelladine

Thereafter, three different modes of phenol coupling are seen:

para-orthocoupling

HO

HO
MeO

MeO
NH
HO
4'-O-methylnorbelladine
OH

HO
MeO

NH

HO

HO
OH

Norpluvine

MeO
NH

HO

NMe

OH
HO

para-paracoupling

HO
MeO

O
ortho-paracoupling

O
N

MeO

Lycorine

O
NH

HO

NMe
HO

OH

MeO

MeO
O

NMe

Oxocrine

MeO
HO

MeO
Galanthamine

OMe
OH

HO

Haemanthamine

8
5.3. Indole Alkaloids (see Nat. Prod. Rep. 2006, 23, 532)
The simplest representative of the indole alkaloids are the natural amines tryptamine und serotonin, which
are biosynthesized from the amino acid tryptophan (Trp):
COO

R = H Tryptamine
R = OH Serotonin

NH2

NH3

N
H

N
H

Serotonin is a monoamine neurotransmitter synthesized in serotonergic neurons in the central nervous

system (CNS), and enterochromaffin cells in the gastrointestinal tract of animals including humans.
Serotonin is also found in many mushrooms and plants, including fruits and vegetables. Serotonin is
believed to play an important role as a neurotransmitter, in the modulation of anger, aggression, body
temperature, mood, sleep, sexuality, and appetite as well as stimulating vomiting.
The vinca alkaloids are a very interesting class of indole alkaloids, and include vinblastine, vincristine,
vindesine and vinorelbine. These alkaloids are produced by plants of the genus Catharanthus. Catharanthus
(Madagascar Periwinkle) is a genus of eight species of herbaceous perennial plants, seven endemic to the
island of Madagascar, the eighth native to the Indian subcontinent in southern Asia. One species, C. roseus,
has been widely cultivated, and after introduction has become
an invasive species in some areas. C. roseus has also gained
interest from the pharmaceutical industry; the alkaloids
vincristine and vinblastine from its sap have been shown to be
an effective treatment for leukaemia. Although the sap is
poisonous if ingested, some 70 useful alkaloids have been
identified from it. In Madagascar, extracts have been used for
hundreds of years in herbal medicine for the treatment of
diabetes, as hemostatics and tranquilizers, to lower blood
pressure, and as disinfectants. The extracts are not without their
side effects, however, which include hair loss.

N
N
H

H
MeO

MeOOC
OH

Geissoschizine

Vindoline

N
OAc
H
Me
OH
MeOOC

Stemmadenine

COOMe

Catharanthine

H N
H

OH

N
H
MeOOC

Strychnine
O

H
H

N
H
MeOOC

Vinblastine

OH

MeO

N
Me
MeOOC

OH

OAc

The structures of these alkaloids reveal that not only Trp is required for the biosynthesis. A C10 fragment is
also needed, and is provided from terpene metabolism. Strychnine biosynthesis also incorporates one
acetate unit (in red above). The important C10 fragment is produced from geraniol, and is called
secologanin:

9
HO

Me
H

OH

CHO
O-Glu

O-Glucose

MeOOC

Secologanin

Loganin

Geraniol

MeOOC

Secologanin is a glucoside, which can be cleaved by hydrolysis under acidic conditions:

HO
CHO

OH
O

OH
OH

H3O

MeOOC

The formation of the indole alkaloids begins with the condensation of tryptamine and secologanin, catalyzed
by strictosidine synthase (STR, see below):

N
H

NH2

NH

N
H

CHO

O-Glu

O-Glu
MeOOC

MeOOC

N
H

MeOOC

NH
OGlu
O

Strictosidine

Strictosidine is then a key intermediate in the formation of over 1000 different indole-terpene alkaloids.

10
For example, the Corynanthe alkaloids:
Glucose
NH

N
H

N
H

O-Glu

MeOOC

N
H

NADPH

OH

MeOOC

MeOOC

OH

MeOOC

OH

OH

MeOOC

N
H

MeOOC

N
H
CHO

Acetal

NH

Geissoschizine

N
H

NADPH

N
H

H
H

Ajmalicine
MeOOC

Me

N
H

(Imine
reduction)
MeOOC

N
H
O

2 NADPH

H
MeOOC
OH

N
H
Yohimbine

H
H

MeOOC
OH

Yohimbine is the principal alkaloid of the bark of the West-African evergreen Pausinystalia yohimbe Pierre
(formerly Corynanthe yohimbe), family Rubiaceae (Madder family). There are 31 other yohimbane
alkaloids found in Yohimbe. In Africa, yohimbine has traditionally been used as an aphrodisiac. Yohimbine
hydrochloride is a standardized form of yohimbine that is available as a prescription drug in the United
States, and has been shown to be effective in the treatment of male impotence. Yohimbine hydrochloride
has also been used for the treatment of sexual side effects caused by some antidepressants, female
hyposexual disorder, as a blood pressure boosting agent in autonomic failure, xerostomia, and as a probe for
noradrenergic activity.
Ajmaline was first isolated from the roots of Rauwolfia serpentina, a species of flowering plant in the
family Apocynaceae. It is one of the 50 fundamental herbs used in traditional
Chinese medicine, where it has the name shgn m () or ynd shm
(). The extract of the plant has also been used for millenia in India
it was reported that Mahatma Gandhi took it as a tranquilizer during his
lifetime. Ajmaline is a class Ia antiarrhythmic agent, a group of
pharmaceuticals that are used to suppress fast rhythms of the heart (cardiac
arrhythmias), such as atrial fibrillation, atrial flutter, ventricular tachycardia,

11
and ventricular fibrillation. Ajmaline functions by blocking Na-channels in cell membranes.
Rauwolfia caffra is the South African quinine tree. Rauwolfia serpentina, or Indian Snakeroot or
Sarpagandha, contains a number of bioactive chemicals, including ajmaline, deserpidine, rescinnamine,
serpentinine, and yohimbine. Reserpine is an alkaloid first isolated from R. serpentina, and was widely used
as an antihypertensive drug. It had drastic psychological side effects and has been now replaced by bloodpressure-lowering drugs that lack such adverse effects. But in herbal use it is a safe and effective resource
for hypertensive patients. The pharmaceutical companies have stopped producing this drug as reserpine or
deserpedine. It is only available currently in the U.S. as a herbal medicine over the Internet.
The pathway to ajmaline has been well documented, although few mechanistic studies have been reported
so far on the biosynthetic enzymes:
OHC

OHC
see above

NH

N
H

N
H

OGlu

Polyneuridine Aldehyde

Dehydrogeissoschizine

Strictosidine

N
H

MeOOC

COOMe

COOMe

MeOH
CO2
O
AcO
N

OH

N
H

Oxidation

H
N
H

16-epi-vellosimine

Vinorine

Reduction

AcO
N
HH

Vomilenine
NADPH

Acetyl-CoA

AcO

N
H

Dihydrovomilenine

HO

AcO

NADPH
OH
Reduction

N
H H

Hydrolysis
N

OH

17-O-Acetylnorajmaline

SAM

H
Me
H

Ajmaline

OH

Catharanthine is a member of the so-called iboga family of indole alkaloids. It is one of the many alkaloids
present in Catharanthus roseus. It is produced along with many other
Catharanthus alkaloids by factory farming in China. It can be used as a
starting material for the synthesis of the anti-tumor drugs, vinblastine
and vincristine. Vindoline (an Aspidosperma alkaloid) is another
important component of the bis-indole alkaloids, typified by vinblastine
and vincristine, also produced by C. roseus. Some of the biosynthetic
steps have been documented, but the enzymes have not yet been studied
in detail. A fascinating proposal was made to explain how catharanthine
and vindoline might be produced from geissoschizine. Tabersonine is a
known intermediate, and the steps from tabersonine have been established; the rest is hypothetical -

12
Hypothetical

N
H

H
MeOOC
Geissoschizine

MeOOC

CHO

MeOOC

CHO

Redox
changes

N
H

CHO

N
N

N
H
MeOOC

N
CH2OH

MeOOC

CHO

CHO

MeOOC

preakuammicine
Hypothetical

NADH
N

N
N
H

N
H

MeOOC

CH2OH

MeOOC

MeOOC
dehydrosecodine

Oxidation

N
H

N
H

CH2OH

stemmadenine

HO

N
H

COOMe

N
N
H
MeOOC

COOMe

Tabersonine

16-Hydroxytabersonine

+ H 2O
2 x SAM
N

N
Oxidation

Catharanthine
Acetyl-CoA

MeO

N
Me HO

MeO
COOMe

Desavetoxyvindoline

COOMe

N
Me HO
Deacetylvindoline

OH

COOMe
MeO

N
Me HO

OCOCH3
COOMe

Vindoline

Vinblastine and vincristine are anti-mitotic drugs used to treat certain kinds of cancer, including Hodgkin's
lymphoma, non-small cell lung cancer, breast cancer and testicular cancer. They bind to tubulin, thereby
inhibiting the assembly of microtubules. They are M phase cell cycle specific, since microtubules are a
component of the mitotic spindle and the kinetochore, which are necessary for the separation of
chromosomes during anaphase of mitosis. Toxicities include bone marrow suppression (which is doselimiting), gastrointestinal toxicity, potent vesicant (blister-forming) activity, and extravasation injury (forms
deep ulcers).

13
The coupling of catharanthine and vindoline can be catalyzed by a relatively non-specific peroxidase (a
hemoprotein). It is possible that a similar enzyme specifically catalyzes this coupling in C. roseus.

Peroxidase
H2O2

HO

N
H

N
N

N
COOMe

Catharanthine

COOMe

COOMe

Coupling
N

N
Me HO

MeO
N
H
MeOOC

OCOCH3
COOMe

Vindoline

Reduction
MeO

N
N
H
MeOOC

N
Me
MeOOC

OH

OAc

N
N

[O]
MeO

N
Me
MeOOC

OH

OAc
Reduction

N
H
MeOOC

OH

MeO
Vinblastine (R = Me)
Vincristine (R = CHO)

N
R
MeOOC

OH

OAc

Vinblastine is only present at low levels in C. roseus (0.0002% of dry leaf wt). Over 500 kg of catharanthus
is needed to produce 1g of pure vincristine. Much effort has been put into the synthesis of the dimeric
alkaloids, starting from the monomers, which can be isolated from the plant in much higher yields. One
example is shown below:

14

N
N
H

N
COOMe

COOMe

COOMe

Catharanthine
+

N
N
H
MeOOC

N
Me HO

MeO

OCOCH3
COOMe

Vindoline

CONH2

MeO

N
Me
MeOOC

OH

OAc

COOH

- 40 oC

N
H
MeOOC

N
MeO
N
H
MeOOC

OH

MeO

N
Me
MeOOC

Vinblastine

N
Me
MeOOC

OH

OH

OAc

1) FeCl3, air
2) NaBH4

OAc

5 steps. 40% yield overall

(see also: J.Am.Chem.Soc., 2008, 130, 420).

Finally, note that strictosidine is also the precursor to the quinoline alkaloids, including the important antimalarial drug quinine.

NH

N
H

O-Glu

CHO

N
H

N
H

H
O

MeOOC

HO

OH

MeOOC

MeO
N
Quinine

NH2

CHO