Phytochemical Screening & in vitro Antioxidant and Thrombolytic Activities of Argemone mexicana ...

89 Pages·2012·1.59 MB·English

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“Phytochemical Screening & in vitro Antioxidant and Thrombolytic
Activities of Argemone mexicana Extracts”

Submitted By
Ahamed Hasan Al-Baizyd
ID: 2010-3-79-013

In Partial Fulfillment of the Requirements for the Award of the Degree
Bachelor of Pharmacy

Under the Guidance of
MR. APURBA SARKER APU
Senior Lecturer

Department of Pharmacy
East West University
Dhaka, Bangladesh.

July, 2012
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DECLARATION BY THE CANDIDATE

I hereby declare that this dissertation entitled “Phytochemical Screening & in vitro
Antioxidant and Thrombolytic Activities of Argemone mexicana Extracts” is an
authentic and genuine research work carried out by me under the guidance of Mr.
Apurba Sarker Apu, Senior lecturer, Department of Pharmacy, East West University,
Dhaka, Bangladesh.

__________________________
Ahamed Hasan Al-Baizyd
ID: 2010-3-79-013
Department of Pharmacy
East West University,
Dhaka, Bangladesh.
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CERTIFICATE BY THE SUPERVISOR

This is to certify that the dissertation entitled “Phytochemical Screening & in vitro Antioxidant
and Thrombolytic Activities of Argemone mexicana Extracts” is a bonafide research work
done by Ahamed Hasan Al-Baizyd, in partial fulfillment of the requirement for the Degree of
Bachelor of Pharmacy.

__________________________________
MR. APURBA SARKER APU
Senior Lecturer
Department of Pharmacy
East West University
Dhaka, Bangladesh.
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ENDORSEMENT BY CHAIRPERSON

This is to certify that the dissertation entitled “Phytochemical Screening & in vitro Antioxidant
and Thrombolytic Activities of Argemone mexicana Extracts” is a bonafide research work
done by Ahamed Hasan Al-Baizyd under the guidance of Mr. Apurba Sarker Apu, Senior
Lecturer, Department of Pharmacy, East West University, Dhaka.

____________________________
DR. SUFIA ISLAM
Associate Professor and Chairperson
Department of Pharmacy
East West University
Dhaka, Bangladesh.
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ACKNOWLEGDEMENTS
By the grace of Allah, I have completed this intellectual academic attempt. I would like to
express my thanks to my research supervisor Mr. Apurba Sarker Apu, Senior Lecturer,
Department of Pharmacy, EWU, who had been always optimistic and full of enthusiasm and
ideas. His generous advice, encouragement, feedback and reminders have not only helped
shape this study but also molded me into being a better researcher. His in-depth thinking
and motivation to guide had provided me with the motivation to continue and complete this
research.
I am gratefully acknowledged to Chairperson Professor Dr. Sufia Islam Associate professor
& Chairperson, Department of Pharmacy, EWU for her inspiration in my study Moreover, I
am grateful to my administration as they provide the facilities to use the laboratory for
research work.
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INDEX

Contents Page no.

List of Tables
List of Figures
Abstract 10
Chapter 1: Introduction 12-13
1.1 Argemone Mexicana 13
1.1.1 Common names 15
1.1.2 Scientific classification 15
1.1.3 Plant description 15
1.1.4 Growth and development 16
1.1.5 Properties of Argemone Mexicana 17
1.1.6 M edicinal uses 18
1.2 Solvent System 20
1.3 Phytochemical Screening 23
1.4 Antioxidants 24
1.4.1 DPPH free radical scavenging assay 25

1.4.2 Nitric Oxide Scavenging Capacity Assay 25
1.5 Total phenolic content 26
1.6 Thrombolytic Activity 26
1.7 Purpose of the Present Study 28

Chapter 2: Literature Review of Argemone mexicana
2.1 Phytochemistry of Argemone mexicana Linn 31
2.2 Pharmacological studies on Argemone mexicana 36

Chapter 3: Method and materials
3.1 Plant extraction 45
3.1.1 Plant collection and identification 45
3.1.2 Chopping, drying and grinding of the plants 46
3.1.3 Selection of Solvent 46
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3.1.4 Methanolic extraction process of Argemone Mexicana 47
3.1.5 Ethyl acetate extraction process of Argemone Mexicana 49
3.1.6 n-hexane extraction process of Argemone mexicana: 49
3.1.7 Apparatus & reagent used for plant extraction 50
3.2 Screening of Phytochemical Constituents 51
3.3 DPPH Free Radical Scavenging Test 56
3.4 Nitric Oxide Scavenging Capacity Assay 57
3.5 Total phenolic content 58
3.6 In Vitro Thrombolytic Activity Test 59

Chapter 4: Result and discussion
4.1 Screening of Phytochemical Constituents 63
4.2 DPPH Free Radical Scavenging Test 64
4.3 Nitric Oxide Scavenging Capacity Assay 70
4.4 Total phenolic content 77
4.5 In Vitro Thrombolytic Activity Test 79

Chapter 5: Conclusion 82

References 83
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LIST OF TABLES
Title Page no.

Table1.1: Different parts of Argemone mexicana in medicinal uses. 19
Table 1.2 Properties of the solvents 22
Table2.1: Summary of the Phytochemical Studies on Argemone mexicana 36
Table2.2: Summary of the pharmacological Studies on Argemone mexicana 41
Table3.1 Properties of the solvents 47
Table3.2 Time and temperature of extraction 47
Table4.1 Data of phytochemical screening of A. mexicana 63
Table4.2 IC50 value of ascorbic acid (AA) 64
Table4.3 IC50 value of the leaf of AMHE 65
Table4.4 IC50 value of the leaf of AMEA 66
Table4.5 IC50 value of the leaf of AMME 68
Table4.6 % inhibition of AMHE, AMEA, AMME and AA 69
Table4.7 IC50 value of ascorbic acid (AA) 74
Table4.8 IC50 value of the leaf of AMEA 76
Table4.9 IC50 value of the leaf of AMME 77
Table4.10 Absorbance of gallic acid 77
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LIST OF FIGURES

Title Page no.

Figure1.1: An antique botanical prints of Argemone mexicana 14
Figure1.2 Different parts of Argemone Mexicana 16
Figure1.3 Mechanism of anticoagulation of streptokinase 28
Figure2.1: Structure of luteolin 31
Figure2.2: Structure of triacontan-11-ol 32
Figure2.3: Structure of jatrorrhizine 33
Figure2.4: Structure of columbamine 33
Figure2.5: Structure of oxyberberine 34
Figure2.6: Graphical view of CYP719A13 participates in both sanguinarine and 35
berberine biosynthesis. Prepared from 2-month-old plantlets of the
Mexican prickly poppy Argemone mexicana
Figure3.1: The identification plate of Argemone mexicana. 45
Figure3.2: Cutting and drying of Argemone mexicana. 46
Figure3.2: Soxhlet apparatus 48
Figure3.3 Extracts (before evaporation) 48
Figure3.4 Rotary Evaporator (IKA ®RV05 Basic, Biometra, Germany) 49
Figure3.5 Crude extracts of Argemone mexicana 50
Figure3.6 Identification of alkaloids by Hager’s reagent 52
Figure 4.1 Clot lysis of blood samples of normal subjects by APHE, APEA, and 79
APME extracts of A. mexicanaleaves, streptokinase and normal
saline
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ABSTRACT
Purpose: In present study the leaves extracts of Argemone mexicana (Papaveraceae) have been
screened for their photochemical constituents, antioxidant, total phenolic content and in vitro
thrombolytic activities.
Methods: The n-hexane, ethyl acetate and methanol extracts were screened for the presence of
phytochemicals, using in vitro assay methods and their inhibition of 2,2-diphenyl-1-
picrylhydrazyl (DPPH) free radical was used to evaluate their free radical scavenging activity
at 517 nm. Nitric oxide scavenging capacity was measured by sodium nitroprusside and griss
reagent. The total phenolic content (TPC) measured by the Folin-Ciocalteu method. An in vitro
thrombolytic model was used to check out the clot lysis effects of three extracts using
streptokinase as a positive control.
Results: Phytochemical screening of the plant showed the presence of alkaloids, flavonoids,
terpenoids, saponins, tannins. All the extracts inhibited DPPH, indicating their antioxidant
activity. In DPPH and nitric oxide scavenging assay both n-hexane extract showed highest IC
50
values 73.73 μg/ml, 152.73μg/ml and the lowest IC value was 12.54 μg/ml of ethyl acetate
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extract and 65.56μg/ml of methanol extract. It appears that, it has greater free radical
scavenging capacity. Ethyl acetate extract had the highest total phenolic content of 106.65 mg
GAE/100 g extract. All the extracts showed significant % of clot lysis effect (p<0.001) with
reference to negative control.
Conclusion: The outcomes of the study are an indication of present of phytochemicals and
may be responsible for some of the therapeutic uses of these plants.
Keywords: Argemone mexicana, Phytochemical screening, DPPH, Nitric oxide, Phenolic
content, In vitro Thrombolytic.