Presentation

In vitro Production of Indole Alkaloids from two Egyptian
Catharanthus roseus cultivars by means of
Biotechnology. gy
Prof. Mohamed M. Yacout
Department of Genetics & Biotechnology
Faculty y of Agriculture g ‐ Alexandria Universityy

pharmaceuticals agrochemicals
colors
flavors fragrances
secondary
metabolites
food additives
MAN KIND AND PLANTS
biopesticides

IIn EEgypt, there h are more than h 835
pharmaceuticals and food companies
ALEXREP2009
I In vitro it PProduction d ti of f
Pharmaceutical Plant Products
By means of Biotechnology

Project Partners
1 .Faculty of Agriculture.
Alexandria University
2 .Faculty of Pharmacy.
Alexandria University
3. High institute of health.
Alexandria Alexandria University
4. Mubarak City for Scientific
Research

Catharanthus roseus (Madagascar Periwinkle)
Produce a wide range of secondary metabolites
Aj Ajmalicine li i and d SSerpentine ti – hhypertension t i
Vinblastine and Vincristine – anticancer drugs used
to treat lymphomas y p and leukemia
vinblastine vincristine

Objectives of the project
study DNA
markers related to
Vincristine
production
To produce in
vitro, C. roseus
in plant cell
cultures from
local plants
d ti varieties
to enhance
production, both
in quality and
quantity of
Vincristine
to optimize plant
cell cultures
growth conditions
for Vincristine
production

Objectives of the project
study DNA TTo produce d iin vitro, it
C. roseus in plant
cell cultures from
local plants varieties
markers related
to Vincristine
production
to enhance
production, d i both b h
in quality and
quantity of
Vi Vincristine
i ti
to optimize plant
cell cultures
growth conditions
for Vincristine
production

Objectives of the project
study DNA
markers related to
Vincristine
production
to enhance
production,
both in quality
and quantity
of Vincristine
To produce in
vitro, Vincristine in
plant cell cultures
from local plants
varieties
to optimize plant
cell ll cultures l
growth conditions
for Vincristine
production

Objectives of the project
study DNA
markers
related to
Vincristine
production varieties
To produce in
vitro, Vincristine in
plant cell cultures
from local plants
varieties
to enhance to optimize plant
production, d i bboth hiin
cell ll cultures l
quality and growth conditions
quantity of for Vincristine
Vincristine
production

MATERIALS
AND
METHODS

Two Egyptian
cultivars ii.e. e Pink
and white winkle.

RESULTS

Cell suspension culture
Cell Cell suspension from leaf explants in MS media media

Cell suspension from leaf explants in MS
media

Figure 1: Callus shape at 10 x power in binocular
microscope formed from leaf explants of C roseus

Figure 2: Induced callus percentage from leaf explants on MS
media with 1-Naphthyl acetic acid (NAA) and kinetin (Kin).

2,4-Dichlorophenoxy
acetic acid (2,4-D)
concentration mg/l
mg/l
2,4 4D1,5
2,4 2,4 4D1
2,4D .5
IAA IAA AA.5 AA.5
IAA1 IAA1 A1 A1
IAA1.5 IAA1.5 .5 .5
90
80
70
60
50 Callus formation
40 percentage
30
20
10
0
Indole 3-acetic acid (IAA) ( )
concentration mg/L
Figure 3: Callus formation percentage from leaf explants on MS media with
2,4-Dichlorophenoxy acetic acid (2,4-D) and Indole 3-acetic acid (IAA).

Cell culture
fresh weight
(g/L)
6
5
4
3
2
1
0
8000
7000
6000
5000
4000
3000
2000
1000
0
N1.5Kin0.5 N0.5Kin0.5 2D1IA0.5 2D1IA1 2D1IA1.5 2D1.5IA0.5 2D1.5IA1 2D1.5IA1.5
2,4 D, NAA, IAA concentrations mg/L
Figure 4: Fresh weight of cells yield of cell suspension from leaf
explants in MS media with 2,4-Dichlorophenoxy acetic acid (2,4-
D), 1-Naphthyl acetic acid (NAA) and/or kinetin (Kin).

Production of alkaloids
Applying TLC technique for extracts of Alkaloids from
cells and callus showed that there was difference
between pink and white plants in types and quantity.
Pink plants cells and callus gave two types of spots
on TLC plates while white plants gave one spot.
Thee intensity e s y of o theespots spo s which c reflect e ec thee
concentration was darker in case of pink plants more
than white ones. On the other hand, , callus spots p
were stronger than that of cells.
•
•
•

Objectives of the project
study DNA
markers related to
Vincristine
production
To produce in
vitro, C. roseus
in plant cell
cultures from
local plants
d ti varieties
to enhance
production, both
in quality and
quantity of
Vincristine
to optimize plant
cell cultures
growth conditions
for Vincristine
production

Dr Fathy El-Fiky &
Dr Hattem Meky
Department of Pharmacognosy,
Pharmacognos
Faculty of Pharmacy,
Alexandria University
Dr. Wafaa Masoud
Biotechnology laboratory,
Faculty of Agriculture,
Alexandria University
From our group
Thanks to:

In vitro Production of Indole Alkaloids from two
Egyptian Catharanthus roseus cultivars by means
of Biotechnology
Biotechnology.
Mohamed M. Yacout,* Fathy El‐Fiky,*** Hattem Mekky*** and Wafaa M. Masoud**
*Department of Genetics and biotechnology, biotechnology Faculty of Agriculture, Agriculture Alexandria University
** Biotechnology laboratory, Faculty of Agriculture, Alexandria University
***Department of Pharmacognosy, Faculty of Pharmacy, Alexandria University
(Email (
to: mohamed.yakout@alex
y
‐ agr.edu.eg g g )

In vitro Production of Indole Alkaloids from two
Egyptian Catharanthus roseus cultivars by means
of Biotechnology
Biotechnology.
Mohamed M. Yacout,* Fathy El‐Fiky,*** Hattem Mekky*** and Wafaa M. Masoud**
*Department of Genetics and biotechnology, biotechnology Faculty of Agriculture, Agriculture Alexandria University
** Biotechnology laboratory, Faculty of Agriculture, Alexandria University
***Department of Pharmacognosy, Faculty of Pharmacy, Alexandria University
(Email ( to: mohamed.yakout@alex
y
‐ agr.edu.eg g g )
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