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Thesis · September 2009
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Mansoura University
Faculty of Science
Chemistry Department
Isolation and Structure Elucidation of Natural Products from

Some Medicinally Important Plant Species
A Thesis
Submitted to the Faculty of Science, Mansoura University

For the Degree of Master of Science in Chemistry
(Organic Chemistry)
By
Amr El Sayed Ahmed El Demerdash

Demonstrator
B. Sc. (Chemistry), Mansoura University (2004)
Supervised by
Prof. Dr. M. Abdel-Mogib Prof. Dr. A. M. Dawidar

Professor of Natural Products' Chemistry Professor of Organic Chemistry
Chemistry Department, Chemistry Department,
Faculty of Science, Faculty of Science,
Mansoura University Mansoura University
Dr. E. M. Keshk
Associate Professor of Organic Chemistry
Chemistry Department,
Faculty of Science,
Mansoura University
2009
Mansoura University
Faculty of Science
Title of the thesis: Isolation and Structure Elucidation of Natural

Products from Some Medicinally Important Plant Species.
Student name: Amr El Sayed Ahmed El Demerdash
Supervised by:
Name Position Signature
Prof. of Natural Products' Chemistry
Prof. Dr. Chemistry Department,
Faculty of Science,
M. Abdel-Mogib Mansoura University
Prof. of Organic Chemistry

Prof. Dr. Chemistry Department,
Faculty of Science,
A. M. Dawidar Mansoura University
Associate professor of Organic Chemistry
Chemistry Department,
Dr. E. M. Keshk Faculty of Science,
Mansoura University
Head of Chemistry Vice-Dean of Faculty of Science Dean of Faculty

Department. for postgraduate and Research of Science.
affairs.
Prof. Dr. A. I. Ahmed Prof. Dr. El-Metwally M. Prof. Dr. Taha Z.

El-Abbasy Sokkar
Examination Report
Title the thesis: Isolation and Structure Elucidation of Natural
Products from Some Medicinally Important Plant Species
Student name: Amr El Sayed Ahmed El Demerdash
Supervisors
Name Position
Prof. of Natural Products' Chemistry
Prof. Dr. M. Abdel-Mogib Chemistry Dept., Faculty of Science,
Mansoura University
Chemistry Dept., Faculty of Science,
Prof. Dr. A. M. Dawidar
Mansoura University
Associate professor of Organic Chemistry
Chemistry Dept.,
Dr. E. M. Keshk Faculty of Science,
Mansoura University
Discussion and judgment team

Name Position
Prof. of Pharmacognosy
Prof. Dr. A. M. El Shami Pharmacognosy Dept., Faculty of Pharmacy,
Cairo University
Prof. Dr. A. A. Fahmi Chemistry Dept., Faculty of Science,
Cairo University
Chemistry Dept., Faculty of Science,
Prof. Dr. A. M. Dawidar
Mansoura University
Prof. Dr. M. Abdel-Mogib Chemistry Dept., Faculty of Science,
Mansoura University
Head of Vice-Dean of Faculty of Science Dean of Faculty of

Chemistry Department. for postgraduate and Research Science.
affairs.
Prof. Dr. A. I. Ahmed Prof. Dr. El-Metwally M. Prof. Dr. Taha Z.

El-Abbasy Sokkar
Mansoura University
Faculty of Science
NOTE
The present thesis is submitted to the Faculty of Science, Mansoura
University for the Degree of Master of Science in Chemistry (Organic
Chemistry).
Beside the work carried in this thesis, the candidate Amr EL Sayed
Ahmed EL Demerdash has pursued the following postgraduate courses for one
academic year:
1- Photochemistry and applied Spectroscopy
2- Advanced Physical organic Chemistry
3- Organic synthesis
4- Dyes
5- Organometalic Chemistry
6- Fluorocarbon Chemistry
7- Natural products Chemistry
8- Biochemistry
9- Petroleum Chemistry
10- Polymer Chemistry
11- Heterocyclic Chemistry
12- Enviromental and Analytical Chemistry
13- Molecular rearrangements
He had successfully passed the final examination of these courses in Jan.

2008.
Head of Chemistry Department
Prof. Dr. A. I. Ahmed

First of all thanks for Allah, the most beneficent, the most
merciful.
I wish to express my sincere appreciation and gratitude to

Prof. Dr. M. Abdel-Mogib, Professor of Chemistry of Natural
products, Faculty of Science, Mansoura University, for introducing
me to the field of natural products, suggesting the research point, his
continuous guidance, help, precious advice, real support and
valuable criticism.
I would like to express my cordial thanks and gratitude to

Prof. Dr. A. M. Dawidar, Emeritus Professor of Chemistry of
Natural products, Faculty of Science, Mansoura University, for his
guidance, and continuous encouragement.
I would like to express my deep appreciation and gratitude to

Dr. E. M. Keshk, Associate professor of organic chemistry for her
valuable guidance, advices and valuable support.
Deepest appreciation and thanks to Dr. Z. Abou El- Naga,

Lecturer of Entomology, Zoology Dep., Faculty of science,
Mansoura University for insecticidal evaluation experiments.
Finally, I would like to thank all the stuff members of

Chemistry Department, Faculty of Science, Mansoura University,
for their support.
AMR El DEMERDASH
CONTENTS
Summary
I- Introduction. 1
1.1- General 1
1.2. Cynanchum acutum L. 1
1.2.1. Botanical aspects, description and distribution in Egypt. 1
1.2.2. Medicinal uses of some Cynanchum species. 3
1.2.3. Medicinal and biological activities of Cynanchum acutum L. 3
1.2.4. Phytochemical constituents of Cynanchum genus. 3
1.2.5. Chemical constituents of Cynanchum acutum L. 11
1.3. Curcuma longa L. 12
1.3.1. Botanical aspects, description and distribution. 12
1.3.2. History and medicinal uses. 13
1.3.3. Biological activities of Curcuma longa L. 14
1.3.4. Phytochemical constituents of Curcuma longa L. 20
1.3.4.1. Constituents of volatile oil of Curcuma longa L. 20
1.3.4.2. Curcuminoids. 23
II- Aim of the Work. 26
III- Results and discussion. 27
2.1. Processing of Cynanchum acutum L. aerial parts. 27
2.1.1. Study of petroleum ether extract. 27
2.1.2. Characterizations of compound 1. 27
2.1.3. Characterizations of compound 2. 30

2.1.4. Structure elucidation of compound 3. 32
2.2. Study of the methylene chloride extract. 35
2.2.3. Study of methylene chloride extract using GC/MS. 38
2.3. Study of Ethyl acetate extract. 47
3.1 Processing of Curcuma longa L powdered rhizomes. 48
3.2. Study of petroleum ether extract of Curcuma longa L. 49
3.2.1. Structure elucidation of sesquiterpene ketone 16. 49
3.2.2. Structure elucidation of sesquiterpene ketone 17. 52
3.3. Study of chloroform extract of Curcuma longa L. 55
3.3.3 Structure elucidation of compound 22. 60
3.4. Insecticidal activities of the volatile oil and 1- dehydrogingerdione (19) isolated from
Curcuma longa against the confused flour beetles Tribolium confusum. 62
IV. Experimental. 69
3.1. Instrumentations. 69
3.2. Plant material. 70
3.3 Spray reagents. 70
3.4. Processing of plant material under investigation. 71
3.4.1. Processing of plant material Cynanchum acutum L. 71

3.4.1.2. Isolation of coumarins from methylene chloride extract. 72
3.4.1.2.1. Chemical composition of methylene chloride extract by GC/MS. 72
3.3.1.3. Investigation of ethyl acetate extract. 72
3.4.2. Charactrization of compounds isolated from Cynanchum acutum L. 73
3.4.3. Processing of plant material of Curcuma long L. 76
3.4.3.1. Chromatography of the petroleum ether extract. 76
3.4.3.2. Chromatography of the chloroform extract. 76
3.4.3. Characterization of isolated compounds from Curcuma longa L. 77
3.5. Insecticidal and antifeedant activities of volatile oil and 79
1-dehydrogingerdione isolated from Curcuma longa L. petroleum ether
extract against the confused flour beetles Tribolium confusum.
References 83
Appendix 95
Publication
Summary in Arabic.
.
Abreviations
Abréviations Scientific meaning

UV Ultraviolet
IR Infrared
MS Mass Spectrometry
GC/MS Gas chromatography/Mass Spectrometry
1
H NMR Proton Nuclear Magnetic resonance
13
C NMR Carbon-13 Nuclear Magnetic resonance
GC Gas chromatography
HPLC High performance liquid chromatography
GLC Gas liquid chromatography
Rf Retardation factor
CC Column Chromatography
TLC Thin Layer Chromatography
PTLC Preparative Thin Layer Chromatography
LC50 Lethal concentration for 50% of individual numbers
LC90 Lethal concentration for 90% of individual numbers
ODI Ovipostion deterrent indices
δ Chemical shift value in ppm
ppm Parts per million
MHz Mega hertz
m/z Mass over charge
Rt Retention time
pet. ether Petroleum ether
MeOH Methanol
CH2Cl2 Dichloloromethane
CDCl3 Deuterated chloroform
(CD3)2CO Deuterated acetone
List of Tables
Tab. Title Page
1
1 H NMR of compound 1 30
1
2 H-NMR of compound 2 31
1
1
1
5 H-NMR of compound 5 46
1
1
1
1
1
1
12 Contact toxicity of the isolated materials, toward the larvae 65
of the confused flour beetle T. Confusum
13 Fumigant toxicity of C. longa oil to adults and larvae of T. 66
confusum.
14 Effect of 24-h exposure to C. longa L. oil impregnated filter 67
papers on oviposition by T. confusem
List of Schemes
No. Title Page
1 Processing of Cynanchum acutum L. 28
2 Proposed fragmentation pattern of compound 5 36

6 proposed fragmentation pattern of compound 10 42

10 Processing of Curcuma longa rhizomes. 48


List of Figures
Fig. Title Page
1 Cynanchum acutum L. 2
2 Curcuma longa L. rhizomes 13
3 IR spectrum of compound 1 95
1
4 H NMR spectrum of compound 1 96
1
1
7 H NMR spectrum of compounds 3, 4 104
9 Mass spectrum of compound 5 109
1
10 H NMR spectra of compound 5 110
1
13 UV spectrum of compound 6 118
14 UV spectrum of compound 6 after the addition of EtONa 119
1
24 H NMR spectrum of unidentified flavonoidal compound 123
1
1
1
1
1
33 LC50 values of Curcuma oil 63
34 LC50 values of 1-dehydrogingerdion 64
35 LC50 values of Curcuma oil mixed with 1-dehydrogingerdione 64
36 Percentage of egg hatch and survival of larvae of Tribolium 67
castaneum exposed to Curcuma longa oil
37 Percentage of adult emergency of T. confusum from eggs 68
exposed to C. longa oil.
SUMMARY
SUMMARY
Isolation and Structure Elucidation of Natural
Products from Some Medicinally Important Plant
Species
The work described in this thesis has been undertaken with the aim of
searching of naturally occurring substances from some available plants in Egypt
as a source of drugs.
Extracts of two selected plant specie were studied, these are Cynanchum
acutum L. (Asclepiadaceae) and Curcuma longa L. (Zingiberaceae).
Through this investigation, useful information was obtained from the
application of various chromatographic methods, such as CC and PTLC, as well
as spectroscopic methods, such as MS, GC/MS, 1H NMR, UV and IR.
Processing of Cynanchum acutum L. revealed the isolation and structure
elucidation of fifteen compounds, which can be classified into two triterpenes
(3-O-acetyl lupane 1 and lupeol 2), two sterols (β-sitisterol 3 and stigmasterol
4), two coumarins (scoparone 5 and scopoletin 6), two shikimats (isovanillin 7
and syringic aldehyde 8), three monoterpens 9-11 and four fatty acids 12-15.
This is the first report indicating the presence of coumarins from the genus
Cynanchum.
Processing of Curcuma longa L. revealed the isolation and structure
elucidation of seven compounds which could be classified into three known
sesquiterpens, (ar-turmerone 16, α-turmerone 17 and caryophyllene oxide 18),
gingerol (1-dehydrogingerdione 19), three diaryl heptanoids (curcuminoids;
curcumin I 20, curcumin II 21 and Curcumin III 22). This is the first report on
isolation of gingerols (compound 19) from Curcuma longa.
I
SUMMARY
The petroleum ether and chloroform extracts of Curcuma longa are

studied for insecticidal and antifeedant activities against the confused flour
beetles, Tribolium confusum, where the petroleum ether extract was proved to be
active while the chloroform fraction was inactive.
The bioassay results showed that both 1-dehydrogingerdione 19 and
curcuma oily fraction which were isolated from the pet. ether fraction of
Curcuma longa were highly effective insecticidal against the confused flour
beetles, Tribolium confusum.
The experimental part describes the methods of plant processing,
extraction, isolation and insecticidal activity assessment.
29 29
20 20
30 20 30 20
27 19 27 19
12 22 12 22
18
25 11
18
25 11 13 13
1
17 28 1
17 28
14 14
2 9 16 2 9 16
10 8 15 10 8 15
O 26 26
4
O 5
6
7
1 HO 3 4 5
6
7
2
H3C 23 24 23
29 24 29
28 28
21 22 22
20 24 27 21 24 27
20
19 19
25 25
12 23 12 23
17 17
18 11 26 18 11 26
13 13
1 16 1 16
2 9 14 2 9
10 15 10 14 15
8 8
HO 3 5
6
7 HO 3 5 7
4 6
4
3 4
II
SUMMARY
O OH
HO O O
5 4 5 4
O 6 O
3 6
3
O 7 O 2 O 7 2
8 1
HO 8
O O
1
CHO H O
5 6 7 8
H OH
O O
O HO O
O
9 10 11
H OH
HO
O 12 O O 13
O O HO
14 15
12
H H
3 O
5 4
2
8 3'
O 7 1 O
2'
10 9 11
H
16 17 18
III
SUMMARY
H
O O
2' 10 4 2
MeO 3' 1'
8 6
9 7 5 3 1
HO 4' 6'
19
5'
H
O O
6 4 4' 6'
MeO 7 5 5' 7' OMe
2 2'
3 1 3'
H
HO 8 10 20 10' 8' OH
9 9'
H
O O
6 4 4'
6'
MeO 7 5 5' 7'
2 2'
3 1 3'
H
HO 8 10 21 10' 8' OH
9 9'
H
O O
6 4 4' 6'
7 5 5' 7'
2 2'
3 1 3'
H
HO 8 10 22 10' 8' OH
9 9'
IV
INTRODUCTION
INTRODUCTION
1.1. General
The use of medicinal plants for the treatment of many diseases is
associated with folk medicine in different parts of the world. Natural products
from some plants, fungi, bacteria and other organisms continue to be used in
pharmaceutical preparations either as pure compounds or as extracts. There is
a great variety of compounds that can be extracted and characterized from
plants. One good example is the harmaline; one of the indole alkaloids found
in Peganum harmala (Zygophyllaceae) used in the treatment of dermatosis.
Another substance that can be found in plants is the morphine from the opium
poppy, which has highly analgesic action and is still used. Its molecule is used
as a model for design to reach new drugs. The isolation of artemisinin showed
the real importance to investigate plants that can be sources of new
compounds with clinical activities. Plants produce a large number of
substances, which can provide a wide spectrum of biological properties.1 In
this thesis, we have select two plant species to be studied for their content of
natural products, these are Cynanchum acutum L. and Curcuma longa L.
1.2. Cynanchum acutum L.

1.2.1. Botanical aspect, description and distribution in Egypt.
Latin name : Cynanchum acutum L.
Arabic name: Modeid or Libbein.
Family : Asclepiadaceae.
1
INTRODUCTION
Cynanchum acutm L. is a wildly growing twining perennial herb, very

commonly distributed all over Egypt including Nile valley, Delta, El-Faiyum
as well as the Mediterranean coastal strip from El-sollum to Rafah and in the
oases of the Libyan desert, the plant flowers during September and October.2
Glabrous or glabrescent perennial; stems to 2.5 m, twining; leaves 3-10 ×
1.5-8 cm, cordate, the apex acute, the base broadly auriculate; petiole 1-3.5
cm; flowers in axillary cymes; peduncle 1-5 cm; calyx 1-2 mm, hairy the teeth
triangular; corolla 0.8-1.2 cm broad, pinkish-white, glabrous; 4-5 mm, linear,
acute; corona 5-lobed, the lobes caudate; follicles 6-12(-18) x 0.6-1 cm; the
apex attenuate, glabrous.3
Fig. 1: Cynanchum acutum L.
2
INTRODUCTION
1.2.2. Medicinal uses of some Cynanchum species.

The family Asclepiadaceae comprises many medicinal plants with a
wide rang of therapeutic activities. The word "Asclepias" means the god of
healing as the ancient Greeks named according to Koieke and his group4.
Cynanchum arnottianum has been used in India and tropical America
as insecticide and parasiticide.5 Cynanchum atratum has been used in China
as antifebrile and diuretic.6 Cynanchum paniculatum dried whole plant has
been used in China as anodyne and for the therapy of chronic tracheitis.7
Cynanchum wilfordi is used in Korea as a substitute for the tonic crude drug
"Ka-Shu-Uh"8. Cynanchum glaucescens dried roots have been used as
antitussive and expectorant.9 Cynanchum otophyllum has been used as a
traditional Chinese medicine for treatment of epilepsy and chronic hepatitis.10
1.2.3. Medicinal and Biological activities of Cynanchum acutum L.
Fawzy and his team11 reported antidiabetic and antioxidant activities
of major flavonoids of Cynanchum acutum L. growing in Egypt. Abou Zeid
and his group12 reported insecticidal and mollusicidal activities of the aerial
parts of Cynanchum acutum L. growing in Egypt. El-Lithi and his workers13
reported some pharmacological and toxicological activities of Cynanchum
acutum L.
1.2.3. Phytochemical constituents of Cynanchum genus.
The phytochemical investigation of different species has led to the
isolation and characterization of different classes of natural products including
steroidal glycoside, triterpenes, phenolic compounds and alkaloids.
1.2.4. Steroidal glycosides
Steroidal glycosides and particularly pregnane series could be
considered as the major phytoconstituents, which had been reported from the
different species of Cynanchum. Mitsuhashi and his group14 reported the
3
INTRODUCTION
isolation of steroids hirundigenin 1 and anhydrohirundigenin 2 from

Cynanchum grandifolium.
CH3 CH3
O O
O O
O O
OH
HO HO
1 2
Warashina and his group15 succeeded in the isolation and identification
of ten new pregnane glycosides 4-12 and 14 which had sarcostin or
deacylmetaplexigenin as the aglycone moiety from the aerial parts of
Cynanchum caudatum.
On the other hand, Warashina and his group16 succeeded in
identification of steroidal glycosides from the roots of Cynanchum caudatum.
These glycosides contain cynanchogenin 15, caudatin 16 and gagaminin 17 as
the aglycone moiety and 2,6-dideoxy-3-O-methylhexopyranose and
glucopyranose as the sugars component.
HO O
OH OH
OH OH
OH OH
OH OH
RO RO
R = H = Sarcostin R = H = Deacylmetaplexigenin
3
4
5
6
7
8
9
10
11
12
13
14
4
INTRODUCTION
O
O O N
O
O
O O
O O
H O
OH
OH OH
OH
OH
H OH
H OH
HO H OH
HO
HO
15 16 17
Yeo and his workers17 reported the isolation of three steroidal

glycosides named cynascyrosides 18-20 from the roots of Cynanchum
ascyrifolium.
OCH3 H3C
O H3C O O
R= 18 HO
H3CO O
O
O HO
OCH3
H3C OCH3 H3C O
OH OCH3 O O
H3C O
R= 19 O O O
HO HO
O HO OH
H3CO
OCH3 H3C
OH H3C O H3C O
RO O O
R= 20 HO HO
O
HO OH
Zhu and his team18 identified two C21-steroidal glycosides named

stauntosides 21, 22 from Cynanchum stauntoi.
21 R =
H HO H
H3C
O O CH3 H3CO O O O
O O O
HO H3C O
O OCH3 H3C H3CO OH
H3C H
H
CH3 H H
22 R=
O
OCH3 H
O HO H3C
HO CH3 O
RO O O
O
H3C HO
O
H H
On the other hand, Fu and his group19 reported successively a C21-

steroidal glycoside 23 determined as glaucogenin-C 3-O-α-L-
cymaropyranosyl-(1→4)-β-D-digitoxopyranosyl-(1→4)-β-D-
canaropyranoside from Cynanchum stauntoni.
5
INTRODUCTION
O
O
O H
CH3
O
OH
CH3
O
23
OH O
CH3 OH
OCH3
Zhan and his team20 reported the isolation of two cytotoxic C21-
steroidal glycosides named as auriculosides 24, 25 from the root of
Cynanchum auriculatum.
OH 24 R =
O O H 3C H3 C
HO H3 C O O
OH O O O
HO O O
O H3CO H3CO
CH3 OCH3
O OCH3
O
OH
OH OH 25 R=
O O H3C H 3C
OH HO H 3C H 3C O
O
OH O O O
RO HO O O
H3CO OH OCH3
H3CO
Kanchanapoom and his team21 reported the isolation of 8, 14-seco-

pregnane and pregnane glycoside named cynaphyllosides 26-35 from
Cynanchum aphyllum. The aglycone moieties are cynaphyllogenin 36 and 37.
6
INTRODUCTION
O
R
O
O 26 : H
H
27 :cym
36 O 28 :cym4- Glc
H O 29 :cym4 cym4 -Glc
RO 30 :cym4 cym4 The
OH
31 :cym4-Dig4 The
O
O 32 :cym4-cym4-The4 Glc
O 33 :cym4-Dig4-The4-Glc
O
37 OH 34 :cym4-Dig4-The4-Glc4-Glc
O 35 :D-Dig4-D-cym4-L-cym
H O
RO
OH
H3C OH
HO H3C H3C
HO HO HO HO
OH HO HO OH
H HO
H3CO H HO OH
OCH3 H H OH
OH
OH
OH
cym. Dig. The. Glc.
Li and his group22 succeeded in the isolation of C21-steroidal

glycosides named chekiangensosides 38-43 from the roots of Cynanchum
chekiangense.
O O
O
O
HO
HO O
O
RO RO
38 : R = S1 41 : R = S1
39 : R = S2 42 : R = S2
40 : R = H 43 : R = H
OH Me
O O
Me OMe
O Me
HO O
S1 = HO
O O O
OH OMe
OMe
Me Me
O O O
OH OMe O
Me
O O
O
S2 = HO
OH OMe
OH
HO
7
INTRODUCTION
Dou and his co-workers23 reported a C21-steroidal glycoside,

neocynapanogenin 3-O-β-D-thevetoside 44 as well as the aglycone
neocynapanogenin 45 from Cynanchum paniculatum.
O
D- thevetose
O
44 R = Me
O
O HO
O MeO
OH
45 R= H
RO
Wang and his team24 reported the isolation of three C21-steroidal

glycosides named inamosides 46-47 from the roots of Cynanchum inamoenum.
O
O 46 R = S1
O 47 R = S2
O 48 R = S3
RO
OH H3C CH3 CH3
O O O O
S1 HO O
HO O
OH OCH3
OCH3
OH
OH
H3C H3C
OCH3
O O
O O
O O
S2 HO
O H3C
O O
HO HO
OH OCH3
OH HO OH
OH
H3C H3C
OCH3 O O
O
S3 O O H 3C
O
O
HO
HO O O
HO OH
OH OH
HO OH
8
INTRODUCTION
1.2.3.2 Triterpenes.
Konda and his group25 reported the isolation of hancokinol 49 from
Cynanchum hancokianum.
30
20
19 21
29
27 18 22
12
11 13 17
25
28
1 14 16
9
2 8 15
A
26
3 4 5 7
HO
6
24 23
49
1.2.3.3 Alkaloids.
An and his group 26 reported the isolation of an alkaloid identified as
2,3-dimethoxy-6-[3-oxobutyl]-7,9,10,11,11a,12-
hexahydrobenzo[f]pyrrolo[1,2-b] isoquinoline 50 from Cynanchum komarovii.
OMe
MeO
50
On the other hand, Staerk and his team27 reported the isolation of
the phenanthroindolizidine alkaloids (-)-13aα-antofine 51, (-)-10β,13aα-
antofine N-oxide 52, (-)-14β-hydroxyl-10β,13aα-antofine N-oxide 53 from
Cynanchum pumilum.
9
INTRODUCTION
OMe
OMe
MeO
MeO R
N
N
MeO
MeO
52 R = H
51
53 R = OH
1.2.3.4 Shikimates and phenolic compounds.

Lou and his co-workers28 reported the isolation of two sinapic acid
esters 6-O-[E]-sinapoyl-(α- and β)-D-glucopyranoside 54, β-D-
frucofuranosyl-α-D-(6-O-[E]-sinapoylglucopyranoside 55 and a phenolic
glycoside 2-acetylphenol-1-β-D-gluco-pyranosyl(1→6)-β-D-xylpyranoside 56
from Cynanchum hancockianum.
OMe
OMe
O
O OH
OH
O
O OMe
OMe
CH2
CH2
O
O
HO
HO HO
OH OH
HO OH
OH OH
O
54 55 OH
OH
Me
O
O H2C
56 HO
HO
O O
OH HO O
HO
OH
Huang and his group29 reported the isolation of acetophenones 4,3'-

diacetyl-2,3,2',6'-tetrahydroxybipheny 57 and 4,3'-diacetyl-2,3,2',6'-
tetrahydroxybiphenyl dimers 58 from roots of Cynanchum taiwanianum.
10
INTRODUCTION
O OH
OH
Me OH O
4'
5'
Me
Me
6' 7'
8'
57 HO
3'
HO 1' 2' O
O
1 OH
2 6
OH
OH 13
Me 7"
O O 3
8" 3" 8 9 5
2"
O 4
7 14
4"
1" 10 15 16 Me
12 17
5" 6" 11
OH
58
1.2.5. Chemical constituents of Cynanchum acutum L.

Literature survey of investigation of Cynanchum acutum L. reveled
the isolation of several secondary metabolites. Halim and his group30 reported
the isolation of lupeol long-chain fatty acid esters and other lipid constituents
from Cynanchum acutum L. The petroleum ether extract of the aerial parts of
Cynanchum acutum L. afforded the triterpenoidal compounds lupeol 63, lupyl
acetate 64, and α-amyrin 65 in addition to the steroidal compounds β-
sitosterol 66 and stigmasterol 67.
O
HO O HO
Me
63 64 65
HO
HO
66 67
11
INTRODUCTION
El Sayed and his team31 reported the isolation of scrostin 68 from the
diethyl ether extract in addition to the isolation and identification of querctin
69 and querctin-3-O- β-D-galactoside 70 from the ethyl acetate extract.
On the other hand, Heneidak and her team32 reported the isolation and
identification of four flavonoid glycosides; quercetin di-O-hexoside 71,
quercetin-3-O-rhamnosyl-xyloside 72, and quercetin 3-O-xyloside 73 using
HPLC.
OH OH
H3C
OH
OH OH
CHOH
OH HO O HO O
OH
OH OR
OH OH O OH O
HO
69 R = di-O-hexoside 70
68
= O-rhamnosyl-xyloside 71
= galactoside 72
= xyloside 73
1.3. Curcuma longa L.

1.3.1. Botanical aspect, description and distribution.
Latin name: Curcuma longa L.
Common name: Turmeric (in India).
Arabic name: korkome.
Family: Zingiberaceae.
Curcuma longa L. is a perennial herb with simple and large leaves. Its
tubers, rhizomes and oil have great importance. Its rhizomes are oblong, ovate
with characteristic odor and slightly pungent bitter taste. Root scars and
annulations are present on the surface of the rhizome. The fracture is horny
and internal surface is orange in color.33
12
INTRODUCTION
Fig. 2 Curcuma longa L. rhizomes
1.3.2. History and Medicinal Uses.

Turmeric (Curcuma longa L.) belongs to family Zingiberaceae along
with the other noteworthy members like ginger, cardamom and galangal. It
belongs to the genus Curcuma that consists of hundreds of species of plants
that possesses rhizomes.
Turmeric is of special importance to humans with the discovery that
its rhizome powder, when added to various food preparations, preserves their
freshness and imparts a characteristic flavor. Turmeric, which belongs to a
group of aromatic spices, had been originally used as a food additive in
curries to improve the storage condition, palatability and preservation of food.
Turmeric is grown in warm, rainy regions of the world such as China, India,
Indonesia, Jamaica and Peru.34
In India, it is popularly known as Haldi (Hindi). In Malaysia,
Indonesia and India, turmeric has been well studied due to its economic
importance.
Its rhizomes are oblong, ovate, pyriform and often short-branched and
these are a household remedy in Nepal.35
In Ayurveda, turmeric has been used internally as a stomachic, tonic
and blood purifier and externally in the prevention and treatment of skin
diseases.36
13
INTRODUCTION
Traditional Indian medicine claims the use of its powder against

biliary disorders, anorexia, coryza, cough, diabetic wounds, hepatic disorder,
rheumatism and sinusitis.37
Modern interest in turmeric began in 1970’s when researchers found
evidence suggesting that the herb may possess anti-inflammatory properties.
Turmeric roots are known to be antiseptic and aromatic. Normally,
curcuminoids were extracted by solvent extraction.38
The significance of turmeric in medicine has changed considerably,
since the discovery of the antioxidant properties of naturally occurring
phenolic compounds in turmeric. The chemical constituents of turmeric have
been studied in relation to the mechanisms of oxygen stress-related processes,
including aging. Products isolated from Curcuma longa show a strong
antioxidant action when tested on the model systems like oxidation of linoleic
acid in air 39, 40, 41 and in vitro peroxidation of brain lipids.42
1.3.3. Biological activities of Curcuma longa L.

Srimal and Dhawan43 reported the pharmacological action of curcumin
e.g. the compound was effective in acute as well as chronic model of
inflammation. The potency of this drug is approximately equal to
phenylbutazone in the carrageenin-induced edema test, but it is only half as
active in the chronic experiments. It was observed that curcumin was less
toxic than the reference drug (no mortality up to a dose of 2 g/kg).
 Antioxidant activity.
Masuda and his group44, 45
reported the antioxidant mechanism of
curcumin, in the presence of ethyl linoleate as one of the polyunsaturated
lipids. During the antioxidation process, curcumin reacted with four types of
linoleate peroxyl radicals. Six reaction products were observed in the reaction
and these have novel tricyclic structures, including a peroxyl linkage. On the
basis of the formation pathway for their chemical structures, an antioxidant
14
INTRODUCTION
mechanism of curcumin in polyunsaturated lipids was proposed, which

consisted of an oxidative coupling reaction at the 3'-position of the curcumin
with the lipid and a subsequent intramolecular Diels–Alder reaction. Further,
a relatively high concentration of curcumin gave three dimers as radical
termination products in addition to the coupling products with curcumin and
the lipid hydroperoxide. The structural analysis of these dimers and
quantitative analysis of their production rates revealed that radical termination
mainly occurred at the 2-position of curcumin. The contribution of the
pathway for production of these dimers to the antioxidant mechanism of
curcumin was estimated from the concentration-dependent data of the
antioxidant activity and formation rates of these termination products. The A–
A termination (dimer formation) was estimated to contribute at least about
40% of the entire antioxidant process against ethyl linoleate oxidation.
Sun and his team 46, 47 used the fractions of turmeric oil to determine
their protective effect against the mutagenicity of sodium azide by means of
the Ames test. All the fractions and turmeric oil exhibited antimutagenicity
markedly. The antioxidant effects of turmeric oil and its fractions may
provide an explanation for their antimutagenic action. Bond dissociation
enthalpies (BDEs) for the curcumin-related compounds have been calculated
using density functional theory methods.
It was reported that the antioxidant mechanism of curcumin was an H-
atom abstraction from the phenolic group, not from the central CH2 group in
the heptadienone link. Curcumin, methylcurcumin, and half-curcumin had
similar O-H BDEs, indicating that the two phenolic groups in curcumin were
independent of each other.
Gayathri and his group48 reported that the loss (27–71%) of β-
carotene in vegetables was observed during the two domestic methods of
cooking commonly used, namely, pressure cooking and open pan boiling.
15
INTRODUCTION
However, presence of antioxidant spice turmeric generally improved the

retention of β-carotene.
Daniel and his workers49 showed that curcumin could chelate toxic
metals (e.g. cadmium and lead), and potentially reduces their neurotoxicity
and tissue damage in rat brain homogenate.
 Anti-protozoal activity.
Arau´jo and his team50,51 reported the anti-protozoa activity of
curcumin and some semi-synthetic derivatives against tripanosomatids in
promastigotes (extracellular) and amastigotes (intracellular) forms of
Leishmania amazonensis. It was reported that curcumin has an excellent
activity (LD50=24µM or 9 mg/ml) and the semi-synthetic derivative,
methylcurcumin (a non-phenolic curcuminoid), has the best action with a
LD50<5 µg/ml and LD90=35µM against promastigotes forms. This derivative
was tested in vivo in mice and showed good activity with 65.5% of inhibition
of the lesion size of the footpad of the animals, when compared with the
group inoculated with the parasites alone. Another interesting point
mentioned is that they did not observe any inflammatory reaction in the area
where the drugs were injected, perhaps because curcuminoids are potent
inhibitors of inflammation.
 Antimicrobial activity.
Chopra and his team52 reported that the Curcuma oil was tested
against cultures of Staphylococcus albus, Staphylococcus aureus and Bacillus
typhosus and the results showed inhibition of the growth of S. albus and S.
aureus at different concentrations.
Bhavanishankar and Srinivasamurthy53 investigated the activity of
turmeric fractions against some intestinal bacteria in vitro. Total inhibition of
growth of lactobacilli in the presence of whole turmeric was reported (4.5–90
µl/100 ml). The alcoholic extract was also effective (10–200 mg/ml), but the
16
INTRODUCTION
inhibition was not equal as the whole turmeric. Curcumin (2.5–50 mg/ml) was
inhibited only S. aureus.
Negi and his team54 reported the antibacterial activity of turmeric oil.
The oil was extracted from the spent turmeric oleoresin and it was separated
into three fractions using column chromatography. These fractions were
tested for antibacterial activity by pour plate method against Bacillus cereus,
Bacillus coagulans, Bacillus subtilis, S. aureus, Escherichia coli, and
Pseudomonas aeruginosa. Fraction eluted with 5% ethyl acetate in hexane
was found to be the most active fraction. The turmeric oil, fractions were
analyzed by GC and GC/MS. Aromatic-turmerone, turmerone, and curlone
were found to be the major compounds present in these fractions along with
other oxygenated compounds.
Jayaprakasha and his group55 reported the antifungal activity of
turmeric oil, which was also isolated from mother liquor after isolation of
curcumin. The turmeric oil was fractionated using fractional distillation under
vacuum to get two fractions. These fractions were tested for antifungal
activity against Aspergillus flavus, A. parasiticus, Fusarium moniliforme and
Penicillium digitatum by spore germination method. Fraction obtained at
110–1200C under vacuum was found to be more active. The chemical
constituents of turmeric oil, fractions were determined by GC and identified
by GC/MS. Aromatic turmerone; turmerone and curlone were major
compounds present in the active fraction along with other oxygenated
compounds.
 Antivenom activity.
Ferreria and his team56 reported the activity of turmeric and its
constituents against snake venom. The fraction consisting of aromatic-
turmerone, isolated from Curcuma longa neutralized both the hemorrhagic
activity and lethal effect of venom in mice. Aromatic-turmerone was capable
of abolishing the hemorrhagic activity of Bothrops venom and about 70% of
17
INTRODUCTION
the lethal effect of Crotalus venom. Immunological studies demonstrated that

aromatic-turmerone inhibited the proliferation and the natural killer activity of
human lymphocytes.
 Anti-HIV.
Sui and his team57 reported inhibition of HIV-1 and HIV-2 proteases
by curcumin and curcumin boron complexes. Simple modification of the
curcumin structure rise the IC50 value complexes of the central dihydroxyl
groups of curcumin with boron lower the IC50 to a value as low as 6 and 55
µm, respectively, for HIV-1 and HIV-2, whereas curcumin showed 100 and
250 µm.
Mazumder and his co-workers58 demonstrated that curcumin has an
antiviral activity, being a HIV-1 integrase inhibitor (IC50=40µM) and
suggested that curcumin analogs could be developed as anti-AID’s drugs.
Data showed that curcumin inhibited the replication of HIV-1 integrase
protein.
 Anti-tumour activity.
Huang and his team59 studied the effect of curcumin, chlorogenic acid,
caffeic acid and ferulic acid on tumour promotion in mouse skin by 12-O-
tetradecanoylphorbol-13-acetate and observed that all these compounds
inhibit the epidermal ornithine decarboxylase and epidermal DNA synthesis,
curcumin being the most effective.
Limtrakul and his group60 showed an inhibitory effect of curcumin on
mouse skin carcinogenesis initiated by 7,12-dimethylbenz (a) anthracene and
promoted by TPA. Thus, curcumin administration decreased both the number
of tumors per mouse and tumour volume.
Ozaki and his team61 studied the action of curcumin on rabbit
osteoclast apoptosis and demonstrated that curcumin drastically inhibits bone
resorption and stimulation of apoptosis in the cells. Since, cancer and bone
18
INTRODUCTION
inflammation are diseases that increase bone resorption, he suggests that

curcumin may be useful in the therapy of these diseases.
Inano and his team62 studied the anti-cancer action of curcumin in a
standard model of radiation-induced tumour in rat mammary gland. He
suggests that curcumin has the potential to be an effective agent for
chemoprevention of radiation-induced initiation stage of mammary
tumourogenesis.
 Anti-inflammatory activity.
Arora and his group63 investigated the anti-inflammatory activity of
different fractions of the rhizomes of turmeric in animals. It was reported that
the extracts reduced the granuloma growth and no toxic effects were observed.
Chandra and Gupta64 demonstrated the anti-inflammatory and anti-
arthritic actions of volatile oil of Curcuma longa L.
Ghatak and Basu65 showed the action of sodium curcuminate as an
anti-inflammatory agent, being better than curcumin and hydrocortisone
acetate, in experimental inflammation induced by carrageenin and formalin in
albino rats (ED50=144 µg/kg).
Ammon and Wahl66 reported that the Curcuma extracts showed a high
anti-inflammatory effect after parenteral application in standard animal
models.
 Insecticidal activity.
A number of anti-insect properties of turmeric have been documented
in the literature. Su and his co-workers67 reported that the insect repellent
components in turmeric are the turmerones and ar-turmerone. The petroleum
ether extract of Curcuma longa rhizome has been reported as repellent to
Tribolium castaneum and insecticidal to Plutella xylostella68 on the other
hand the hexane extract of rhizome of Curcuma longa reduced progeny
production in T. castaneum at 200 ppm concentration.69 The aqueous extract
of Curcuma longa L. rhizome acted as repellent against Callosobruchus
19
INTRODUCTION
chinensis70, insecticidal to Myzus persicae71 and reduced larval populations of

Anopheles stephensi Liston72. The acetone extract of Curcuma longa L.
rhizome acted as repellent to T. castaneum.73
1.3.4. Phytochemical constituents of Curcuma longa L.

The constituents of Curcuma longa L. involving, the constituents
of the volatile oil and the constituents of curcuma resin.
1.3.4.1. Constituents of volatile oil of Curcuma longa L.
The aroma of turmeric is due to its volatile oil, while the phenolic
compounds and its analogues account for its bright yellow colour. Due to its
lower commercial importance, the chemistry of turmeric oil has not received
much attention earlier.
Kelkar and Sanjeev Rao74 reported that steam distilled volatile oil is
predominantly a mixture of sesquiterpene ketones and alcohols.
Malingre75 reported p-cymene 74, β-sesquiphellandrene 75, turmerone
76, and aromatic-turmerone 77 from Curcuma longa L.
O
O
74 75 76 77
Chen and his group76 compared the composition of the volatile oils of
rhizome and tuber of Curcuma longa of Chinese origin. Turmerone (24%),
aromatic-turmerone (8.4%) and curdione 78 (11.58%) are found to be the
major compounds in both the oils. However, aromatic-curcumene 79 was
found in rhizome oil to the extent of 12.2%, but it was not reported in tuber
oil.
20
INTRODUCTION
Kiso and his team77 examined an aqueous ethanolic extract of

Curcuma longa rhizomes. A new oxygenated sesquiterpene, curlone 80 was
isolated. Structural elucidation of curlone was achieved by dehydrogenation
to ar-turmerone as well as by mass and NMR spectral studies.
Imai and his group78 reported the two sesquiterpene keto-alcohols
turmeronol-A 81 and turmeronol-B 82 from the dried rhizomes of Curcuma
longa L.
O O
78 79 80
OH
HO
H3C O
H3C O
81 82
Ohshiro and his team79 examined sesquiterpenoidal constituents from

methanolic extract of Curcuma longa L. Five sesquiterpenes germacron-13-al
83, 4-hydroxybisabola-2,10-dien-9-one 84, 4-methoxy-5-hydroxybisabola-
2,10-dien-9-one 85, 2,5-dihydroxybisabola-3,10-diene 86 and
procurcumadiol 87 were isolated and identified by NMR spectroscopy. It was
concluded that the high content of bisabolene type sesquiterpenes is
characteristic for Curcuma longa L. compared with other Curcuma species.
OH
OH
O
O
CHO
83 84 85
21
INTRODUCTION
OH
OH
OH
OH
86 87
Uehara and his group80 analyzed hexane extracts of the rhizomes of a
number of cultivars of turmeric using GC/MS. It was reported that, the
percentages of major components vary e.g. aromatic-turmerone (2.6–70.3), α-
turmerone (trace—46.2%) and zingiberene 88 (trace—36.8%).
Hisashige and his group81 reported the presence of (+)-aromatic-
curcumene and (-)-β-bisabolene 89 in turmeric oil.
Zhu and his team82 investigated the rhizome oil of Curcuma longa L.
of Chinese origin. The oil was reported to contain seventeen chemical
constituents of which turmerone (24%), aromatic-turmerone (18%) and
germacrone 90 (11%) as the major compounds.
H
H
O
88 89 90
Sharma and his group83 analyzed oil produced from 5-10 month old
Curcuma longa L. rhizomes that were grown in Bhutan using GC and
GC/MS and the major compounds were found to be aromatic-turmerone
(16.7–25.7%), α-turmerone 91 (30.1–32.0%) and β-turmerone 92 (14.7–
18.4%).
H H
O O
91 92
22
INTRODUCTION
Garg and his geoup84 reported the volatile compounds from leaves of
turmeric Curcuma longa L. Twenty chemical constituents were identified
using GC and GC/MS, which comprised 72% of oil contents. Predominant
chemical groups included monoterpene hydrocarbons, oxygenated
monoterpenes, sesquiterpene hydrocarbons and oxygenated sesquiterpenes
(57, 10, 3.3 and 2.1%, respectively). Of the individual volatile compounds
identified, p-cymene, 1,8-cineole 93 and β-pinene 94 were the major
constituents, accounting for 25.4, 18, 7.4 and 6.3% of the oil, respectively.
93 94
1.3.4.2 Curcuminoids.
The coloring principle of turmeric was isolated in the 19th century and
was named curcumin. Curcuminoids refer to a group of phenolic compounds
present in turmeric, which are chemically related to its principal ingredient
curcumin. Three curcuminoids were isolated from turmeric, curcumin 95
demethoxycurcumin 96 and bisdemethoxycurcumin 97.
HO OH
MeO OMe
O O
HO 95 OH
OMe
O O
HO 96 OH
O O
97
23
INTRODUCTION
All three impart the yellow pigmentation to the Curcuma longa L.

plant and particularly to its rhizomes. Although the chemical structure of
curcumin was determined in the 1970’s and 1980’s, recently the potential uses
of curcuminoids in medicine have been studied extensively.
Majeed and his co-workers 85 proved that the structure of curcumin is
diferuloylmethane by the degradative work.
Kiuchi and his group86 reported the isolation of a curcuminoid,
cyclocurcumin 98 and it was isolated from the nematocidally active fraction
of turmeric along with other known curcuminoids.
HO OH
MeO OMe
98
Park and Kim87 reported the isolation of two curcumonoids

4''-(3''methoxy-4'''hydroxyphenyl)-2''-oxo-3''-enebutanyl-3-(3'-methoxy-4'
hydroxyphenyl) propenoate (calebin-A) 99 and 1,7-bis(4-hydroxy-3-
methoxyphenyl)-1,4,6-heptatrien-3-one 100 and seven known compounds,
curcumin, demethoxycurcumin, bisdemethoxy-curcumin, 1-hydroxy-1,7-
bis(4-hydroxy-3-methoxyphenyl)-6-hepten-3,5-dione 101, 1,7-bis(4
hydroxyphenyl)-1,4,6-hepten-3,5-dione 102, 1,7-bis(4-hydroxyphenyl)-1,4,6-
heptatrien-3-one 103 and 1,5-bis(4-hydroxy-3-methoxyphenyl)-1,4-pentadien-
3-one 104 from Curcuma longa L.
OMe
OH
99 O
HO
OMe
O
H3CO OCH3
100
HO OH
24
INTRODUCTION
OH
O O
H3CO OCH3
101
HO OH
O O
102
HO OH
O
H3CO
103
HO OH
O
H3CO OCH3
104
HO OH
Yao and his group88 report the isolation of a diarylheptanoid 105

identified as 1,5-epoxy-3-carbonyl-1,7-bis(4-hydroxylphenyl)-4,6 heptadiene.
HO OH
105
Numerous methods are available for isolating curcuminoids from

Curcuma longa L. Isolation of pure curcumin from plant material is time
consuming and pure curcumin sold on the market is therefore, a purified
extract containing a mixture of the three curcuminoids i.e. curcumin (75–
81%), demethoxycurcumin (15–19%) and bisdemethoxycurcumin (2.2–6.6%).
Except by the chromatographic routes, all other methods generally provide
several curcuminoids, with curcumin as the major constituent.
25
AIM of THE WORK
AIM of THE WORK
Plants have been one of the important sources of medicines even since
the dawn of human civilization. Recently, in spite of tremendous development
in the field of allopathy, plants still remain one of the major sources of drug in
the modern as well as traditional system of medicine throughout the world.
Over 60% of all pharmaceuticals are plant-based.
In this work we tried to reinvestigate the two medicinal plants

(Cynanchum acutum L.) and (Curcuma longa L.) hoping for isolation and
characterization some further bioactive natural products.
Our research plan included the following:
 Literatures survey for the two selected plant species.
 Collection and identification of the two plant species Cynanchum acutum
L. and Curcuma longa L.
 Fractionation and preparation of their.
 Isolation and purification of the natural organic plant constituents.
 Structure elucidation and characterization of the isolated products using
spectral methods.
 Laboratory bioassay for some isolated natural products whenever possible.
26
RESULTS AND DISCUSSION

As we mentioned before, our target concerns with the evaluation of
some natural extracts of some plants available in Egypt as a source for drugs.
The literature survey for the medicinal plants found in Egypt enabled us to
select two plant species which are available in the nature or the local markets.
These are Cynanchum acutum L. (Asclepiadaceae) and Curcuma longa L.
(Zingiberaceae) to be studied in details.
2.1. Processing of Cynanchum acutum aerial parts.
The powdered air dried aerial parts of Cynanchum cutum L. was processed
According to scheme 1.
2.1.1 Study of petroleum ether fraction.
The petroleum ether fraction (cf. scheme 1) was evaporated to dryness
where a dark green residue was obtained. The residue was saponified using
alcoholic sodium hydroxide. The unsaponifiable material was subjected to
column chromatograph using silica gel. Three fractions were obtained by the
solvent system hexane / EtOAc containing compound 1 at ratio of 49:1,
compound 2 at 24:1 and compounds 3 and 4 at 13:1.
2.1.2 Characterization of compound 1.
Thin- layer chromatographic study of compound 1 showed that it was
not homogenous and contaminated with other minor constituents. Therefore,
it was purified by preparative thin-layer chromatography, using silica gel and
the same solvent system of CC (pet. ether / EtOAc 49:1). Compound 1 was
obtained as white needles, m.p. 218-220 oC, Rf = 0.24 (silica gel, pet. ether /
chloroform 3:4). It gave a violet color upon spraying with p-anisaldehyde-
sulphoric acid reagent indicating it's steroidal or triterpenoidal nature.
27
Cynanchum acutum
plant material (aerial parts)
1- Extraction by Methanol
2- Filtration and evaporation to dryness
3- Defatting with cold Methanol
4- Filtration
Defatted extract ppt (Long chain fatty acids & esters)

Filtrate
1- Evaporation, residue
2- Solvent fractionation
with different polarities
Pet. ether CH2Cl2 Ethyl acetate fraction

fraction fraction
1- saponification, NaOH PTLC
2- Extraction with pet. ether GC/MS
10 Cpds EtoAc / MeOH
( 19 : 1 )
1- C C silica gel
2- pet.ether / EtoAc
Saponifiable Unsaponifiable 3- PTLC Flavonoidal compound
fraction fraction
C.C silica gel
pet. ether / EtOAc
PTLC
Fr.30 Fr.45
Compd.5 Compd.6
Fr-4 Fr-7 Fr-13

Compd.1 Compd.2 Comp.3, 4
Scheme1: Processing of Cynanchum acutum L.
28
The IR spectrum (Fig. 3) revealed the presence of absorption bands at

2985 cm-1 (CH-stretching), 1732 cm-1 (C=O group), 1252 cm-1 (C-O of
acetate group). The 1H NMR spectrum (table 1 and Fig. 4) revealed the
presences of six methyl group signals in the up field region showing that the
compound may be steroidal or triterpenoidal compound. The presence of two
olefinic protons as broad singlets at 4.55 and 4.67 ppm is characteristic for
pentacyclic lupane triterpene series (H-29, 29').89 A down field H-3 as double
of doublet at δ 4.46 ppm indicates the probable link to an ester group and this
was confirmed by the presence of acetoxyl group as singlet at 3.6 ppm. An
olefinic methyl group as a broad singlet at 1.67 ppm was consistent to the C-
30 methyl group. The H-19 appears as ddd at 2.38 ppm due to its allylic
position. The Me-23 and 24 appear as singlets at 0.94, 0.76 ppm, respectively.
The spectrum indicated the presence of four tertiary methyl groups (Me-25,
26, 27, and 28) as singlets at 0.83, 1.03, 0.96, 0.79 ppm, respectively. Based
on the previous spectral data, compound 1 which was isolated from
unsaponifiable part is 3-O-acetyl lupane which was isolated previously from
the same plant species by Halim and his team30.
29
20
30 20
27 19
12 22
18
25 11 13
1
17 28
14
2 9 16
10 8 15
O 3 26
4 5 7
O 6
H3C 23 1
24
29
Table 1: 1H NMR of compound 1

H atom δ value, ppm Integration, multiplicity
(J, Hz)
3 4.46 1H, dd, 5.53
19 2.38 1H, ddd, 10.6, 10.6, 5.3
23 0.94 3H, s
24 0.76 3H, s
25 0.83 3H, s
26 1.03 3H, s
27 0.96 3H, s
28 0.79 3H, s
29 4.55 1H, br s
29' 4.67 1H, br s
30 1.67 3H, br s
CH3CO 3.6 3H, s
2.1.3 Characterization of compound 2

Compound 2 was isolated as white needle crystals, m.p. 211-213 oC,
Rf = 0.24 from (silica gel, pet. ether-chloroform 3:4). It gave a violet color
upon spraying with p-anisaldehyde-sulphoric acid reagent indicating it's
steroidal or triterpenoidal nature. The IR spectrum (Fig 5) revealed the
presence of absorption bands at 3313 cm-1 (OH group), 2985 cm-1 (C-H
stretching). The 1H NMR spectrum of compound 2 (table 2, Fig.6) is identical
with the 1H NMR spectrum of compound 1 except the position of proton H-3.
H-3 appears as double of doublet at a chemical shift 3.2 ppm which indicating
that H-3 is attached to a free hydroxyl group characteristic for H-3 in sterols
and triterpens. Thus compound 2 is lupan-3-ol which was isolated previously
from the same plant species by Halim and his team30.
30
29
20
30 20
27 19
12 22
18
25 11 13
1
17 28
14
2 9 16
10 8 15
26
3 4 5 7
HO 6
23 2
24

(J, Hz)
3 4.46 1H, dd, 5.53
19 2.38 1H, ddd, 10.6, 10.6, 5.3
23 0.94 3H, s
24 0.76 3H, s
25 0.83 3H, s
26 1.03 3H, s
27 0.96 3H, s
28 0.79 3H, s
29 4.55 1H, br s
29' 4.67 1H, br s
30 1.67 3H, b s
31
2.1.4 Structure elucidation of compound 3

Compound 3 was obtained as white needles, m.p. 138-139 oC; it gave
a violet color upon spraying with p-anisaldehyde-sulphoric acid reagent
indicating it’s steroidal or triterpenoidal nature. The 1H NMR spectra of
compound 3 (table 3, Fig.7) revealed the presences of six methyl groups in
the up field region showing that the compound is aliphatic and may be
steroidal or triterpenoidal compound. The spectrum indicated the presence of
a multiplet at δ 3.52 ppm, which was assigned for H-3 indicating its steroidal
nature. An olefinic proton appeared as broad singlet at 5.33 ppm, which was
assigned for H-6 in ring B suggesting the presence of a Δ5 -3-hydroxy sterol.
The spectrum indicated the presence of two tertiary methyl protons signals at
0.68, 1.01 ppm, corresponding to (Me-18 and Me-19) respectively. The side
chain signals appeared at δ 0.92 (3H, d, J = 6.4 Hz, Me-21), 0.83 (3H, d, J =
6.8 Hz, Me-26), 0.81 (3H, d, J = 6.9 Hz, Me-27), 0.85 (3H, t, J = 7.8 Hz, Me-
29) suggesting that the sterol has a stigmast-5-en-3-ol skeleton. Based on all
these spectral data, compound 3 is β-sitosterol which was isolated previously
from same plant species by Halim and his co-workers30.
29
28
21 22 24 27
20
19 23 25
17
18 12 26
11 13
1 16
2 9 14
10 8 15
7
HO 3 5
4 6 3
32

(J, Hz)
3 3.52 1H, m, 1H
6 5.35 1H, m, 1H
Me-18 0.69 3H, S
Me-19 1.01 3H, S
Me-21 0.92 3H, d, 6.4
Me-26 0.83 3H, d, 6.8
Me-27 0.81 3H, d, 6.9
Me-29 0.85 3H, t, 7.8

Compound 4 was isolated as white crystals; m.p 170 oC also it gave a
violet color upon spraying with p-anisaldehyde-sulphoric acid reagent
indicating it's steroidal or triterpenoidal nature. The 1H NMR spectrum of
compound 4 (table 4, Fig.7) is identical with the spectrum of compound 3 in
addition to two olefinic protons which appeared as double of doublet at δ 5.1,
5.00 ppm, respectively, which were assigned for (H-22 and H-23). The
spectrum suggesting the presence of a Δ5, 22 -3-hydroxy sterol. The H-20 and
H-24 appeared as a multiplet at δ 2.24 and 2.00 ppm, respectively due to their
allylic position. Thus, all the previous data support that the compound 4 is
stigmast-5, 22-dien-3-ol which is known as stigmasterol which was isolated
previously from the same plant species by Halim and his group.30
33
29
28
21 22 24 27
20
19
23 25
17
18 12 26
11 13
1 16
2 9 14
10 8 15
7 4
HO 3 5
4 6
Table4: 1H NMR of compound 4

H atom δ, value, ppm integration, multiplicity
(J, Hz)
3 3.52 1H, m
6 5.35 1H, m
Me-18 0.69 3H, s
Me-19 1.01 3H, s
Me-21 0.92 3H, d, 6.4
22 5.00 1H, dd, 8, 14
23 5.21 1H, dd, 8, 14
Me-26 0.82 3H, d, 7
Me-27 0.83 3H, d, 7
Me-29 0.97 3H, t,7
34
2.2. Study of the methylene chloride extract.

The CH2Cl2 extract was separated over silica gel column, eluted with
petroleum ether and ethyl acetate, with increasing the polarities (c.f. scheme
1). About seven fractions were collected. Fraction 30 was further purified
using preparative layer chromatography using the solvent system petroleum
ether/ethyl acetate (silica gel, 11: 9, Rf = 0.39), where compound 5 was
obtained. Fraction 45 also was further purified using PTLC using the same
solvent system (silica gel, 3:2, Rf = 0.21) to give compound 6.
2.2.1. Structure elucidation of compound 5

Compound 5 was obtained as colorless needles material, m.p = 144 oC,
it gave a violet fluorescence in UV light. The IR spectrum (Fig. 8) showed a
carbonyl band at 1713 cm-1 (δ-lactone), also an ethylinic band at 1611 cm-1
corresponding to (CH=CH) group and 1565, 1514 cm-1 bands due to aromatic
benzene ring. The 1H NMR spectrum (table 5, Fig.10) showed two doublets
with coupling constant 9.6 Hz at δ 6.28 and 7.6 ppm, which are characteristic
for coumarins. Additionally two methoxy group singlets at 3.91, 3.94 ppm.
Other two aromatic protons singlets at 6.85 and 6.84 ppm were viewed. These
two aromatic protons singlets are explained by 6,7-disubstitution indicating
that the characterized compound is 6,7-dimethoxycoumarin (scoparone,5). Its
mass spectrum (Fig. 9) showed a base ion peak at m/z 206 (100%)
corresponding to C11H10O4 .The spectrum also showed an peak at m/z 191
(50%) due to [M-CH3]+ and other ion peaks at m/z 178 (30%) due to [M-
CO]+, 163 (40%) due to [M-CH3, CO] +, 135 (25) due to [M-CH3, 2CO]+.
Scoparone was isolated previously from Ethulia vernonioides by Schuster and
his group90.
35

H atom δ value, ppm integration, multiplicity
(J, Hz)
3 6.28 1H, d, J= 9.6 Hz
4 7.6 1H, d, J= 9.6 Hz
5 6.85 1H, s
8 6.84 1H, s
C-6-(O-Me) 3.91 3H, s
C-7-(O-Me) 3.94 3H, s
5 4
O
6
3
O 7 O 2 O
8 1
-CO -CH3
M+ m/z 206 (100)
+ 5
O O
O O O O
O
.+
m/z 178 m/z 191
-CO
-CO
m/z 150 -CO

m/z 135
[C9H10O2]+
[C8H7O2]+
O
O .+
m/z 163
Scheme 2: proposed fragmentation pattern of compound 5
36
2.2.2. Structure elucidation of compound 6

Compound 6 was obtained as yellow needles material, m.p. = 204 oC, it
gave a violet fluorescence in UV light. The IR spectrum (Fig.11) showed a
hydroxyl absorption band at 3408 cm-1 , a carbonyl band at 1713 cm-1 (δ-
lactone), also an ethylenic band at 1611 cm-1 corresponding to (CH=CH)
group and 1565, 1514 cm-1 bands due to aromatic benzene ring. The 1H NMR
spectrum (table 6 Fig.11) showed two doublets at δ 6.28 and 7.6 ppm, with
coupling constant J = 9.6 Hz which are characteristic for coumarins.
Additionally one methoxy group singlet at δ 3.96 ppm. Other two aromatic
protons singlets at 6.91 and 6.84 ppm were viewed. These two aromatic
protons singlets are explained by 6,7-disubstitution indicating either
scopoletin 6 or isoscopoletin 6a. Using UV spectroscopy (Fig.13&14), by
addition of sodium acetate solution according to Horowitz and his team91, a
notable bathochromoic shift from 340 to 390 nm with increase in the intensity
of the band indicated that the isolated compound is 6-methoxy-7-
hydroxycoumarin which known as scopoletin 6.
Scopoletin 6 was isolated and characterized from Artemisia annua L. by
Tzeng and his co-workers92. This is the first report indicating the isolation
and identification of coumarins (5 and 6) from the genus Cynanchum.
5 4 5 4
O HO
6 6
3 3
HO 7 O 2 O O 7 O 2 O
8 1 8 1
6 6a
37

(J, Hz)
3 6.28 1H, d, J= 9.6 Hz
4 7.6 1H, d, J= 9.6 Hz
5 6.91 1H, s
8 6.84 1H, s
C-6-(O-Me) 3.96 3H, s
2.2.3 Study of methylene chloride extract using GC/MS.

The methylene chloride extract was analyzed by the familiar GC/MS
technique. The GC chromatogram showed nine peaks corresponding to nine
compounds. These nine compounds were identified on the basis of a high
percentage of matching with authentic spectrum using NIST library. The
literature survey revealed no information about the following compounds
(7-15) characterized by GC/MS from methylene chloride extract.
2.2.3.1 Structure elucidation of compound 7
HO
7
CHO
The MS spectrum (Fig.15) of compound 7 showed [M] + at m/z 152

(95%) corresponding to C8H8O3 and [M-1]+at m/z 151 as a base peak (100%)
corresponding to C8H7O3. the fragment peaks at m/z 137 (5%) due to the loss
of methyl group from M+, 123 (20%) due to the loss of CHO from M+, 109
38
(25%) due to the loss of CH3, CO groups were in agreement with isovanillin
(3-hydroxy-4-methoxy benzaldehyde)93.
+
O
2 O
HO
O HO
HO
1 2
+
O
1 H O
[C8H7O3]+ O 3 [C7H5O3]+
H
m/z 151 m/z 137
+
3 -CH3, CO +
O
[C8H8O3]+
O
m/z 152 (95%)
HO HO
+
[C7H7O2]+ [C6H5O2]+
m/z 123 m/z 109
2.2.3.2 Structure elucidation of the fatty acid 8
OH
H
O
8 O
39
The fatty acid derivative 8 was identified from comparing its MS

spectrum (Fig.16) with the NIST library. The fragment peaks were in
agreement with 9-oxononanic acid.
1 2 3 4 5 7
6
OH
H
O O
1
7
2 3 4 5 6
[C8H15O2]+ [C2H3O2]+
m/z 144 m/z 60
[C7H13O2]+ [C3H5O2]+
m/z 129 + + +
[C6H11O2] [C5H9O2] [C4H7O2] m/z 73
m/z 115 m/z 101 m/z 87
The mass spectrum of compound 9 (Fig.17) showed M+ at m/z 180

(20%) as aperant peak corresponding to C11H16O2. The other fragment peaks
at m/z 165(5%) due to loss of methyl group from M+ , 152 (10%) due to the
40
loss of CO from M+, 137 (40%) due to the loss of CH3, CO respectively
followed by losing CO to give the base peak at m/z 111 corresponding to
[C8H15] + which undergo retro-Diels Alder reaction to gave a peak at m/z 81
+
(10%) corresponding to [C6H9] were in agreement with (R)-5,6,7,7a-
tetrahydro-4,4,7a-trimethylbenzofuran-2(4H)-one.
-CO O -CH3
[C11H16O2]+
m/z 180
+.
O
O
+ O
[C10H16O]+ [C10H13O2]+
m/z 152 m/z 165
-CH3
+ + +
-CO
O O + 2H
[C9H13O]+
m/z 137 [C8H15]+
m/z 111 (100%)
[C6H9]+ retro-Diels alder

m/z 81

OH
H
10
The mass spectrum of compound 10 (Fig.18) showed M at m/z 208

corresponding to [C13H20O2] +. The other fragment peaks at m/z 193 (3%)
corresponding to [C12H17O2] + due to the loss of methyl group from M +, 165
(4%) corresponding to [C11H17O] + due to the loss of methyl group followed
41
by losing of CO group, 137 (5%) corresponding to [C9H13O]+. The fragment

at m/z 108 (100%) which corresponding to [C8H12]+ representing the base
peak as a result of losing of CHO. The fragmentation pattern of 10 was in
agreement with 3-oxo-α-ionol from NIST library.
OH
H
[C13H20O2]+
m/z 208
-CH3
O
OH
+
+ H
O
O -CO
+
[C12H17O2]
m/z 193
+
+ +
-C2H4
O
O
[C9H13O]+ [C11H17O]+
m/z 137 -CHO m/z 165
[C8H12]+
m/z 108 (100%)
Scheme 6: proposed fragmentation of compound 10
OH
O O
H O
11
42
The mass spectrum of compound 11 (Fig.19) showed a parent peak at

m/z 182 (100%) corresponding to [C9H10O4]+. The other fragmentation peaks
at m/z 181 (62%) as a base peak corresponding to [M-H]+, m/z167 (13%) due
to the loss of methyl group from M, 153 (8%) due to the loss of CHO from
M+, m/z 139 (17%) due to the loss of CH3, CO groups from M+, 123 (7%)
corresponding to [C7H7O2]+, m/z 111 (22%) corresponding to [C6H7O2]+ due
to the loss of CH3, 2CO from M+, m/z 96 (15%) corresponding to [C5H4O2]+,
94
were in agreement with the MS spectrum of syringic aldehyde (NIST
library).
OH
O O
OH -H OH +
O O -CH3 O O
H O
+
[C9H9O4] [C9H10O4]+
m/z 181 m/z 182
+
O
H O
-CH3, CO [C8H7O4]+
-CHO
+ m/z 167
OH OH
+
O O O O
-CH3, 2 CO
[C8H9O3]+ [C7H7O3]+
m/z 153 [C6H7O2] + m/z 139
m/z 111
-O +
-CH3 OH
O
[C5H4O2]+
m/z 96
[C7H7O2]+
m/z 123
Scheme 7: proposed fragmentation of compound 11
43
HO
O 12
The mass spectrum of compound 12 (Fig. 20) showed a peak at m/z

228 corresponding to [M]+. The fragment peak at m/z 199 is due to the lose of
[C2H5]+. The fragment peak at m/z 73 which represents the base peak (100%)
is corresponding to [C3H5O2]+ resulting from McLafferty rearrangement due
to the presence of γ-hydrogen in a carbonyl compound. The saturated fatty
acid was identified by comparing its MS spectrum with the NIST library.
Compound 12 was in agreement with tetradeconic acid.
HO
O
60 73 87 101 115 129 143 157 171 185 199
[C14H28O2]
m/z 208
44
2.2.3.7 Structure elucidation of terpenoide 13
O
HO
13
The MS spectrum of compound 13 (Fig.21) showed M+ at m/z 196

(5%) corresponding to [C11H16O3]+ and a base peak m/z 111 due to [C6H7O2]+.
The fragment peaks at m/z 178 is due to the losing of water molecule from
M+, 153 (8%) due to [C9H13O2]+, 125 (5%) due to [C7H9O2]+ were in
agreement with the spectrum of loliolide 95 (NIST library).
O O
14
The MS spectrum of compound 14 (Fig.22) gave a fragment peak M+ at

m/z 168 (2%) corresponding to [C10H16O2]. The fragment peak at m/z 97
(100%) which represent the base peak is due to the loss of [C5H11]. The other
fragment peaks at m/z 81 (12%) is due to [C5H5O]+, 68 (50%) is due to
[C4H4O]+, 69 (20%) is due to [C4H5O]+ were in agreement with the spectrum
of massoilactone 96 (NIST library).
45
O O
[C10H16O2]
m/z 168 -C5H11
-CHO -O
O O
[C5H5O2]
m/z 97
+
[C4H4O]+ -CO
m/z 68
O
+ [C5H5O]
m/z 81
[C4H5O]
m/z 69
O
HO
15
The MS spectrum of compound 15 (Fig.23) was in agreement with the

MS spectrum of palmitic acid from NIST library.
46
2.3. Study of Ethyl acetate extract.

The chromatographic separation of ethyl acetate extract using TLC,
(EtOAc: MeOH, 19:1) solvent system, afforded a yellow solid polar material,
with a yellow color in the visible region upon spraying with sodium
hydroxide indicating a flavonoidal compound. In the 1H NMR spectrum
(DMSO) (Fig. 24), a signal appeared at δ 13 ppm which characteristic for
flavonides in addition to a characteristic signals for sugar moiety in range 4
and 5.3 ppm. This material needs further investigation.
47
3.1 Study of Curcuma longa L.

The powdered air dried rhizomes of Curcuma longa was processed
according to scheme 10.
Curcuma longa L
plant material (rhizomes)
1- Extraction by ethanol
2- Filtration and evaporation to dryness
3- Solvent fractination
Pet. ether fraction CHCl3 fraction
CC, silica gel CC, silica gel

benzene / EtOAc benzene / EtOAc
100 % benzene
Yellow-orange fraction 31 21 22
oily fraction 20
1
H NMR
GC/MS
19
16 18
17
Scheme 10: processing of Curcuma longa rhizomes
48
3.2. Study of the petroleum ether extract of Curcuma longa L.

The petroleum ether extract (cf. scheme 10) was evaporated to dryness
where a faint green oily material was obtained which was subjected to column
chromatograph using silica gel as stationary phase. Two fractions were
obtained. Fraction 1, a yellow orange oily material by the solvent 100%
benzene, where compounds 16, 17 and 18 were identified. Fraction 31
obtained using the solvent system benzene/EtOAc, 9:1 where compound 19
was obtained as yellow crystals.
3.2.1. Structure elucidation of the sesquiterpene ketone 16

12
3
6 5 4
2
8 3'
O 7 1
2'
10 9 11
16
Compound 16 present as viscous yellow orange oily material. The
literature survey of the phytochemical constituents of Curcuma longa oil
indicated the presence of many of sesquiterpene ketones and alcohols of
bisabolane skeleton97. The 1H NMR spectrum of compound 16 was consistent
with the expected signal pattern of bisabolane. It’s spectrum (table 7, Fig. 25)
showed three singlets for three methyl groups at 1.71, 1.95 and 2.18 ppm,
(Me-10, 11 and 1) respectively indicating that it had three quaternary methyl
groups. Also, the singlet at 2.18 ppm, integrated for three protons suggested
an aromatic methyl group (Me-1). A doublet at 1.13 ppm with coupling
constant J= 6.9 Hz integrated for three protons, indicating of a methyl group
49
C-12. One proton singlet at δ 5.90 ppm was assigned for proton at C-8. A
double of doublet at 2.45 and 2.55 ppm with coupling constant J= 8.4 Hz
were assigned for the two protons H-6 and H-6'. A multiplet at 3.2 ppm with
coupling constant J= 6.9 Hz were assigned for benzylic proton at C-5. A
double of doublet integrated for four protons at 6.94, 6.98 ppm with coupling
constant J= 8.45 Hz indicate the presence of AA'BB' system for (H-2, H-2',
H-3 and H-3') protons. The MS spectrum (Fig.26) gave M+ at m/z 216 (5%)
corresponding to [C15H20O]. The other fragment peaks at 201 (5%) due to the
lose of methyl group from M+, 119 (35%) due to the loss of C6H9O from M+,
91 (20%) due to the loss of C8H13O from M+, 83 (55%) due to [C5H7O]+, 98
(5%) due to McLafferty rearrangement, 55 (20%) due to [C4H7]+ confirmed
the structure of ar-turmerone which was isolated previously from Curcuma
longa L. by Geoffrey and his group97.

H atom δ, value, ppm Integration,
multiplicity (J, Hz)
1 2.18 3H, s
2, 2', 3, 3' 6.94, 6.98 AA'BB', 8.45
5 3.2 1H, m, 6.9
6 2.45 1H, dd, 8.4
6' 2.55 1H, dd, 8.4
8 5.9 1H, s
10 1.71 3H, s
11 1.95 3H, s
50
+
+
O
1 [C14H17O]+
+
[C9H11] 1 m/z 201
m/z 119 -CH3 +
3
2
O
+
2 [C7H7]+
m/z 91
O
[C15H20O] McLafferty rearrangment
m/z 216
+
3
[C5H7O]+ OH
m/z 83
[C4H7]+
m/z 55
[C6H10O]+
m/z 98
Scheme 11: Proposed fragmentation pattern of compound 16
51
3.2.2 Structure elucidation of sesquiterpene 17
17
The MS spectrum of sesquiterpene ketone 17 (Fig.27) showed M+ at

m/z 218 (8%) corresponding to [C15H22O], 119.99 (100%) due to [C9H12]+, 93
(2%) due to [C7H9] +, 91(10%) due to [C7H7]+, 83 (20%) due to [C5H7O]+, 55
(10%) due to [C4H7]+. The MS spectrum of 17 was in agreement with α-
Turmerone from (NIST library). α-Turmerone was reported previously as one
of the major constituents of Curcuma oil by Sharma and his co-workers83.
+ +
[C7H9]+ -2H
m/z 93 1
+ 2
2 1 [C9H12]+
m/z 119.99
4
[C7H7]+ 3
O
m/z 91 + 3 [C4H7]+
4 m/z 55
O
[C15H22O]
m/z 218
[C5H7O]+
m/z 82.99
52

H
O
H
18
Compound 18 gave a molecular ion peak, M+, at m/z 220

corresponding to C15H24O and fragment ion peaks at m/z 205 and 177 due to
the loss of CH3 and C3H7, respectively. The cyclic ion 177 gave a series of
fragment ion peaks at m/z 159, 149, 135, 121, 107, 95 and 79 due to the loss
of H2O, CO, CH2CO, C4H8, C5H10, C6H12 and C7H14, respectively.
Good agreement with the expected fragmentation pattern as well as
with an authentic spectrum and with that of the NIST library (Fig. 28) has led
to it's identifying as caryophyllene oxide98.
H
O
C3H5]+ 18
m/z 149
m/z 41 -CH3
-C3H7 H
-H2 -CO
C15H24O, M+, m/z 220
+ m/z 205
C3H7 + m/z 177 -C4H14
m/z 79 -H2O
-C6H12
-C
-H2O 5H m/z 187
10 m/z 93
-C4H8
m/z 159 -CH2CO
m/z 121 m/z 107

m/z 135
53
H
O O
2' 10 4 2
MeO 3' 1'
8 6
9 7 5 3 1
HO 4' 6' 19
5'
Compound 19 exists as yellow crystals with m.p. 84-85 oC. The

literature survey of Curcuma longa L. indicated the presence of diaryl
heptanoids that represent the major secondary metabolites that responsible for
the yellow color of its powder. On the other hand the literature survey of
ginger (Zingiber officinale) indicated the presence of gingerols and shogaols
that are responsible for the taste of ginger. The 1H NMR spectrum of
compound 19 was consistent with the expected signal pattern of gingerols.
The 1H NMR spectrum of compound 19 (table 8, Fig.29) showed only one
aromatic benzene ring containing (a broad singlet at δ 7.3 ppm corresponding
to H-2', a double of doublet at δ 7.14 ppm with coupling constants J= 8.05, 1.5
Hz corresponding to H-6', a doublet at δ 6.84 ppm with coupling constant J=
8Hz corresponding to H-5'). The presence of a down field doublet at δ 7.58
ppm, with coupling constant J= 15.45 Hz in a complement with an up field
doublet at δ 6.66 ppm with coupling constant J=15.45 Hz suggested the
presence of an AB system for two trans protons coupled with the aromatic
system suggesting the presence of a vinyl benzene system. The singlet at 5.95
ppm was assigned to the H-7, because the methylene protons at C-7 is
enolyzed to α, β- unsaturated carbonyl and appeared as singlet, thus this
suggested the presence of 1,3-diketone moiety. The singlet at δ 3.89 ppm,
integrated for three protons, was assigned to the methoxy protons at C-3’
position. Based on these evidences, compound 19 was identified as 1-
54
dehydrogingerdione which has been isolated previously from ginger (Zingiber

officinale) by Charles99. This is the first report indicating the isolation of 1-
dehydrogingerdione from Curcuma longa L. However it may be present in
the commercial processed sample as a result of either adulteration or
contamination by ginger (Zingiber officinale), but Zaeoung and his group100
reported the isolation of 1-dehydrogingerdione 19 with 6-gingerol and 6-
shogaol from Zingiber officinale in quantities 0.032, 0.078 and 0.042 g,
respectively. Thus 19 is the minor constituent in Zingiber officinale. Seeing
that the two other constituents, which are greater in quantity than 19 not
isolated with it from the sample under investigation, consequently, it is a
constituent of Curcuma longa and canceled the probability of being
originated from adulteration or contamination.

H atom δ, value, ppm Integration, multiplicity
(J, Hz)
2' 7.3 1-H, br s
5' 6.84 1-H, d, 8.00
6' 7.14 1-H, dd, 8.05, 1.5
10 7.55 1-H, d, 15.45
9 6.66 1-H, d, 15.45
7 5.95 1-H, s
O-Me 3.89 3-H, s
3.3 Study of chloroform extract of Curcuma longa L.

The chloroform extract (cf. scheme 10) was evaporated to dryness
where a red orange solid material was obtained which was subjected to
column chromatograph using silica gel and benzene/ethyl acetate as solvent
system. Only one major fraction (0.7g) was obtained which was further
55
purified using PTLC where the three major curcuminoides have been isolated
using the same solvent system (6:1) followed by elution with CHCl3.
H
O O
6 4 4' 6'
MeO 7' OMe
7 5 2' 5'
2
3 1 3'
H
8 10 10' 8'
HO OH
9 9'
20
Compound 20 exists as yellow-orange prismes; m.p. 183oC. The

literature survey of Curcuma longa L. indicated the presence of diaryl
heptanoids that considered the main active phytoconstituents responsible for
the yellow color of its powder. The 1H NMR spectrum of 20 was consistent
with the expected signal pattern of curcuminoids. Compound 20 is
symmetrical, so only half of the signals were evident in its 1H NMR spectra.
In the 1H NMR of compound 20 (table 9, Fig. 30), the assignment of H-3 and
H-4, as well as the H-3' and H-4' protons as trans to each others was based on
the large coupling constant between them where a doublet at δ 7.62 ppm with
coupling constant J=16.05 Hz is corresponding to H-4 and H-4', and another
doublet at 6.70 ppm with coupling constant J=16.05 Hz corresponding to H-3
and H3'. A singlet at 5.95 ppm is corresponding to H-1, because the
methylene protons at C-1 in the curcuminoids are enolyzed to α-β-
unsaturated carbonyl and appear as singlet. A broad singlet at 7.3 ppm,
integrated for two protons, is corresponding to H-6 and H-6', a broad doublet
at 7.14 ppm with coupling constant J=7.65 Hz, integrated for two protons, is
corresponding to H-10 and H-10'. Another doublet at 6.85 ppm with coupling
constant J= 7.65 Hz, integrated for two protons, is corresponding to H-9 and
56
H-9'. The singlet at 3.89 ppm, integrated for six protons, was assigned to the
two methoxy groups at C-7 and C-7'. Based on the previous spectral data,
compound 20 was found to be in agreement with 1,7-bis(4-hydroxy-3-
methoxyphenyl)1,6-heptadien-3.5-dione, known as curcumin I
(diferuloylmethane) which was isolated previously from Curcuma longa L.
by Geoffrey97.

H atom δ, value, ppm integration,
4, 4' 7.62 2H, d, 16.05
3, 3' 6.70 2H, d, 16.05
1 5.95 1H, s
6, 6' 7.31 2H, br s
9, 9' 6.86 2H, d, 7.65
10, 10' 7.14 2H, dd, 7.65
7,7'- OCH3 3.89 6H, s
57
3.3.2 Structure elucidation of Compound 21
H
O O
6 4 4'
6'
MeO 7'
7 5 2' 5'
2
3 1 3'
H
8 10 10' 8'
HO OH
9 9'
21
Compound 21 exists as yellow-orange powder, m.p 168 oC, also the 1H

NMR spectrum of 21 was consistent with the expected signal pattern of
curcuminoids. It lacked the symmetry of compound 20. In the 1H NMR of 21
(table 10, Fig.31), the assignment of H-3and H-4, as well as the H-3' and H-4'
protons as trans to each other, was based on the large coupling constant
between them where a two downfield doublets at δ 7.55 ppm with coupling
constant J= 16.8 Hz, integrated for one proton, corresponding to H-4. A
doublet at 7.56 ppm with coupling constant J=16.05 Hz, integrated for one
proton, corresponding for H-4'. On the other hand, the presence of two upfiled
doublets, one at δ 6.63 ppm, with coupling constant J=16.05 Hz integrated
for one proton was assigned to H-3. The other doublet at δ 6.68 ppm, with
coupling constant J=15.3 Hz integrated for one proton, was assigned to H-3'.
A singlet at δ 5.95 ppm corresponding to H-1. A doublet at δ 6.85 ppm with
coupling constant J= 8.4 Hz, integrated for two protons, corresponding to H-7'
and H-9'. A doublet at δ 7.53 ppm with coupling constant J=9.2 Hz, integrated
for two protons, corresponding to H-6' and H-10'. A doublet at δ 7.32 ppm
with coupling constant J=1.5 Hz, (meta coupling) integrated for one proton,
corresponding to H-6. A double of doublet at δ 7.14 ppm with coupling
constant J=8.4, 1.5 Hz, integrated for one proton, corresponding to H-10. A
doublet at δ 6.87 ppm with coupling constant J=8.4 Hz, integrated for one
proton, corresponding to H-9. A singlet at 3.89 ppm, integrated for three
58
protons, corresponding to methoxy group at C-7. Based on the discussed

spectral date, compound 21 was found to be in agreement with 1-(4-hydroxy-
3-methoxyphenyl)-7-(4-hydroxyphenyl)1,6-heptadien-3.5-dione, known as
curcumin II (demethoxycurcumine) which has been isolated from Curcuma
longa by Geoffrey97.
Table 10: 1H NMR date of Compound 21

1 5.95 1H, s
3 6.62 1H, d, 16.05
3' 6.68 1H, d, 15.3
4 7.55 1H, d, 16.8
4' 7.56 1H, d, 16.05
6 7.32 1H, d, 1.5
9 6.88 1H, d, 8.4
10 7.14 1H, d, 8.4, 1.5
6', 10' 7.53 2H, d, 9.2
7', 9' 6.85 2H, d, 8.4
7-OMe 3.89 3H, s
59
H
O O
6 4 4' 6'
7 5 5' 7'
2 2'
3 1 3'
H
8 10 10' 8'
HO OH
9 22 9'
Compound 22 exists as yellow-orange material, m.p. 224 oC; it is

symmetrical as compound 20, so only half of the signals was evident in its 1H
NMR spectra. In the 1H NMR of compound 22 (table11, Fig.32), the
assignment of H-3, H-3', H-4, H-4' and H-1 were concluded by the same
manner as for compound 20. A downfield doublet at δ 7.56 ppm with
coupling constant J=15.45 Hz, integrated for two proton, was assigned to H-4
and H-4'. An upfield doublet at δ 6.63 ppm with coupling constant J=16.05
Hz, integrated for two protons, was assigned to H-3 and H-3'. A singlet at δ
5.95 ppm corresponding to H-1. The 1H NMR spectrum of compound 22 gave
two types of aromatic protons which characteristic of AA'BB' system
represented by a downfield doublet at δ 7.53 ppm with coupling constant J=
8.6 Hz, integrated for four protons, corresponding to H-6, 6', 10, 10' and an
upfield doublet at δ 6.86 ppm with coupling constant J=8.6 Hz, integrated for
four protons, corresponding to H-7, 7', 9, 9'. Based on the previous spectral
data compound 22 was found to be in agreement with 1,7-bis(4-
hydroxyphenyl)1,6-heptadien-3.5-dione, known as curcumin III
(bisdemethoxycurcumin) which was isolated previously from Curcuma longa
L. by Geoffrey97.
60

1 5.95 1H, s
3, 3' 6.63 2H, d, 16.05
4, 4' 7.59 2H, d, 15.45
6, 6', 10, 10' 7.53 4H, d, 8.6
7, 7', 9, 9' 6.86 4H, d, 8.6
61
3.4. Insecticidal activities of the volatile oil and 1- dehydrogingerdione

isolated from Curcuma longa against the confused flour beetles,
Tribolium confusum.
The major compounds like ar-turmerone and α-turmerone which are
reported in the literature as insect repellent or insecticidal and are also present
in higher concentration compared with other in addition to 1-
dehydrogingerdione.
 Contact and Fumigant Toxicity.
Contact Toxicity.
The crude extracts from Curcuma longa rhizomes were analyzed for
insecticidal activity towards the T. Confusum larvae. After 4, 8, & 12 days of
exposure to the treated diet, larval survival was monitored and compared to
controls. Petroleum ether extract exhibited significant insecticidal activity and
caused strong larval mortality. Bioassay-guided isolation of the petroleum
ether extract afforded two main active sesquiterpene ketone 16 and 17 as oily
fraction in addition to 1-dehydrogingerdione 19 as yellow crystalline material
from fraction 31. These two main active sesquiterpene ketone 16 and 17 in
addition to compound 19 were shown to be responsible for the insecticidal
activity of the crude extract (table 12).
The oily fraction of Curcuma longa was relatively more active than 1-
dehydrogingerdion. The copulated mixture of both tested materials, resulting
in complete larval mortality even at the lowest concentration analyzed (4.86
mg/200g flour). Whereas, a significant reduction of larval survival by 1-
dehydrogingerdione was only observed at the highest concentration analyzed
where the LC50 values of Curcuma oil and gingerdione were (11.o8, 6.72, &
4.86 mg/200g flour) (Fig.33) & (30.57, 18.49, & 15.53 mg/200g flour) (Fig.
34), after exposure time of 4, 8 & 12 days of treatment, respectively (Table
12). Whereas, the LC50 values of the mixture from oil and gingrdione were
(18.44, 14.49 & 9.03) at the same exposure times, respectively (Fig.35). This
62
results is in agreement with the reported data by Tripathi and his team101 on
the insecticidal activity of the turmeric leave essential oil against three stored-
producr beetles, the oil was insecticidal in both contact and fumigant toxicity
assays. The adults of R. dominica were highly susceptible to contact action of
C. longa leaf oil, with LD50 value of 36.71 mg/g weight of insect, whereas in
the fumigant assay, adults of S. oryzae were highly susceptible with LC50
value of 11.36 mg/liter air, by the same application methods used in this
study.
Fig.33. LC50 values of Curcuma oil
63
Fig.34. LC50 values of 1-dehydrogingerdion
Fig.35. LC50 values of Curcuma oil mixed with gingerdione

 Fumigant toxicity.
The essential oil of C. longa rhizomes showed a strong toxicity to the
tested species (Table13). Adults of T. confusum were more susceptible than
its larvae (Table 13). In addition, susceptibilities of the various larval stages
(14 -18 days old) to the fumigating toxicity of C. Longa oil were variable and
the susceptibility decreased with the age increasing (Table13). Results of the
64
present investigation indicate that the essential oil of C. longa rhizome has got
high bioactivities against adults of T. confusum. However, Tawatsin and his
team102 have reported repellent activity against A. aegypti, A. dirus and C.
quinquefasciatus which is due to 5% vanillin which has been added to the
essential oil of C. longa. Therefore the repellent activity observed by
Tawatsin and his team is due to synergistic action of vanillin.
Table 12: Contact toxicity of the isolated materials, toward the larvae of the
confused flour beetle T. Confusum
Tested Line Line LC50 Lowe Uppe 1 2 3 Index LC90 RR Slope±
material No name (mg/200g r limit r (mg/100g SE
m) limit m)
C. longa
Oil
1 12 4.86 ----- ----- 533.02 17.67 0.50 2.29±0.
days 3 7 001
treatme
nt
2 8 days 6.72 6.003 7.626 * 211.83 23.96 0.46 2.32±0.
treatme 3 003
nt
3 4 days 11.08 9.245 15.07 * 100 49.95 1 1.96±0.
treatme 9 001
nt
Gingerdione
1 12 15.53 14.36 16.87 * 196.89 45.56 0.50 2.74±0.
days 8 021
treatme
nt
2 8 days 18.49 17.13 20.52 * 165.26 49.23 0.60 3.02±0.
treatme 8 5 018
nt
3 4 days 30.57 25.25 44.53 * 100 101.88 1 2.45±0.
treatme 009
nt
Gingerdione
Mixure with C.
longa oil
1 12 days 9.03 8.15 9.92 * 204.23 33.29
0.49 2.261±0.0
treatment 01
2 8 days 14.49 12.51 18.09 * * 127.19 28.57 0.79 1.63±0.00
treatment 2
3 4 days 18.44 15.20 25.73 * * 100 126.04 1 1.535±0.0
treatment 11
Index compared with 4 days treatment RR: Resistance Ratio compared with 4 days treatment
65
Table 13: Fumigant toxicity of C. longa oil to adults and larvae of T.

confusum.
Life n LC50 95% Þducial X2 LC90 95% Þducial Slope±SE
stage (mg/ liter limits (mg/ liter limits
air) air)
Adults 600 18.315 16.097- 1.47 85.842 60.319- 1.91±0.02
20.838 157.981
Larvae
14 d old 600 25.903 22.975- 0.672 99.57 70.27- 2.192±0.0
30.441 177.313 28
16 d old 600 30.825 26.557- 2.906 131.35 85.382- 2.036±0.0
38.683 279.26 1
18 d old 600 34.351 29.379- 1.609 131.699 86.835- 2.196±0.0
43.812 270.035 5
X2 is significant (P˃0.05)
 Effect of C. longa rhizomes Oil on Oviposition, Egg Hatchability and

Larval Survival of T. confusum.
Oviposition was significantly (F=50; df =5, 56; P<0.05) reduced by C.
longa rhizomes oil at the concentrations of 6.42 and 8.26 mg/cm2 (Table 14).
However, as there was 12.6 and 17.9% mortality, respectively, observed in
these two concentrations after 24 h, the decrease in egg production was
probably due to fewer surviving insects laying eggs or the weakened physical
stage of the insects. Since the oviposition dropped to 26% at 5.2 mg/cm2
concentration of treatment with C. longa rhizome oil, the oil probably acted
as an oviposition deterrent at high concentrations. The hatching of T.
confusum eggs and emergence of larvae were adversely affected by C. longa
leaf oil (Fig.36). At 5.4 mg/cm2, egg hatching was reduced by 70% and no
larvae were observed the 9th day after hatching at the concentration of 12.6
mg/cm2 and above. Thus, C. longa rhizome oil was ovicidal also to the eggs
of T. confusum. When the T. confusum eggs were exposed to C. longa
rhizome oil, the number of adults that emerged from the treated eggs was
significantly (F =89; df = 5, 42; P<0.05) decreased (Fig. 37). At 16.2 mg/cm2
concentration of C. longa rhizome oil, no adults emerged, since no larvae
survived at this concentration in the previous experiment (Fig. 6). Therefore,
66
C. longa leaf oil prevented the oviposition of T. confusum and it suppressed

the egg hatching along with the subsequent development of eggs to larvae and
adults.
Table 14: Effect of 24-h exposure to C. longa L. oil impregnated filter papers
on oviposition by T. confusem
Conc., Mortality of adults at 24 h No. eggs laid per ♀
mg/cm2 (mean±SE) (mean ±SE)
0.0 0.0±0.0 15.4±0.2
0.092 0.0±0.0 13.7±0.5
0.92 0.3±0.0 11.9±0.3
2.92 4.1±0.1 10.8±0.5
5.20 6.3±0,2 7.0±0.6
6.42 12.6±0.1 5.2±0.3
8.26 17.9±0.3 2.1±0.5
F 50.97
Fig.36: Percentage of egg hatch and survival of larvae of Tribolium

castaneum exposed to Curcuma longa oil.
67
Fig.37: Percentage of adult emergency of T. confusum from eggs exposed to

C. longa oil.
68
EXPERIMENTAL
EXPERIMENTAL
3.1 Instrumentations.
 1
H NMR.
1
H NMR spectra were recorded on 500 MHz (JEOL). Chemical shifts are
given in δ (ppm) relative to TMS as internal slandered material at Faculty of
Science, Alexandria University.
 Infrared spectra (IR).

Infrared spectra (IR) were recorded as thin film cast from CHCl3, and
performed on Mattson 5000 FT-IR spectrometer at Faculty of Science, Mansoura
University.
 GC/MS.
GC/MS analyses were performed on a Varian GC interfaced to Finnegan
SSQ 7000 mass selective Detector (SMD) with ICIS V2.0 data system for MS
identification of the GC components. The column used was DB-5 (J&W
Scientific, Folosm, CA) cross-linked fused silica capillary column (30 m.long,
0.25mm. internal diameter) coated with ploydimethylsiloxane (0.5µmfilm
thickness). The oven temperature was programmed from 50 oC for 3 min., at
isothermal, then heating by 7 oC/ min. to 250 oC and isothermally for 10 min., at
250 oC. Injector temperature was 200 oC and the volume injected was 0.5µl.
transition-line and ion source temperature was 250 oC and 150 oC respectively.
The mass spectrometer had a delay of 3 min. to avoid the solvent pead and then
scanned from m/z 50 to m/z 300. Ionization energy was set at 70eV. (Agriculture
Research Center, National Research Center (NRC), Dokki, Cairo, Egypt). Thin
layer chromatography and preparative (TLC) were performed on silica gel
(Kieselgel 60, GF 254) of 0.25 thickness.
69
EXPERIMENTAL
 Solvents.
Petroleum ether (60-80oC), hexane, methylene chloride, chloroform,
ethyl acetate and methanol were obtained from ADWIC company; diethyl ether
was obtained from NICE chemical company. Acetones, Methanol HPLC were
obtained from TEDIA and SDS companies respectively.
3.2. Plant materials.

Cynanchum acutum L. was collected from new Damitta, Egypt in May
2005 by prof. Dr. Mamdouh Abdel-Mogib then identified by prof. Dr. Ibrahim
Mashaly, Botany Dept., Faculty of Science, Mansoura University.
Curcuma longa L. a commercial powder sample was purchased from a
local market in Mansoura, Egypt in December 2007.
3.3. Spray .
reagents
P-anisaldehyde – sulphuric acid reagent.103
A freshly prepared solution was made by adding conc. sulphuric acid (8
ml) and 0.5 ml anisaldehyde under cooling with ice to a mixture of 85 ml
methanol and 10 ml glacial acetic acid. The chromatogram after spraying was
heated at 100 oC until the spots attained the maximum color.
70
EXPERIMENTAL
3.4 processing of plant materials under investigation.

3.4.1. Cynanchum acutum L.
Air dried aerial parts of Cynanchum acutum (800 g) were extracted by
soaking at room temperature in methanol. After evaporization, the crud extract
(40 g) was dissolved in the least amount of methanol and diluted with water and
successively extracted with petroleum ether (40-60 oC) followed by extraction
with methylene chloride and finally with EtOAc.
3.4.1.1. Saponification and chromatography of petroleum ether extract.

The petroleum ether extract (6.7g) was saponified using alcoholic
aqueous sodium hydroxide (5 %) and stirring for 2 hours then re-extracted by
pet. ether to give the saponifiable part (1.5 g). Acidification by conc. HCl (5 %)
and re-extraction by chloroform afforded the non saponifiable part (2 g). The non
saponafiable part of petroleum ether fraction (2 g) was separated over neutral
aluminum oxide (Al2O3) as adsorbent. The column was eluted with hexane /
ethyl acetate solvent system with increasing in polarity. Lupyl acetate was
isolated by (hexane / EtOAc, 49:1), then lupeol by hexane / EtOAc (24: 1), and
finally a mixture of β-sitosterol and stigmasterol by hexane / EtOAc (13: 1).
Compounds were further purified using precoated TLC (silica gel,
hexane/EtOAc 49:1) using the same solvent system of CC to give lupyl acetate
(0.25g, Rf = 0.72), lupeol (0.15g, Rf = 0.3), mix of β-sitosterol and stigmasterol
(0.1g, Rf = 0.25).
71
EXPERIMENTAL
3.4.1.2. Isolation of coumarins from methylene chloride extract.

The fraction of CH2Cl2 (2.6 g) was resolved over a silica gel CC which
was eluted with petroleum ether / ethyl acetate solvent system with increasing
polarity. Scopoletin (pet. ether / EtOAc 1: 1), and 6,7-dimethoxycoumarin
(scoparone) (pet. Ether/EtOAc 3:2) were isolated. The coumarins were further
purified by TLC (silica gel). Scopoletin was purified using petroleum ether /
EtOAc (6:5), (0.198 g, Rf = 0.21) while scoparone was purified by the same
solvent system (3:2), (0.2 g, Rf = 0.39).
3.4.1.2.1 Chemical composition of methylene chloride extract by GC/MS.

A sample of CH2Cl2 fraction was analyzed by GC/MS where the following
known natural compounds were characterized; isovanillin (Rt = 12.68 min., 1.08
%), 9-oxononanic acid (14.03, 0.63%), (R)-5,6,7,7a-tetrahydro-4,4, 7a-
trimethylbenzofuran-2(4H)-one (14.81, 1.00%), 3-oxo-α-Ionol (16.42, 1.96 %)
syringic aldehyde (16.61, 2.54%), tetradeconic acid (16.61, 2.54%), (-)-Loliolide
(17.99, 2.05 %), massoilactone (19.9, 0.78 %) and hexadeconic acid (20.59,
2.46).
3.4.1.3. Investigation of the ethyl acetate extract.

Chromatography of this fraction using TLC, (EtOAc: MeOH) (19:1)
solvent system, afforded a yellow solid polar substance, that showing a yellow
color in the visible region upon spraying with NaOH indicating a flavonidal
compound. In the 1H NMR spectroscopy (DMSO), a signal appeared at δ 13 ppm
which characteristic for flavonides in addition to a characteristic signals for sugar
moiety in range 4 and 5.3 ppm. This material needs further investigation.
72
EXPERIMENTAL
3.4.2. Characterization of the isolated compounds from Cynanchum acutum.
 Acetyl-3-O-Lupane (1)
White needles , m.p. 218-220 oC, (0. 25 g, 12.5%), 1H NMR (CDCl3): δH 4.46
(1H, dd, J = 10.6 Hz, H-3), 2.38 ( 1H, ddd, J = 10.6 Hz, H- 19), 0.94 ( 3H, s,
Me-23), 0.76 ( 3H, s, Me-24), 0.83 ( 3H, s, Me-25), 1.03 ( 3H, s, Me-26),
0.96 (3H, s, Me-27), 0.79 ( 3H, s, Me-28), 4.55 ( 1H, brd, H-29), 4.67 ( 1H,
brd, H'-29), 1.67 ( 3H, brs, Me-30), 3.6 ( 3H, s, CH3-CO ).
 Lupeol (2)
White needles, m.p. 211-213 oC(0.15 g, 7.5%), 1H NMR (CDCl3): δH 3.2
(1H, dd, J = 10.6 Hz, H-3), 2.38 ( 1H, ddd, J = 10.6 Hz, H- 19), 0.94 ( 3H, s,
Me-23), 0.76 ( 3H, s, Me-24), 0.83 ( 3H, s, Me-25), 1.03 ( 3H, s, Me-26),
0.96 (3H, s, Me-27), 0.79 ( 3H, s, Me-28), 4.53 ( 1H, brd, H-29), 4.66 ( 1H,
brd, H'-29), 1.67 ( 3H, brs, Me-30).
 β-sitosterol (3)
White needles, m.p 136-137 oC 1
H NMR (CDCl3): δH 3.52 (1H, m, H-3),
5.35 (1H, m, H-6), 0.69 (3H, s, Me-18), 1.01 (3H, s, Me-19), 0.92 (3H, d, J =
6.4 Hz, Me-21), 0.83 (3H, d, J = 6.8 Hz, Me-26), 0.81 (3H, d, J = 6.9 Hz, Me-
27), 0.85 (3H, t, J = 7.8 Hz, Me-29).
 Stigmasterol (4)
White needles, m.p. 170 oC, 1H NMR (CDCl3): δH 3.52 (1H, m, H-3), 5.35
(1H, m, H-6), 0.69 (3H, s, Me-18), 1.01 (3H, s, Me-19), 0.92 (3H, d, J = 6.4
Hz, Me-21) 5.00 ( 1H, dd, J = 8, 14 Hz, H-22), 5.21 (1H, dd, J = 8, 14 Hz, H-
23), 0.82 (3H, d, J = 7 Hz, Me-26), 0.83 (3H, d, J = 7 Hz, Me-27), 0.97 (3H, t,
J = 7 Hz, Me-29).
73
EXPERIMENTAL
 Scopoletin (5)
Yellow crystalline material, m.p. = 204 oC, (0.19 g, 7.5%) I.R.cm-1: 3396
(OH hydrogen bonded), 1713 (Carbonyl group, α- lactone), 1611 (CH=CH),
1565, 1514 (aromaric benzene ring). ; UV, λ max, MeOH: 340 nm; 1H NMR
(CDCl3), δ 7.6 (1H, d, J = 9.6 Hz, H-4), 6.92 (1H, s, H-5), 6.86 (1H, s, H-8),
6.28 (1H, d, J = 9.6 Hz, H-3), 3.96 (3H, s, O-Me).
 Scoparone (6)
Colorless needles material, m.p = 144 oC, (0.2 g, 7.6%), GC/MS, m/z (rel.
int): 206 (100) [M,]+, 191 (50) [M-CH3]+, 178 (30) [M-CO]+, 163 (40) [M-
CH3, CO]+, 135 (25) [M-CH3, 2CO]+; 1H NMR (CDCl3): δ 7.6 (1H, d, J =
9.6 Hz, H-4), 6.92 (1H, s, H-5), 6.86 (1H, s, H-8), 6.28 (1H, d, j = 9.6 Hz, H-
3), 3.91 (3H, s, OCH3), 3.94 (3H, s, OCH3).
 Isovanillin (7)
(Rt = 12.68 min., 1.08 %), GC/MS, m/z (rel. int): 152 (100) [M]+, 151 (80)
[M-H]+, 137 (8) [M-CH3]+, 123 (20) [M-CHO]+, 109 (25) [C6H5O2]+, 108
(10) [C6H4O2]+, 81 (30) [C5H5O]+, 65 (10) [C5H5]+, 50 (7) [C4H2]+.
 9-Oxononanic acid (8)

(Rt= 14.03, 0.63%), GC/MS, m/z (rel. int): 144 (20) [C9H16O2]+, 129 (27)
[C7H13O2]+, 115 (10) [C6H11O2]+, 101 (30) [C5H9O2]+, 87 (25) [C4H7O2]+, 73
(83) [C3H5O2]+, 60 (45) [C2H3O2]+.
 (R)-5, 6, 7, 7a-tetrahydro-4, 4, 7a-trimethylbenzofuran-2(4H)-one (9)

(Rt= 14.81, 1.00%), GC/MS, m/z (rel. int): 180 (20) [M]+, 165(5) [M-CH3]+,
152 (10) [M-CO]+, 137 (40) [M-CH3, CO]+, 111 (100) [C8H15]+, 81 (10%)
[C6H9]+ .
74
EXPERIMENTAL
 3-oxo-α-Ionol (10)
(Rt = 16.42, 1.96 %), GC/MS, m/z (rel. int): 208 (2) [M]+, 193 (3) [M-CH3]+,
165 (4) [M-CH3, CO]+, 173 (5) [C9H13O]+, 108 (100) [C8H12]+.
 Syringic aldehyde (11)

(Rt = 16.61, 2.54 %), GC/MS, m/z (rel. int): 182 (100) [M,]+, 181 (60) [M-
H]+, 167 (13) [M-CH3]+, 153 (8) [M-CHO]+, 139 (17) [M-CH3, CO]+, 123 (7)
[C7H7O2]+, 111 (22) [C6H7O2]+, 96 (15) [C5H4O2]+. 109 (25) [C6H5O2]+, 81
(30) [C5H5O]+, 65 (10) [C5H5]+, 50 (7) [C4H2]+.
 Tetradeconic acid (12)

(Rt = 16.61, 2.54 %), GC/MS, m/z (rel. int): 228 (10) [M]+, 199 (5)
[C12H23O2]+ .Other fragment peaks at different m/z values obtained by losing
CH2 group each time.
 (-)-loliolide (13)
(Rt = 17.99 min, 2.05 %), GC/MS, m/z (rel. int): 196 (5) [M]+, 178 (44)
[C11H14O]+, 153 (8) [C9H13O2]+, 125 (5) [C7H9O2]+ and a base peak m/z 111
due to [C6H7O2]+.
 Massoilactone (14)
(Rt = 19.9 min, 0.78 %) GC/MS, m/z (rel. int): 168 (2) [M]+, 97 (100) which
represent the base peak is due to the loss of [C5H11]+, 81 (12) [C5H5O]+, 68
(50) [C4H4O]+, 69 (20) [C4H5O]+.
 Hexadeconic acid (15)

(Rt = 20.59 min., 2.46) GC/MS, m/z (rel. int): 256 (22) [M]+, 227 (22)
[C14H27O2]+. Other fragment peaks at different m/z values obtained by losing
CH2 group each time.
75
EXPERIMENTAL
3.4.3 Curcuma longa L.

Fresh rhizomes of Curcuma longa L. (375 g) were extracted by soaking
at room temperature in ethanol. The ethanolic extract was evaporated under
reduced pressure till a suitable volume then successively extracted by petroleum
ether (40-60 oC) afford about (7g) oily materials. Extraction by chloroform gave
yellow orange solid (1.34 g).
3.4.3.1 Chromatography of the petroleum ether extract.
The petroleum ether extract (2 g) was resolved over a silica gel column,
which was eluted by 100% benzene solvent system to afford the red-yellow oily
fraction (1.69g) (aromatic turmerone and α- turmerone). Increasing polarity
using CH2Cl2 till 100% then using EtoAc to give 1- dehydrogingerdione at 90%
benzene: 10% EtoAc. Isolated compounds were further purified using precoated
TLC plates. 1-dehydrogingerdione was purified by elution with 100% benzene
followed by chloroform two times where yellow crystals (0.25g, Rf 0.5 benzene)
were obtained.
3.4.3.2 Chromatography of chloroform extract.
The yellow orange solid extract (1.34g) was resolved over a silica gel
column. Elution with benzene / EtOAc solvent system with increasing polarity
till 80% benzene / 20% EtOAc, where three major curcuminiods were isolated as
orange solid precipitate after evaprization (fraction 25, 0.75g). Isolated
compounds were further purified using TLC (silica gel, benzene / EtOAc, 6:1).
Elution with chloroform gave the three major diarylheptanoids, curcumin I (0.6g)
(hexane/EtOAc, 2:1, Rf = 0.24), curcumin II (demethoxycurcumin) (0.24g)
(hexane/EtOAc, 2: 1, Rf = 0.27), curcumin III (bisdemethoxycurcumin) (0.1g)
(hexane-EtOAc, 2: 1, Rf = 0.25).
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EXPERIMENTAL
3.4.4 Charactrization of compounds isolated from Curcuma long L.
 ar- Turmerone (16)

Oily material, 1H NMR (CDCl3): δ 1.13 (3H, d, J=6.9Hz, H-12), 1.71 (3H, s,
H-13), 1.99 (3H, s, H-14), 2.18 (3H, s, H-1), 2.48 (1H, dd, J=8.4, 6.15 Hz,
H-8), 2.58 (1H, dd, J=8.4, 6.15 Hz, H-8'), 3.17 (1H, m, H-7), 5.9 (1H, s, H-
10), 6.97 ( 4H, dd, J=8.45, H-2, 3, 4, 5). GC/MS: (Rt = 28.68, Area = 0.41%),
m/z (rel.Int); 216 (5) [M] +, 201 (5) [M-CH3] +, 119 (35) [M-C6H9O] +, 91 (20)
[M-C8H13O] +, 83 (55) [C5H7O] +, 95 (5) [C6H10O] +, 55 (20) [C4H7] +.
 Turmerone (17)
Oily material, GC/MS: (Rt = 32.52, Area = 23.42%), m/z (rel.Int); 218 (8)
[M]+ , 119.99 (100) [C9H12]+, 93 (2) [C7H12]+, 91 (10) [C7H7]+, 83 (20)
[C5H7O]+, 55 (10) [C4H7]+.
 (-)-caryophyllene oxide (18)

Oily material, GC/MS: (Rt = 28.95, Area = 0.40%), 220 (5) [M]+, 205 (10)
[M-CH3]+, 177 (10) [M-CH3, CO]+, 151 (15) [C11H19]+, 123 (25) [C9H15]+.
 1-Dehydrogingerdione (19)
yellow crystals with m.p. = 84-85 oC, (0.25g, 12.5%), 1H NMR (CD3)2CO: δ
7.58 (1H, d, J=15.45Hz, H-1), 7.14 (1H, dd, J=8.05, 1.5 Hz, H-6'), 7.30 (1H,
d, J=1.5Hz, H-2'), 6.84 (1H, d, J=8Hz, H-5'), 6.66 (1H, d, J=15.45Hz, H-2),
5.95 (1H, s, H-4), 3.89 ( 3H, s, OMe), 2.37 ( 2H, m, H-6), 1.65 ( 2H, m, H-7),
1.32 ( 4H, m, H-8, H-9), 0.91 ( 3H, t, H-10).
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EXPERIMENTAL
 Curcumin I (20)
yellow-orange prismes; m.p. 183 oC, (0.5g, 66% ) 1H NMR (CD3)2CO: δ 3.89
(6H, s, 2 OMe), 5.95 (1H, s, H-1), 6.70 (2H, d, J=16.05 Hz, H-3, 3'), 6.86 (2H,
d, J=7.65 Hz, H-9, 9'), 7.16 (2H, dd, H-10, 10'), 7.31 (2H, d, H-6, 6'), 7.62 (2H,
d, J=16.05 Hz, H-4, 4').
 Curcumin II (21) (demethoxycurcumin)

yellow-orange powder, m.p 168 oC, (0.24g, 32%) 1H NMR (CD3)2CO: δ 3.89
(3H, s, OMe), 5.95 (1H, s, H-1), 6.62 (1H, d, J=16.05 Hz, H-3), 6.68 (1H, d,
J=15.3 Hz, H-3'), 6.85 (2H, d, J=8.4 Hz, H-7', 9'), 6.88 (1H, d, J=8.45, H-9),
7.14 (1H, dd, J=8.4, 1.5 Hz, H-10), 7.32 (1H, d, J=1.5 Hz, H-6), 7.53 (2H, d,
J=9.2 Hz, H-6', 10'), 7.55 (1H, d, J=16.8 Hz, H-4), 7.56 (1H, d, J=16.05 Hz, H-
4').
 Curcumin III (22) (bisdemethoxycurcumin)

yellow-orange residue, m.p. 224 oC, (0.1g, 13%) 1H NMR (CD3)2CO: δ 5.95
(1H, s, H-1), 6.63 (2H, d, J=16.05Hz, H-3, 3'), 6.88 (4H, d, J=8.6Hz, H-7, 7', 9,
9'), 7.53 (4H, d, J=8.6, H-6, 6', 10, 10'), 7.56 (2H, d, J=15.45Hz, H-4, 4').
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EXPERIMENTAL
3.5 Insecticidal activities of volatile oil and 1-dehydrogingerdione isolated

from Curcuma longa L. petroleum ether extract against the confused flour
beetles Tribolium confusum.
 Insect culture.
Eggs, larvae and adults of the confused flour beetle, T. confusum, were
obtained from laboratory- bred culture, maintained in the laboratory of
entomology for three successive years. Stock flour (sterilized at 60 ºC for 30
min) was used as a diet. By separating the eggs and incubated for about 15±2
days we can obtain a 3rd instars larvae of T. Confusum. Then the bioassay can be
start.
 Contact toxicity bioassay.

In this experiment, the sterilized flour was treated with a series of
dilutions of essential oil of C. longa and 1-dehydrogingerdione, were prepared in
acetone with some modifications104. The wheat flour was left to dry at ambient
conditions, in uncovered containers, to allow the solvent to evaporate, after
complete solvent evaporation, blending with electrical blender to ensure compete
mixing and spreading of the tested materials in the wheat flour. Control flour
treatment (-ve control) and control flour with acetone only treatment (+ve
control) were exactly handled in a similar manner. Twenty five equal-size larvae
(15±2 day-old), in four replicates, were used for each concentration and a similar
number was used in the control. The dead larvae were counted every four days
through a period of 12 days. The experiment was conducted under laboratory
conditions of 24±2 ºC and 60-70% RH.
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EXPERIMENTAL
 Fumigant toxicity.
According to Tripath and his team101, the fumigant toxicities of C. longa
oil were tested. Filter paper strips (6 x 8 cm) were impregnated with 10 μl of one
dilution of C. longa oil or acetone (control). After evaporating the solvent for 5
min in air, the treated filter paper strip was placed at the bottom of a 1-liter glass
bottle. Twenty adults of the tested insect species and/or larvae of T. confusum
along with culture media were kept separately in small vials that had both open
ends covered with 40-mesh copper wire net. Each small vial was hung with thin
metal wire at the geometrical center of the glass bottle and the bottle tightly
closed with the lid. Each set was considered as one replication. There were 10
replicates for each dilution and control. After 24 h, insects were transferred to
fresh vials containing culture media. Mortality data were recorded at 24-h
intervals up to 7 d. The LC50 and LC90 values were determined after 7 days.
 Effect of essential oil of Curcuma longa on oviposition of T. confusum.

Aliquots of 0.5 ml of various concentrations (0.092, 0.92, 2.75, 5.20,
6.42, and 8.26 mg/cm2) of C. longa rhizomes oil and the same concentrations
were used with gingerdione treatment, the different concentrations of the tested
materials were applied to pleated black filter papers (7.0 cm), dried for 30 min,
and placed in a glass bowl (7.0 cm diameter by 5 cm high).100 Twenty adult
insects were introduced on each treated filter paper. Some wheat flour was added
to the filter paper to provide food for the insects. Acetone-treated filter papers
were used as controls. Five replicates were prepared for each concentration and
control. The number of eggs laid and adult insects was recorded after 24 h.
80
EXPERIMENTAL
 Effect of essential oil of Curcuma longa on egg hatching and subsequent

larval and adult survival of T. confusum.
Eggs (0-24 h old) were collected on the pleated black filter papers by
placing each filter paper with 50 adult insects in a bowl (7 cm diameter by 5 cm
high) half-filled with wheat. There were 42-80 eggs per filter paper. These
papers were treated with 1.0-ml aliquots of essential oil of C. longa (0.18, 1.8,
5.4, 9.0, 12.6, and 16.2 mg/cm2) and allowed to air-dry for 1 h.
The treated filter papers were transferred to Petri dishes (7 cm diameter)
having 200 g of wheat flour and the mouth of dishes were sealed with black filter
papers using the wax method to prevent the insects crawling out. The number of
eggs that hatched was recorded and the number of live larvae was counted after 7
d. The number of adults that emerged from the live larvae was recorded until no
further adults had emerged. There were 10 replicates for each treatment
concentration and control.
 Progeny production of T. confusum in treated medium.

One hundred grams of wheat were sprayed with 3.0 ml of various
concentrations (0.18, 1.8, 5.4, 9.0, 12.6, and 16.2 mg/cm2) of C. longa rhizomes
oil and gingerdione each are separately. After thorough mixing and evaporation
of the solvent, 20 adults of T. confusum were added to 20 g treated wheat flour in
crystallizing dishes. The dishes were sealed with wax and kept in an incubator at
30 ºC and 65±70% RH without light. After 48 h, the adults were removed and
the number of dead adults was recorded. The dishes were placed back in the
incubator until F1 adults emerged. The F1 adults were counted and weighed. Ten
replicates were set up for each concentration and control.
81
EXPERIMENTAL
 Data analysis.
Probity analysis (Finney 1971) was used to analyze the dosage-mortality
data, using computerized programme (Ldp-line software) whereas the linear
regression analysis using SPSS (1999) was carried out.
82
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94
APPENDIX
APPENDIX 1: Spectra of compound 1
Fig.3 IR Spectrum of Compound 1
95
APPENDIX
Fig. 4: 1H NMR Spectrum of compound 1
96
APPENDIX
Fig.4a: Expanded 1H NMR Spectrum of compound 1
97
APPENDIX
Fig. 4b: Expanded 1H NMR Spectrum of compound 1
98
APPENDIX
Fig. 4c: Expanded 1H NMR Spectrum of compound 1
99
APPENDIX
Fig. 5 IR Spectrum of compound 2
100
APPENDIX
Fig. 6: 1H NMR Spectrum of compound 2
101
APPENDIX
Fig. 6a: Expanded 1H NMR Spectrum of compound 2
102
APPENDIX
Fig. 6b: Expanded 1H NMR Spectrum of compound 2
103
APPENDIX
APPENDIX 3: Spectra of compounds 3, 4
Fig. 7: 1H NMR Spectrum of compounds 3, 4
104
APPENDIX
Fig. 7a: Expanded 1H NMR Spectrum of compounds 3, 4
105
APPENDIX
Fig. 7b: Expanded 1H NMR Spectrum of compounds 3, 4
106
APPENDIX
Fig. 7c: Expanded 1H NMR spectrum of compounds 3, 4
107
APPENDIX
Fig. 8: IR Spectrum of compound 5
108
APPENDIX
Fig.9: Mass Spectrum of compound 5
109
APPENDIX
Fig. 10: 1H NMR of compound 5
110
APPENDIX
Fig. 10a: Expanded 1H NMR spectrum of compound 5
111
APPENDIX
Fig. 10b: Expanded 1H NMR spectrum of compound 5
112
APPENDIX
Fig. 10c: Expanded 1H NMR spectrum of compound 5
113
APPENDIX
Fig.11: IR spectrum of compound 6
114
APPENDIX
Fig. 12: 1H NMR spectrum of compound 6
115
APPENDIX
116
APPENDIX
117
APPENDIX
Fig. 13: UV Spectrum of compound 6
118
APPENDIX
Fig. 14: UV Spectrum of compound (6) after the addition of EtONa
119
APPENDIX
APPENDIX 6: Mass Spectra of compounds identified by GC/MS of

Cynanchum acutum CH2Cl2 fraction.
Fig. 15: Mass Spectrum of compound 7
Fig.17: Mass Spectrum of compound 9
120
APPENDIX
121
APPENDIX
122
APPENDIX
APPENDIX 7: 1H NMR Spectra of unidentified flavonoidal compound from

Ethyl acetate extract of Cynanchum acutum L.
Fig. 24: 1H NMR Spectrum of unidentified flavonoidal compound
123
APPENDIX
Appendix 8: Spectra of ar- turemerone 16
124
APPENDIX
Fig.25a: Expanded 1H NMR spectrum of compound 16
125
APPENDIX
Fig.25b: Expanded 1H NMR spectrum of compound 16
126
APPENDIX
Fig.25c: Expanded 1H NMR spectrum of compound 16
127
APPENDIX
Fig.26: Mass spectrum of compound 16
Appendix 9: Spectra of compound 17
128
APPENDIX
129
APPENDIX
130
APPENDIX
131
APPENDIX
132
APPENDIX
133
APPENDIX
134
APPENDIX
Fig.30b: Expanded 1H NMR spectrum of compound 20
135
APPENDIX
Fig.31: 1H NMR spectrum of compound 21
136
APPENDIX
Fig.31a: Expanded 1H NMR spectrum of compound 21
137
APPENDIX
138
APPENDIX
139
APPENDIX
Appendix 14: spectra of compound 22
Fig.32: 1H NMR spectrum of compound 22
140
APPENDIX
141
APPENDIX
Fig.32c: Expanded 1H NMR spectrum of compound 22
142
‫اﻟﻤﻠﺨﺺ اﻟﻌﺮﺑﻲ‬
‫ﻓﺼﻞ وﺗﻌﺮﯾﻒ ﻣﻨﺘﺠﺎت طﺒﯿﻌﯿﺔ ﻣﻦ ﺑﻌﺾ اﻟﻨﺒﺎﺗﺎت ذات‬

‫اﻷھﻤﯿﺔ اﻟﻄﺒﯿﺔ‬
‫ﻣﺎزاﻟﺖ اﻟﻨﺒﺎﺗﺎت واﺣﺪة ﻣﻦ أھﻢ ﻣﺼﺎدر اﻷدوﯾﺔ ﻋﻠﻰ اﻟﺮﻏﻢ ﻣﻦ اﻟﺘﻘﺪم اﻟﻜﺒﯿﺮ ﻓﻲ ﻣﺠﺎل اﻟﻌﻼج‬
‫ﺧﻼل اﻟﻘﺮن اﻟﺤﺎدي واﻟﻌﺸﺮون ﺣﯿﺚ ان ﻣﺎ ﯾﻘﺮب ﻣﻦ ﺳﺘﻮن ﺑﺎﻟﻤﺎﺋﺔ ﻣﻦ اﻷدوﯾﺔ واﻟﻤﺴﺘﺤﻀﺮات اﻟﻄﺒﯿﺔ‬
‫ذات أﺻﻞ ﻧﺒﺎﺗﻲ‪.‬‬
‫ﻟﺬا ﺟﺎء اﻟﻌﻤﻞ اﻟﻤﻮﺻﻮف ﻓﻲ ھﺬة اﻟﺮﺳﺎﻟﺔ ﺑﮭﺪف اﻟﺒﺤﺚ ﻋﻦ ﻣﻮاد طﺒﯿﻌﯿﺔ ﻣﻦ اﻟﻨﺒﺎﺗﺎت اﻟﻤﺘﺎﺣﺔ ﻓﻲ‬
‫ﻣﺼﺮ ﻛﻤﺼﺪر ﻟﻠﺪواء‪.‬‬
‫ﻟﺬﻟﻚ ﺗﻢ اﺧﺘﯿﺎر ﻧﺒﺎﺗﯿﻦ ﻟﻤﺰﯾﺪ ﻣﻦ اﻟﺪراﺳﺔ ھﻤﺎ ﺳﯿﻨﻨﻜﻢ اﻛﯿﻮﺗﻢ )ﻣﻦ اﻟﻌﺎﺋﻠﺔ اﻟﻌﺸﺎرﯾﺔ(‪ ،‬ﻛﺮﻛﻮﻣﺎ‬
‫ﻟﻮﻧﺠﺎ )ﻣﻦ اﻟﻌﺎﺋﻠﺔ اﻟﺰﻧﺠﺒﯿﻠﯿﺔ(‪.‬‬
‫وﻓﻲ ھﺬة اﻟﺪراﺳﺔ ﺗﻢ اﻟﺤﺼﻮل ﻋﻠﻲ ﻣﻌﻠﻮﻣﺎت ﻣﻔﯿﺪة ﻛﻨﺘﯿﺠﺔ ﻟﺘﻄﺒﯿﻖ طﺮق ﻓﺼﻞ ﻛﺮوﻣﺎﺗﻮﺟﺮاﻓﯿﺔ‬
‫‪1‬‬
‫ﻋﺪﯾﺪة‪ ،‬ﻣﺜﻞ أﻋﻤﺪة وطﺒﻘﺎت اﻟﻔﺼﻞ اﻟﻜﺮوﻣﺎﺗﻮﺟﺮاﻓﯿﺔ‪ ،‬وطﺮق طﯿﻔﯿﺔ ‪ ،‬ﻣﺜﻞ ‪ MS‬و ‪H NMR GC/MS‬‬
‫و ‪ UV‬و ‪.IR‬‬
‫اﻟﻨﺒﺎت اﻷول ‪ :‬ﻧﺘﺞ ﻋﻦ دراﺳﺔ ﻧﺒﺎت ﺳﯿﻨﻨﻜﻢ اﻛﯿﻮﺗﻢ ﻓﺼﻞ وﺗﻌﺮﯾﻒ ﺧﻤﺴﺔ ﻋﺸﺮ ﻣﺮﻛﺒﺎ ﻣﻌﻈﻤﮭﺎ‬
‫ﻣﻌﺮوف واﻟﺘﻲ ﯾﻤﻜﻦ ﺗﺼﻨﯿﻔﮭﺎ اﻟﻲ اﺛﻨﯿﻦ ﻣﻦ اﻟﺘﺮﺑﻨﺎت اﻟﺜﻼﺛﯿﺔ ‪ 2-1‬واﺛﻨﯿﻦ اﺳﺘﯿﺮوﯾﺪات ‪ 4-3‬و اﺛﻨﯿﻦ‬
‫ﻛﻮﻣﺎرﯾﻨﺎت ‪ 6-5‬و اﺛﻨﯿﻦ ﺷﯿﻜﯿﻤﺎت ‪ 8-7‬و ﺛﻼﺛﺔ ﺗﺮﺑﯿﻨﺎت اﺣﺎدﯾﺔ ‪ 11-9‬و ارﺑﻊ أﺣﻤﺎض دھﻨﺒﺔ ‪ .15-12‬ھﺬا‬
‫ھﻮ أول ﺗﻘﺮﯾﺮ ﯾﺪل ﻋﻠﻰ وﺟﻮد ﻛﻮﻣﺎرﯾﻨﺎت ﯾﺘﻢ ﻓﺼﻠﮭﺎ وﺗﻌﺮﯾﻔﮭﺎ ﻣﻦ ﺟﻨﺲ اﻟﺴﯿﻨﻨﻜﻢ‪.‬‬
‫اﻟﻨﺒﺎت اﻟﺜﺎﻧﻲ‪ :‬ﻧﺘﺞ ﻋﻦ دراﺳﺔ ﻧﺒﺎت ﻛﺮﻛﻮﻣﺎ ﻟﻮﻧﺠﺎ ﻓﺼﻞ وﺗﻌﺮﯾﻒ ﺳﺒﻌﺔ ﻣﺮﻛﺒﺎت واﻟﺘﻲ ﯾﻤﻜﻦ‬
‫ﺗﺼﻨﯿﻔﮭﺎ اﻟﻲ ﺛﻼﺛﺔ ﺗﺮﺑﯿﻨﺎت ﻧﺼﻒ ﺛﻼﺛﯿﺔ ‪ 18-16‬و ﻣﺮﻛﺐ واﺣﺪ ﺟﯿﻨﺠﺮول ‪ 19‬و ﺛﻼﺛﺔ ﻣﺸﺘﻘﺎت داي ﻓﯿﻨﯿﻞ‬
‫ھﺒﺘﺎﻧﻮﯾﺪ ‪ .22-20‬ھﺬا ھﻮ أول ﺗﻘﺮﯾﺮ ﯾﺪل ﻋﻠﻰ وﺟﻮد ﺟﯿﻨﺠﺮول ﻣﻦ ﻧﺒﺎت ﻛﺮﻛﻮﻣﺎ ﻟﻮﻧﺠﺎ‪.‬‬
‫ﺗﻢ اﺧﺘﺒﺎر ﺧﻼﺻﺎت اﻹﺛﯿﺮ اﻟﺒﺘﺮوﻟﻲ و اﻟﻜﻠﻮروﻓﻮرم ﻟﻨﺒﺎت ﻛﺮﻛﻮﻣﺎ ﻟﻮﻧﺠﺎ ﻛﻤﺒﯿﺪات ﺣﺸﺮﯾﺔ ﺿﺪ‬
‫ﺣﺸﺮة ﺳﻮﺳﺔ اﻟﺪﻗﯿﻖ ﺗﺮﺑﻮﻟﯿﻢ ﻛﻮﻧﻔﯿﻮزم ﻓﺎﺗﻀﺢ ﻓﻐﺎﻟﯿﺔ ﺧﻼﺻﺔ اﻹﯾﺜﯿﺮ اﻟﺒﺘﺮوﻟﻲ ﺑﯿﻨﻤﺎ ﻋﺪم ﻓﻌﺎﻟﯿﺔ ﺧﻼﺻﺔ‬
‫اﻟﻜﻠﻮروﻓﻮرم ‪.‬‬
‫ﻟﻘﺪ أوﺿﺤﺖ اﻟﻨﺘﺎﺋﺞ اﻟﺤﯿﻮﯾﺔ ان ﻛﻞ ﻣﻦ اﻟﺰﯾﺖ اﻟﻄﯿﺎر واﻟﻤﺮﻛﺐ ‪ 19‬اﻟﻤﻔﺼﻮﻟﺔ ﻣﻦ ﺧﻼﺻﺔ‬
‫اﻹﺛﯿﺮ اﻟﺒﺘﺮوﻟﻲ ﻟﻨﺒﺎت ﻛﺮﻛﻮﻣﺎ ﻟﻮﻧﺠﺎ ﻟﮭﺎ ﻓﺎﻋﻠﯿﺔ ﻋﺎﻟﯿﺔ ﻛﻤﺒﯿﺪ ﺣﺸﺮي ﺿﺪ ﺣﺸﺮة ﺳﻮﺳﺔ اﻟﺪﻗﯿﻖ ﺗﺮﺑﻮﻟﯿﻢ‬
‫ﻛﻮﻧﻔﯿﻮزم‪.‬‬
‫‪1‬‬
29 29
20 20
30 20 30 20
27 19 27 19
12 22 12 22
18
25 11
18
25 11 13 13
1
17 28 1
17 28
14 14
2 9 16 2 9 16
10 8 15 10 8 15
O 26 26
4
O 5
6
7
1 HO 3 4 5
6
7
2
H3C 23 24 23
29 24 29
28 28
21 22 22
20 24 27 21 24 27
20
19 19
25 25
12 23 12 23
17 17
18 11 26 18 11 26
13 13
1 16 1 16
2 9 14 2 9
10 15 10 14 15
8 8
HO 3 5
6
7 HO 3 5 7
4 6
4
3 4
O OH
HO O O
5 4 5 4
O 6 O
3 6
3
O 7 O 2 O 7 2
8 1
HO 8
O O
1
CHO H O
5 6 7 8
H OH
O O
O HO O
O
9 10 11
2
H OH
HO
O 12 O O 13
O O HO
14 15
12
H H
3 O
5 4
2
8 3'
O 7 1 O
2'
10 9 11
H
16 17 18
H
O O
2' 10 4 2
MeO 3' 1'
8 6
9 7 5 3 1
HO 4' 6'
19
5'
H
O O
6 4 4' 6'
MeO 7 5 5' 7' OMe
2 2'
3 1 3'
H
HO 8 10 20 10' 8' OH
9 9'
H
O O
6 4 4'
6'
MeO 7 5 5' 7'
2 2'
3 1 3'
H
HO 8 10 21 10' 8' OH
9 9'
H
O O
6 4 4' 6'
7 5 5' 7'
2 2'
3 1 3'
H
HO 8 10 22 10' 8' OH
9 9'
3
‫ﻛﻠﯿﺔ اﻟﻌﻠﻮم‬
‫ﻗﺴﻢ اﻟﻜﯿﻤﯿﺎء‬
‫ﻓﺼﻞ وﺗﻌﺮﯾﻒ ﻣﻨﺘﺠﺎت طﺒﯿﻌﯿﺔ ﻣﻦ ﺑﻌﺾ اﻟﻨﺒﺎﺗﺎت ذات اﻷھﻤﯿﺔ اﻟﻄﺒﯿﺔ‬

‫رﺳﺎﻟﺔ ﻣﻘﺪﻣﺔ‬
‫ﻟﻠﺤﺼﻮل ﻋﻠﻲ درﺟﺔ اﻟﻤﺎﺟﺴﺘﯿﺮ ﻓﻰ اﻟﻌﻠﻮم‪ -‬ﻛﯿﻤﯿﺎء‬
‫)ﻛﯿﻤﯿﺎء ﻋﻀﻮﯾﺔ(‬
‫ﻣﻦ‬
‫ﻋﻣرو اﻟﺳﯾد أﺣﻣد اﻟدﻣرداش‬

‫اﻟﻤﻌﯿﺪ‬
‫ﺑﻘﺴﻢ اﻟﻜﯿﻤﯿﺎء‪ -‬ﻛﻠﯿﺔ اﻟﻌﻠﻮم – ﺟﺎﻣﻌﺔ اﻟﻤﻨﺼﻮرة‬
‫ﺑﻜﺎﻟﻮرﯾﻮس ﻋﻠﻮم )ﻛﯿﻤﯿﺎء(‪ -‬ﺟﺎﻣﻌﺔ اﻟﻤﻨﺼﻮرة )‪(2004‬‬
‫اﺷﺮاف‬
‫اﻷﺳﺗﺎذ اﻟدﻛﺗور‪ /‬ﻣﻣدوح ﻋﺑد اﻟﻣﺟﯾب ﻣﺣﻣد‬

‫أﺳﺗﺎذ ﻛﯾﻣﯾﺎء اﻟﻣﻧﺗﺟﺎت اﻟطﺑﯾﻌﯾﺔ‬
‫ﻗﺳم اﻟﻛﯾﻣﯾﺎء – ﻛﻠﯾﺔ اﻟﻌﻠوم – ﺟﺎﻣﻌﺔ اﻟﻣﻧﺻورة‪.‬‬
‫اﻷﺳﺗﺎذ اﻟدﻛﺗور‪ /‬ﻋﺑد اﻟﻌزﯾز ﻣﺣﻣود دوﯾدار‬

‫أﺳﺗﺎذ اﻟﻛﯾﻣﯾﺎء اﻟﻌﺿوﯾﺔ اﻟﻐﯾر ﻣﺗﻔرغ‬
‫اﻟدﻛﺗورة‪ /‬اﯾﻣﺎن ﻣﺣﻣد ﻣﺣﻣود ﻛﺷك‬

‫أﺳﺗﺎذ ﻣﺳﺎﻋد اﻟﻛﯾﻣﯾﺎء اﻟﻌﺿوﯾﺔ‬
‫‪2009‬‬
‫ﻧﻣوذج رﻗم )‪(1‬‬
‫ﺻﻔﺣﺔ اﻟﻣﺷرﻓون‬
‫ﻋﻧوان اﻟرﺳﺎﻟﺔ‪ :‬ﻓﺼﻞ وﺗﻌﺮﯾﻒ ﻣﻨﺘﺠﺎت طﺒﯿﻌﯿﺔ ﻣﻦ ﺑﻌﺾ اﻟﻨﺒﺎﺗﺎت ذات اﻷھﻤﯿﺔ اﻟﻄﺒﯿﺔ‬
‫اﺳﻢ اﻟﺒﺎﺣﺚ‪ :‬ﻋﻤﺮو اﻟﺴﯿﺪ أﺣﻤﺪ اﻟﺪﻣﺮداش‬
‫اﺷﺮاف‪:‬‬
‫اﻟﺗوﻗﯾﻊ‬ ‫اﻟوظﯾﻔﺔ‬ ‫اﻻﺳم‬ ‫م‬

‫‪ 1‬ا‪.‬د‪ .‬ﻣﻤﺪوح ﻋﺒﺪ اﻟﻤﺠﯿﺐ ﻣﺤﻤﺪ أﺳﺗﺎذ ﻛﯾﻣﯾﺎء اﻟﻣﻧﺗﺟﺎت اﻟطﺑﯾﻌﯾﺔ ﺑﻛﻠﯾـﺔ اﻟﻌﻠـوم‪ -‬ﺟﺎﻣﻌـﺔ‬
‫اﻟﻣﻧﺻورة‬
‫‪ 2‬ا‪.‬د‪ .‬ﻋﺒﺪ اﻟﻌﺰﯾﺰ ﻣﺤﻤﻮد دوﯾﺪار أﺳﺗﺎذ اﻟﻛﯾﻣﯾﺎء اﻟﻌﺿوﯾﺔ اﻟﻐﯾرﻣﺗﻔرغ ﺑﻛﻠﯾﺔ اﻟﻌﻠوم‪-‬‬
‫ﺟﺎﻣﻌﺔ اﻟﻣﻧﺻورة‬
‫أﺳﺗﺎذ ﻣﺳﺎﻋد اﻟﻛﯾﻣﯾﺎء اﻟﻌﺿوﯾﺔ ﺑﻛﻠﯾﺔ اﻟﻌﻠوم‪ -‬ﺟﺎﻣﻌﺔ‬ ‫‪ 3‬د‪ .‬اﯾﻤﺎن ﻣﺤﻤﺪ ﻣﺤﻤﻮد ﻛﺸﻚ‬
‫اﻟﻣﻧﺻورة‬
‫ﻋﻣﯾد اﻟﻛﻠﯾﺔ‬ ‫وﻛﯾل اﻟﻛﻠﯾﺔ ﻟﻠدراﺳﺎت اﻟﻌﻠﯾﺎ‬ ‫رﺋﯾس اﻟﻘﺳم‬
‫أ‪.‬د‪ .‬طﮫ زﻛﻲ ﺳﻛر‬ ‫أ‪.‬د‪ .‬اﻟﻣﺗوﻟﻲ ﻣﺣﻣد اﻟﻌﺑﺎﺳﻲ‬ ‫أ‪.‬د‪ .‬ﻋوض اﺑراھﯾم أﺣﻣد‬
‫ﻧﻣوذج رﻗم )‪(2‬‬
‫ﺻﻔﺣﺔاﻟﺳﺎدة أﻋﺿﺎء ﻟﺟﻧﺔ اﻟﻣﻧﺎﻗﺷﺔ واﻟﺣﻛم‬
‫ﻋﻧوان اﻟرﺳﺎﻟﺔ‪ :‬ﻓﺼﻞ وﺗﻌﺮﯾﻒ ﻣﻨﺘﺠﺎت طﺒﯿﻌﯿﺔ ﻣﻦ ﺑﻌﺾ اﻟﻨﺒﺎﺗﺎت ذات اﻷھﻤﯿﺔ اﻟﻄﺒﯿﺔ‬
‫اﺳﻢ اﻟﺒﺎﺣﺚ‪ :‬ﻋﻤﺮو اﻟﺴﯿﺪ أﺣﻤﺪ اﻟﺪﻣﺮداش‬
‫ﻟﺠﻨﺔ اﻹﺷﺮاف‪:‬‬

‫وم‪-‬‬ ‫‪ 1‬ا‪.‬د‪ .‬ﻣﻤﺪوح ﻋﺒﺪ اﻟﻤﺠﯿﺐ ﻣﺤﻤﺪأﺳ‬
‫ﻛﯾﻣﯾ ﺗﺎذﺎء اﻟﻣﻧﺗﺟ ﺎت اﻟطﺑﯾﻌﯾﺑﻛﻠﯾ ﺔ ﺔ اﻟﻌﻠ‬
‫‪ 2‬ا‪.‬د‪ .‬ﻋﺒﺪ اﻟﻌﺰﯾﺰ ﻣﺤﻤﻮد دوﯾﺪار أﺳﺗﺎذ اﻟﻛﯾﻣﯾﺎء اﻟﻌﺿوﯾﺔ اﻟﻐﯾرﻣﺗﻔرغ ﺑﻛﻠﯾﺔ‬
‫اﻟﻌﻠوم‪ -‬ﺟﺎﻣﻌﺔ اﻟﻣﻧﺻورة‬
‫أﺳﺗﺎذ ﻣﺳﺎﻋد اﻟﻛﯾﻣﯾﺎء اﻟﻌﺿوﯾﺔ ﺑﻛﻠﯾﺔ اﻟﻌﻠوم‪-‬‬ ‫‪ 3‬د‪ .‬اﯾﻤﺎن ﻣﺤﻤﺪ ﻣﺤﻤﻮد ﻛﺸﻚ‬
‫ﻟﺟﻧﺔ اﻟﻣﻧﺎﻗﺷﺔ واﻟﺣﻛم‪:‬‬

‫أﺳﺗﺎذ اﻟﻌﻘﺎﻗﯾر ﺑﻛﻠﯾﺔ اﻟﺻﯾدﻟﺔ‪ -‬ﺟﺎﻣﻌﺔ اﻟﻘﺎھرة‬ ‫ا‪.‬د‪ .‬ﻋﻠﻰ ﻣﺣﻣد اﻟﺷﺎﻣﻲ‬ ‫‪1‬‬
‫أﺳﺗﺎذ اﻟﻛﯾﻣﯾﺎء اﻟﻌﺿوﯾﺔ ﺑﻛﻠﯾﺔ اﻟﻌﻠوم‪ -‬ﺟﺎﻣﻌﺔ‬ ‫ا‪.‬د‪ .‬ﻋﺑد اﻟﺟواد ﻋﻠﻰ ﻓﮭﻣﻲ‬ ‫‪2‬‬
‫اﻟﻘﺎھرة‬
‫ا‪.‬د‪ .‬ﻋﺒﺪ اﻟﻌﺰﯾﺰ ﻣﺤﻤﻮد دوﯾﺪار أﺳﺗﺎذ اﻟﻛﯾﻣﯾﺎء اﻟﻌﺿوﯾﺔ اﻟﻐﯾرﻣﺗﻔرغ ﺑﻛﻠﯾﺔ‬ ‫‪3‬‬
‫اﻟﻌﻠوم‪ -‬ﺟﺎﻣﻌﺔ اﻟﻣﻧﺻورة‬
‫ا‪.‬د‪ .‬ﻣﻤﺪوح ﻋﺒﺪ اﻟﻤﺠﯿﺐ ﻣﺤﻤﺪ أﺳﺗﺎذ ﻛﯾﻣﯾﺎء اﻟﻣﻧﺗﺟﺎت اﻟطﺑﯾﻌﯾﺔ ﺑﻛﻠﯾﺔ اﻟﻌﻠوم‪-‬‬ ‫‪4‬‬
‫ﻋﻣﯾد اﻟﻛﻠﯾﺔ‬ ‫وﻛﯾل اﻟﻛﻠﯾﺔ ﻟﻠدراﺳﺎت اﻟﻌﻠﯾﺎ‬ ‫رﺋﯾس اﻟﻘﺳم‬
‫أ‪.‬د‪ .‬طﮫ زﻛﻲ ﺳﻛر‬ ‫أ‪.‬د‪ .‬اﻟﻣﺗوﻟﻲ ﻣﺣﻣد اﻟﻌﺑﺎﺳﻲ‬ ‫أ‪.‬د‪ .‬ﻋوض اﺑراھﯾم أﺣﻣد‬
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Natural Products Chemistry: Isolation and Structure Elucidation of Natural

Thesis · September 2009

Microbial Natural Proucts View project

Bioactive Plant-Based Natural Products View project

The user has requested enhancement of the downloaded file.

Isolation and Structure Elucidation of Natural Products from

Submitted to the Faculty of Science, Mansoura University

Amr El Sayed Ahmed El Demerdash

B. Sc. (Chemistry), Mansoura University (2004)

Prof. Dr. M. Abdel-Mogib Prof. Dr. A. M. Dawidar

Title of the thesis: Isolation and Structure Elucidation of Natural

Prof. of Organic Chemistry

Head of Chemistry Vice-Dean of Faculty of Science Dean of Faculty

Prof. Dr. A. I. Ahmed Prof. Dr. El-Metwally M. Prof. Dr. Taha Z.

Student name: Amr El Sayed Ahmed El Demerdash

Discussion and judgment team

Head of Vice-Dean of Faculty of Science Dean of Faculty of

Prof. Dr. A. I. Ahmed Prof. Dr. El-Metwally M. Prof. Dr. Taha Z.

2- Advanced Physical organic Chemistry

7- Natural products Chemistry

10- Polymer Chemistry

11- Heterocyclic Chemistry

12- Enviromental and Analytical Chemistry

13- Molecular rearrangements

He had successfully passed the final examination of these courses in Jan.

Prof. Dr. A. I. Ahmed

I wish to express my sincere appreciation and gratitude to

I would like to express my cordial thanks and gratitude to

I would like to express my deep appreciation and gratitude to

Deepest appreciation and thanks to Dr. Z. Abou El- Naga,

Finally, I would like to thank all the stuff members of

1.2. Cynanchum acutum L. 1

1.2.1. Botanical aspects, description and distribution in Egypt. 1

1.2.2. Medicinal uses of some Cynanchum species. 3

1.2.3. Medicinal and biological activities of Cynanchum acutum L. 3

1.2.4. Phytochemical constituents of Cynanchum genus. 3

1.2.5. Chemical constituents of Cynanchum acutum L. 11

1.3. Curcuma longa L. 12

1.3.1. Botanical aspects, description and distribution. 12

1.3.2. History and medicinal uses. 13

1.3.3. Biological activities of Curcuma longa L. 14

1.3.4. Phytochemical constituents of Curcuma longa L. 20

1.3.4.1. Constituents of volatile oil of Curcuma longa L. 20

II- Aim of the Work. 26

III- Results and discussion. 27

2.1. Processing of Cynanchum acutum L. aerial parts. 27

2.1.1. Study of petroleum ether extract. 27

2.1.2. Characterizations of compound 1. 27

2.1.3. Characterizations of compound 2. 30

2.1.5. Structure elucidation of compound 4. 33

2.2. Study of the methylene chloride extract. 35

2.2.1. Structure elucidation of compound 5. 35

2.2.2. Structure elucidation of compound 6. 37

2.2.3. Study of methylene chloride extract using GC/MS. 38

2.3. Study of Ethyl acetate extract. 47

3.1 Processing of Curcuma longa L powdered rhizomes. 48

3.2. Study of petroleum ether extract of Curcuma longa L. 49

3.2.1. Structure elucidation of sesquiterpene ketone 16. 49

3.2.2. Structure elucidation of sesquiterpene ketone 17. 52

3.2.3. Structure elucidation of compound 18. 53

3.2.4. Structure elucidation of compound 19. 54