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UNIVERSITI PUTRA MALAYSIA INSULINOTROPIC PROPERTIES OF SEVERAL Ganoderma Extract

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					             UNIVERSITI PUTRA MALAYSIA


INSULINOTROPIC PROPERTIES OF SEVERAL MALAYSIAN HERBS
               AND GANODERMA SPECIES




                MOHD SAUFI BASTAMI.


                     FBSB 2006 2
     INSULINOTROPIC PROPERTIES OF SEVERAL MALAYSIAN
              HERBS AND GANODERMA SPECIES




                                     BY


                         MOHD SAUFI BASTAMI




Thesis submitted to School of Graduate Studies, Universiti Putra Malaysia, in
     fulfilment of the Requirements for the Degree of Master of Science


                               January 2006
                                                                                    of
Abstract of thesis presented to the Senate of Universiti Putra Malaysia in f~dfilment the
                      requirement for the degree of Master of Science


  INSULINOTROPIC PROPERTIES OF SEVERAL MALAYSIAN HERBS AND
                                 GANODERMA SPECIES


                                             BY
                                MOHD SAUFI BASTAMI
                                       January 2006


Chairman       : Muhajir Hamid, PhD
Faculty        : Biotechnology and Biomolecular Sciences


Diabetes Mellitus incident is currently one of the major and common diseases in
Malaysia and increasing every year. To date, no herbs or drugs are able to completely
cure the disease. In this study, insulinotropic properties of Malaysian herbs were
investigated using pancreatic   P cells BRIN-BD1 1. Three out of   13 herbs samples studied
were belongs to fungi (Ganoderma sp). The studied showed seven samples are able to
induce insulin secretion over 200 % by BRIN-BD11 cell lines. Based on traditional
practitioner reports, Gynuraprocumbens was choosed for further investigation on its anti-
hyperglycemic properties. Glucose tolerance test were carried on normal and
streptozotocin-induced diabetic rats showed at 1000 mglkgl b.w Gynura procumbens
methanolic extract was able to reduce blood glucose level in rats. Toxicity towards
BRIN-BD11 cells was analyzed using MTT assay and alkaline comet assay. Toxicity
levels at ICS0was 300 uglml using MTT assay. Furthermore, Alkaline Comet Assay has
proved this toxicity is not due to DNA damage in BRIN-BDl 1 cells. Acute toxicity was
carried on ICR strain mice for 14 days. Gynuraprocumbens did not show any toxicity at
concentration 7 glkg b.w after administered orally. These results suggest Gynura
procumbens and other herbs contain active constituents and have potentials for diabetes
treatment.
    Abstrak tesis yang dikemukakan kepada Senat Universiti Putra Malaysia sebagai
                    memenuhi keperluan untuk ijazah Master Sains

CIRI-CIIU INSULINOTROPIK BEBERAPA HERBA MALAYSIA DAN SPESIES
                                   GANODERMA




                                         Oleh
                             MOHD SAUFI BASTAMI
                                    Januari 2006




Pengerusi     : Muhajir Hamid, PhD
Fakulti       : Bioteknologi dan Sains Biomolekul



Kes penyakit Diabetis Mellitus atau Kencing Manis kini menjadi penyakit utama
dihadapi oleh kebanyakkan pesakit manakala bilangannya semakin meningkat setiap
tahun di Malaysia. Sehingga kini masih belum ada ubat atau herba yang dapat
memulihkan daripada penyakit ini sepenuhnya. Dalam kajian ini, ciri-ciri insulinotropik
dari tumbuhan herba Malaysia di jalankan bagi melihat kebolehan tumbuhan herba
merangsang penghasilan insulin oleh sel selanjar model sel    P   pankreas BRIN-BDll.
Sejumlah tiga dari 13 sampel yang di kaji terdiri dari golongan kulat (Ganoderma sp).
Kajian mendapati tujuh daripada sample kajian berupaya merangsang penghasilan insulin
lebih dari 200 % oleh sel BRIN-BD1 1. Gynura procumbens dipilih berdasarkan laporan
pengamal perubatann tradisional untuk ujian lanjut ciri-ciri anti hyperglycemia.
Seterusnya, kajian toleransi glukosa ekstrak methanol Gynura procumbens terhadap tikus
normal dan tikus diabetik yang di aruh oleh streptozotocin mendapati 1000 mglkg b.w
ekstrak berupaya menurunkan paras glukosa darah. Kajian toksisiti terhadap sel BRIN-
BDl l mengunakan asai MTT dan Asai Komet Beralkali dijalankan. Asai MTT
memberikan keputusan paras toksisiti Gynura procumbens pada ICsoadalah 300 uglml.
Walaubagaimanapun, Asai Komet Beralkali membuktikan toksisiti ini berlaku bukan
disebabkan oleh pemusnahan DNA di dalam sel BRIN-BD11. Tahap toksisiti akut
dijalankan kepada mencit dari strain ICR untuk tempoh 14 hari. Gynura procumbens
didapati tidak memberi kesan toksik pada kepekatan 7 glkg b.w selepas ekstrak di
masukkan secara oral. Keputusan ini menunjukkan Gynura procumbens dan tumbuhan
herba yang lain mengandungi komponen aktif dan berpotensi dalam rawatan kencing
manis.
                                ACKNOWLEDGEMENTS



I would like to express my sincere thanks and appreciation to my supervisor Dr. Muhajir

Hamid for his advice, support, encouragement and his patience. I am especially grateful

to him for his intellectual guidance and constructive comments throughout the course of

this study and during the preparation of the manuscript and for being so freely available

at all stages for discussion.



Sincere appreciations are extended to my supervisory committee, Prof Dr. Abd. Manaf

Ali and Associate Prof Dr. Khozirah Shaari for their advice and support throughout the

past years and for imparting valuable knowledge. Without their help, this study could not

be completed successfully.



Many thanks are also due to my lab mates, Pn. Nazrien Kaman, Ms. Siti Pauliena Bohari

and Mr. Tajul Anuar which giving support and sharing their time during the previous

studies which help on numerous improvements. An acknowledgement is due to En.

Hussain Jiragon on helping me on providing materials needed and not forgotten to Ms.

Hairiah Yunus for all the guidance and support on completing this thesis.



My special thanks also goes to Head Department of Microbiology, Faculty of

Biotechnology and Biomolecular Sciences, Associate Prof. Dr Raja Noor Zaliha and to

all staff at the department.
To the Graduate School Office of Universiti Putra Malaysia, grateful acknowledged for

providing me with a Master Science studentship to perform this research.



Finally, sincere thanks are fondly expressed to my family and my beloved wife Noazira

Idris for their love, patience and support throughout this work. You never know how

much I appreciate it - a big thank you.
I certify that an Examination Committee met on 13'" Febuary 2006 to conduct the final
examination of Mohd Saufi Bastami on his Master of Science thesis entitled
"Insulinotropic Properties of Several Malaysian Herbs and Ganoderma Species" in
accordance with Universiti Putra Malaysia (Higher Degree) Act 1980 and Universiti
Putra Malaysia (Higher Degree) Regulations 1981. The committee recommends that the
candidate be awarded the relevant degree. Members of the Examination Committee are as
follows:



Johari Ramli, PhD
Associate Professor
Faculty of Biotechnology and Biomolecules Science
Universiti Putra Malaysia
(Chairman)


Mohd Noor Abd. Wahab, PhD
Associate Professor
Faculty of Biotechnology and Biomolecules Science
Universiti Putra Malaysia
(Internal Examiner)


Amin Ismail, PhD
Lecturer
Faculty of Medicine and Health Sciences
Universiti Putra Malaysia
(Internal Examiner)


Jamaludin Hj Mohamed, PhD
Professor,
Faculty of Allied Health Sciences
Universiti Kebangsaan Malaysia
(External Examiner)




                                          School of Graduate Studies
                                          Universiti Putra Malaysia

                                          Date :




                                            vii
This thesis submitted to Senate of Universiti Putra Malaysia has been accepted as
fulfillment of the requirement for the degree of Master of Science. The members of the
Supervisory Committee are as follows:



MUHAJIR HAMID, PhD
Lecturer
Faculty of Biotechnology and Biomolecular Sciences
Universiti Putra Malaysia
(Chairman)


ABDUL MANAF ALI, PhD
Professor,
Faculty of Biotechnology and Biomolecular Sciences
Universiti Putra Malaysia
(Member)


KHOZIRAH SHAARI, PhD
Associate Professor
Faculty of Science
Universiti Putra Malaysia
(Member)




                                                         AINI IDERIS, PhD
                                                         ProfessorIDean
                                                         School of Graduate Studies
                                                         Universiti Putra Malaysia




                                          ...
                                         Vlll
                                  DECLARATION

I hereby declare that the thesis is based on my original work except for quotations and
citations which have been duly acknowledged. I also declare that is has not been
previously or concurrently submitted for any other degree at UPM or other institutions.




                                                Date:
                        TABLE OF CONTENTS


                                                                    Page
DEDICATION                                                              1
                                                                        ..
ABSTRACT                                                                11
                                                                        ...
ABSTRAK                                                                 111
ACKNOWLEDGEMENTS                                                        iv
APPROVAL SHEETS                                                         vii
DECLARATION                                                             1X
TABLES OF CONTENTS                                                      X
LIST OF FIGURES                                                         xii
LIST OF TABLES                                                          xv
ABBREVIATION                                                            xvi


CHAPTER
   1      INTRODUCTION

          LITERATURE REVIEW                                             5
          2.1  Diabetes Mellitus                                        5
               2.1.1 Types of Diabetes                                  6
               2.1.2 Type 1 : Insulin -Dependent Diabetes Mellitus      6
               2.1.3 Type 2 : Non-Insulin Dependent Diabetes Mellitus   7
               2.1.4 Other Types of Diabetes
               2.1.5 Complications of Diabetes
               2.1.6 Diabetes and Its Impact
               Insulin
               2.2.1 Insulin Synthesis and Characteristic
               2.2.2 Insulin Mechanism and Action
               2.2.3 Other Usage of Insulin
               Treatment Strategies For Diabetes
               2.3.1 Insulin Injection 1 Replacement
               2.3.2 Pancreas or Islet Transplantation in Diabetes
               2.3.3 Diet Control on Diabetic Patient
               2.3.4 Commonly Used Drugs on Diabetes Treatment
               Medicinal Plants As Alternative Disease Treatments
               Insulin Secreting Cell Lines

          MATERIALS AND METHODS
          3.1 Plant Samples
          3.2 Animals
          3.3 Plants Sample Extraction
          3.4 BIUN-BD 11 Cells Culture Passage
                Cytotoxicity Assay Towards Plants And Ganoderma
                Samples                                                      28
                Insulin Release Activity From Insulin Secreting Cell Lines   30
                Rat Insulin Elisa Assay                                      31
                Streptozotocin Induce Diabetic Rats                          31
                Oral Glucose Tolerance Test By Gynura Procumbens
                Methanol Extract                                             32
                The SCGE / Comet Assay                                       33
                BRIN-BDI 1 Cells Morphological Examination By AOPI
                Assay                                                        34
                Acute Toxicity Study Of Gynura Procumbens Methanol
                Extract Towards ICR Mice                                     35
                Statistical Analysis                                         35

         RESULTS AND DISCUSSION
              Cytotoxicity Effects of Plants And Ganoderma Samples
              Towards BRIN-BD 1 1 Cells                                  36
              Insulin Secretion Responses on Malaysian Traditional
              Plants.                                                    39
              Insulin Secretion Response by Ganoderma Species.           68
              Insulinotropic Properties of Malaysian Herbs As Alternative
              In Diabetes Treatment.                                     74
              Comparison Between Glybenclamide, Tolbutamide And
              Plants Sample On Percentage Insulin Release.               80
              Oral Glucose Tolerance Test On Gynura Procumbens.          84
             In Vitro Cytotoxicity Study (MTT Assay) Of BRIN-BD1 1
              Cells Treated By Gynura Procumbens                         89
              Single Cells Electrophoresis Assay (Comet Assay) of
              Gynura Procumbens Extract On BRIN-BD 1 1 Cells             91
             Morphological Examination of BRIN-BD 1 1 Cells By
             AOPI Assay                                                  93
             Acute Toxicity Study of Gynura Procumbens Extract On
             ICR Mice                                                    97

         CONCLUSION

REFERENCES
APPENDIX
BIODATA OF THE AUTHOR
                                  LIST OF FIGURES
Figure                                                                            Page

 2.1     Human insulin amino acids sequences on chain A and chain B                 11

 3.1     Morphology of BRIN-BD1 1 cells as assessed by phase contrast              29
         microscopy (X 200 magnification.)

 4.1     The effect of Glybenclamide drugs on insulin secretion by BRIN-
         BDl 1 cell lines after 30 minutes incubation.

 4.2     Percentage insulin release by BRIN-BDll cell lines compared to
         control after 30 minutes treatment with Glybenclamide.

 4.3     The effect of Tolbutamide drugs on insulin secretion by BRIN-BDl 1
         cell lines after 30 minutes incubation.

 4.4     Percentage insulin release by BRIN-BDl1 cell lines compared to
         control after 30 minutes treatment with Tolbutamide.

 4.5     The effect of A.paniculata methanol extract on insulin secretion by
         BRIN-BD1 1 cell lines after 30 minutes incubation.

 4.6     Percentage insulin release by BRIN-BD11 cell lines compared to
         control after 30 minutes treatment with A.paniculata.

 4.7     The effect of different A.paniculata fractions on insulin secretion by
         BRIN-BD1 1 cell lines after 30 minutes incubation.

 4.8     The effect of F.deltodia methanol extract on insulin secretion by
         BRIN-BD 11 cell lines after 30 minutes incubation.

 4.9     Percentage insulin release by BRIN-BDl1 cell lines compared to
         control after 30 minutes treatment with F. deltodia methanol extract.

4.10     The effect of Gynura procumbens methanol extract on insulin
         secretion by BRIN-BD 11 cell lines after 30 minutes incubation.

4.1 1    Percentage insulin release by BRIN-BDll cell lines compared to
         control after 30 minutes treatment with G.procumbens methanol
         extract.

4.12     The effect of Labisia pumila methanol extract on insulin secretion by
         BRIN-BDl 1 cell lines after 30 minutes incubation.



                                           xii
4.13   Percentage insulin release by BRIN-BD1 1 cell lines compared to
       control after 30 minutes treatment with L.pumila methanol extract.

4.14   The effect of Momordica charantia methanol extract on insulin
       secretion by BRIN-BD1 1 cell lines after 30 minutes incubation.

4.15   Percentage insulin release by BRIN-BD11 cell lines compared to
       control after 30 minutes treatment with Momordica charantia
       methanol extract.

4.16   The effect of Morinda citrifolia methanol extract on insulin secretion
       by BRIN-BD1 1 cell lines after 30 minutes incubation.

4.17   Percentage insulin release by BRIN-BDl 1 cell lines compared to
       control after 30 minutes treatment with Morinda citrifolia methanol
       extract.

4.18   The effect of Orthosiphon spicatus methanol extract on insulin
       secretion by BRIN-BD1 1 cell lines after 30 minutes incubation.

4.19   Percentage insulin release by BRIN-BD11 cell lines compared to
       control after 30 minutes treatment with Orthosiphon spicatus
       methanol extract.

4.20   The effect of Tinospora cripsa methanol extract on insulin secretion
       by BRlN-BD1 1 cell lines after 30 minutes incubation.

4.21   Percentage insulin release by BRIN-BD11 cell lines compared to
       control after 30 minutes treatment with Tinospora crispa methanol
       extract.

4.22   The effect of Zingiber officinale methanol extract on insulin secretion
       by BRIN-BDl I cell lines after 30 minutes incubation.

4.23   Percentage insulin release by BRIN-BDll cell lines compared to
       control after 30 minutes treatment with Zingiber ofJicinale methanol
       extract.

4.24   The effect of Phylantus nirruri methanol extract on insulin secretion
       by BRIN-BD1 1 cell lines after 30 minutes incubation.

4.25   The effect of G.lucidum extract on insulin secretion by BRIN-BD11
       cell lines after 30 minutes incubation.




                                           ...
                                         Xlll
4.26   The effect of G.tropicum extract on insulin secretion by BRIN-BDl 1
       cell lines after 30 minutes incubation.

4.27   The effect of G.tsugae extract on insulin secretion by BRIN-BD11
       cell lines after 30 minutes incubation.

4.28   Percentage insulin release by BRIN-BDI 1 cell lines compared to
       control after 30 minutes treatment with various species of Ganodenna
       extract.
4.29
       The effect of G.procumbens methanol extract leaves on oral glucose
       tolerance test (OGTT) in normal rats

4.30   The effect of G.procumbens methanol extract leaves on oral glucose
       tolerance test (OGTT) in diabetic rats

4.31   The effect of Gynura procumbens methanol extract on percentage
       relative viability of BRIN-BD1 1 cells line.

4.32   SCGE of BRIN-BDI 1 after treatment with Gynura procumbens
       methanol extract.

4.33   BRIN-BDI 1 cells processed in the comet assay. Image of target cells
       DNA after micro electrophoresis below fluorescent microscope.




                                        xiv
                                   LIST OF TABLES
Table                                                                              Page

 2.1    An insulin amino acids sequence in vertebrates.                               13

 3.1    List of plants used in the experiment, source, part used and their local     25
        name.

 4.1    Inhibition concentration 50 % (ICSo) of Malaysians herbs and
        Ganoderma sp. on BRIN-BDl 1 cells by cytotoxicity assay compared
        to tolbutamide drugs.

 4.2    Comparison percentages of insulin released between glybenclamide
        and tolbutamide.

 4.3    Potentiating activity of insulin-secretory activity by methanol extract
        and drugs

 4.4    Morphological examination of BRIN-BD1 1 cells by AOPI assay

 4.5    Acute toxicity study of Gynuraprocumbens methanol extract on ICR
        mice
            LIST OF ABBREVIATIONS
            Body weight
            Carbon dioxide
cmL         Centimeter square
DMSO        Dimethyl sulfoxide
DNA         Deoxyribonucleic Acid
"C          Degree Celsius
EDTA        Ethylenediamine tetraacetic acid
ELISA       Enzyme-Linked Immunosorbent Assay
EtBr        Ethidium Bromide
FBS         Feotal bovine serum
g           Gram
IC50        Concentration of 50% inhibitory
KRBB        Krebs Ringer bicarbonate buffer
LMA         Low melting agarose
M           Molar
mA          mili Ampere
ml          Milliliter
mM          milimolar
MTT         3-[4,5-dimetiltiazol-2-yl]-2,5-dimetiltetrazolium
            bromide
N           Normality
NaCl        Sodium Chloride
NMA         Normal melting agarose
nrn         Nanometer
OGTT        Oral glucose tolerance test
PBS         Phosphate buffer saline
RPMI 1640   Roswell Park Memorial Institute 1640 Medium
W           Watt



                          xvi
v/v    Volume /volume
ul     Microliter
mg     Miligram
w/v    Weight / volume
mmol   Milimole




                    xvii
                                     CHAPTER 1



                                  INTRODUCTION

Herbal and natural products have been used for centuries throughout the world in every

culture especially on disease treatments by traditional method. Traditional medicines are

medicine is made from natural resources whether plant, fauna or mineral. It can be found

throughout the world either in the water or land. Malaysia is known for its diversity

which are claimed to possess medicinal value. The Malaysians also practice traditional

and herbal remedies as an alternative choice for disease treatment. Traditional medicines

are still a source of disease treatment of the world population. It has been shown by

Phillipson and Wright that 80 % of the world populations depends on traditional

medicine (Rahrnan et al., 1999). In two separate surveys recently conducted in Australia

and United States respectively that almost 48.5 % and 34 % of respondent have used at

least one form of unconventional therapy including herbal medicine (Grover et al., 2001).

Furthermore, many of the current commercial drugs are derived from traditional plants

based.   In the United States, approximately 25 % of prescription contains active

ingredients derived from plants. However the proportion in others countries are varies,

such as in Germany, 40 % of drugs are based on plant materials (Lee, 1999). Well

known drug for headache, aspirin, was originally created from two herbs, - white willow

bark and meadowsweet (Lee, 1999). Traditional herbs were also generating an economic

value to some countries in the world on medicinal aspect. About 30 % of the worldwide

sales of drugs are based on natural product or herbs. Developing new potential drugs
might cost millions dollars. Pharmaceutical companies have to spend IJS$ 350 million

dollars to develop new drugs (Grabley & Thiericke, 1999).



Malaysian herbal market is valued at about US$ 20 billion and growing at a rate of 20 %

annually. Much of it is imported either from Indonesia, China or India (Kadir & Lope

Pihie, 2001). Malaysian herbal research and development are focusing into few herbs

discoveries such as anti-malarial, cytotoxicity, anticancer agents from Goniothalamus

and male aphrodisiacs agent from the root extract of Eurycoma lingofolia (Kadir &

Shaari, 2000). There are many more potential herbs which were used by traditional

practitioner which are undiscovered.



Noor and Ashcroft (1989) described that over the past years; scientific and medical

knowledge on the role of plant-derived product in the treatment of diabetes mellitus has

advanced and created an exciting new area of research which could provide valuable

information for the development of alternative drugs. With rich of natural resources,

almost 6800 species of seed plants and 600 seedless plants, Malaysia may able to develop

drugs towards many diseases.



Most studies on remedies usage are not really understood. Scientific studies have been

carried out on several plants which believed to possess anti-diabetic properties.

Andrographis paniculata, the king of bitter is used since immemorial times as Chinese

and Ayuverdic medicine mainly for liver disease and dysentery was combined with

Orthosiphon spicatus on diabetes treatment (Akowuah et al., 2004).            In India,
hypoglycaemic activity of Momordica charantia or bitter gourd, known as 'peria katak'

in Malaysia, was studied since 1981 by Virdi et al., (2003).          Meanwhile, Gynura

procumbens, found in various part of Southeast Asia, has been used for the treatment of

eruptive fevers, rash and kidney disease (Zhang & Tan, 2000). Other potential herbs

reported to have anti hyperglycaemic activity are Alium cepa, Allium sativum, Murraya

koeinigii, Viscum album, Sumbucus nigra, Tinospora crispa and Morinda citrifolia

(Grover et al., 2002). Most of these plants are easily found in Malaysia.



Development for potential drugs on diabetes treatments are not limited to plants, but also

able to be produced from Ganodenna. Ganoderma lucidum which known as 'Ling Zhi '

in China or 'reishi' in Japan was studied by Tomoda et al., (1986), is reported to have

hypoglycaemic activity. It is also deemed as an elixir of life that could augment good

health and well-being.     However, hypoglycaemic activity and medicinal value of

Ganoderma tropicum and Ganoderma tsugae are unknown.



About 1200 species of Malaysian plants have been reported to have medicinal value.

Phytochemical screening, which aimed at scoring the presence of different types of

chemical compounds such as alkaloids, flavonoids, saponins, terpenoids, eugenols,

polyphenols, sterols, tannins, aloin, plysacharides, diterpenes and glycosides have been

carried out (Kadir and Shaari, 2000; Grover et al., 2002).          These phytochemical

compounds have been reported to have anti-tumor, anti-oxidant also male and women

aprodiasiac properties (Kadir and Shaari, 2000). It is also believed to have potential on
enhancing insulin secretion from pancreatic       cells since some Malaysian herbs were

used by traditional practioner on diabetes treatment.



The main objectives of the experiment are to study insulinothropic properties of some

Malaysian Herbs including some species of Ganoderma. Plants that have potential on

diabetes treatment as reported by traditional practioner will be used for insulinotrophic

acting using rat pancreatic p cell lines. Measurement was carried on insulin secreting cell

lines (BRIN-BDII cell lines) using insulin ELISA methods.                 Various insulin

concentrations are released by BRIN- BDl 1 cell lines affect from its stimulator. Plants

and Ganoderma samples was extracted and tested towards BRIN-BDI 1 cells on their

ability in enhancing insulin secretion and determine the cytotoxicity level and      value

by [3-(4, 5-dimethylthiazol-2-y1)-2,5-dimethyltetrazolium bromide (MTT) assay.



The second objective is to study the anti-hyperglycemic properties of Gynura

procumbens.       A further study of Gynura procumbens methanolic extract on

hypoglycaemic activities and its toxicity was carried as Gynura procumbens was reported

by traditional practitioner to be safe taken as edible and useful in controlling diabetes.

Study were carried to determine the maximum insulin levels secreted by BRIN-BDI 1 cell

lines and in vivo study of OGTT (oral glucose tolerance test) on normal and induced

diabetic rats by Gynura procumbens methanolic extract. Toxicity level of Gynura

procumbens methanolic extract towards BRIN-BDl I cell lines and ICR strain mice will

be evaluated. The genotoxicity towards BRIN-BDl 1 also were studied by Comet Assay

and AOPI assay.
                                      CHAPTER 2



                               LITERATURE REVIEW



2.1 Diabetes Mellitus



Diabetes mellitus (DM) is a complex disorder of carbohydrate, fat and protein

metabolism. It is primarily a result of a defect in secretion or action of insulin, the

hormone that facilitates and control the use of glucose in the cells. Due to the deficiency

of insulin, diabetic patients have an impaired tolerance to glucose that leads to numbers

of short term and long term complications. Early diagnosis of diabetes is determined by

blood glucose level. Glucose levels before meal are generally run between 4-7 mmolll or

60-100 mgldl in normal people (Williams & Porte, 1974; Dagget, 1981). Diabetes not

only limited to human being but also occurred on canine especially on type 1 diabetes,

insulin dependent diabetis mellitus (IDDM) (Davison et al., 2002).



Diabetes Mellitus is the commonest endocrine disorder that affects more than 100 million

people worldwide (6 % of the population) and projected to grow over 220 millions within

40-50 years (Proietto et al., 1999). In next 10 years, diabetes may affect about five times

more people than it does now (Bailey, 2000; Grover et al., 2002).        Diabetes derived

fiom Greek for siphon, which refers to the copious excretion of water that characterizes

the disease, meanwhile mellitus came fiom Latin word for honey, characterized by the

high sugar content in urine.
In Malaysia, diabetes mellitus is increasing towards serious level. The Ministry of Health

has recorded 657,988 people in Malaysia suffering from diabetes in 2002 increasing from

525,858 in year 2001 (Abdullah, 2003). The number of diabetics estimated to be

increased to 0.8 million cases in 2025. From the reports of admissions to government

hospitals in Peninsular Malaysia, diabetes mellitus had increased from 5024 cases in

1979 to 17808 cases in year 1990, and in year 2020 about 144 600 Malaysians will

suffering diabetes (Amos et al., 1997).




2.1.1 Types of Diabetes



Diabetes can be divided into three distinct types, in which subtypes have been identified.




2.1.2 Type 1: Insulin -Dependent Diabetes Mellitus (IDDM)



Type 1 or insulin-dependant diabetes mellitus (IDDM) occurs in approximately 10 % of

all diabetics' patients in western world.    The symptoms are hyperglycaemia due to

secondary insulin deficiency, occurring as a result of autoimmune destruction of

pancreatic endocrine   p cells (Perfetti & Ahmad, 2000). Type 1 commonly occurs on
childhood (Docherty, 2001). This type of diabetes accounts for 3 % of all diabetes

worldwide in 1997, meanwhile type 2 is by far the most common type (Proietto et al.,

1999).
2.1.3 Type 2: Non-Insulin Dependent Diabetes Mellitus (NIDDM)



Non-insulin dependent diabetes mellitus occurs when insulin is no longer reacting as

metabolic hormone in reducing blood sugar level (insulin resistant) or insufficient ability

to secrete insulin (Proietto et al., 1999; Perfetti & Ahrnad, 2000). It is reported that type

2 diabetes has strong genetic tendency (Proietto et al., 1999). Combination of insulin

resistant and insufficient insulin released has accounting 90-95% of diabetes in developed

countries (Husen et al., 2004). About 90 % of diabetics in western world are categorized

by type 2 rather than type 1. Type 2 diabetic shows direct contact with obesity. Hartz et

al., 1983 and Kissebah et al., 1984, reports that a close relationship between upper body

obesity with type 2 diabetes by increased free fatty acid oxidation could impair glucose

oxidation thus leading to insulin resistance (Proietto et al., 1999). The risk of diabetes

seems to be increased ten times in women with both severe obesity and high waist to hips

ratio.




2.1.4 Other Types of Diabetes



Other types of diabetes including entities secondary to or associated with certain other

conditions or syndromes. Diabetes may be secondary to pancreatic disease removal of

pancreatic tissues, endocrine disease such as acromegaly, Cushing's syndrome,

glucagonoma, somatostatinoma, or the administration of certain drugs, hormones or

chemicals that causing hyperglycaemia (Hamid, 1999).

				
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