ﻢﻴﺣﺮﻟا ﻦﻤﺣﺮﻟا ﷲا ﻢﺴﺑ THE EFFECT OF FRESH CRUSHED GARLIC ... · THE...

بسم اهللا الرحمن الرحيمTHE EFFECT OF FRESH CRUSHED GARLIC BULBS (ALLIUM SATIVUM) ON PLASMA CHOLESTEROL

AND TRIGLYCERIDES IN CHOLESTEROL FED RATS By

Maha Mohamed Khalil Mohamed Salih B.Sc. (2002)

Khartoum University Faculty of Science

Supervisor Dr. Barakat El Hussein Mohamed

A thesis submitted to the University of Khartoum in partial fulfillment for the requirement of the Degree of Master of Science in Biochemistry

Department of Biochemistry Faculty of Veterinary Medicine

University of Khartoum

July 2006

DEDICATION

This study is dedicated

To my mother,

Family, friends

And darlings

With respect

Maha

ACKNOWLEDGEMENTS

Praise is to Allah, the lord of the creation, although I find it

possible to mention only a few, I sincerely appreciate each one who

contributed towards the completion of this thesis.

Special thanks and deep appreciation to my university supervisor

Dr. Barakat El Hussein Mohamed, the Head Department of

Biochemistry, Faculty of Veterinary Medicine, University of Khartoum,

for his guidance and support.

My deepest gratitude and sincere appreciation to Dr. Abd Elwahab

Hassan Mohamed, the Head Department of Pharmacology, Medicinal and

Aromatic Plants Research Institute (MAPAI), National Center for

Research for suggestions and beneficial criticism during the period of this

work.

Thanks should be extended to the staff member of Research and

Laboratory Units, Khartoum Teaching Hospital for laboratory facilities.

I would like to thank Mr. Hamza Ibrahim for his patience and

typing of the thesis, thanks is due to Miss Nisreen Fathi for her

considerable assistance in the statistical analysis of data.

The endless thanks are due to my kind uncle Asmaa, my cousin

Amal Omer and my dear friends Shihab El Deen and Mai Sinada for their

support and encouragement.

My deep respect and gratitude are due to the members of my

family for their patience and continuous encouragement.

CONTENTS

Page DEDICATION ……………………………………………..... i ACKNOWELDGEMENTS .………………………………… ii CONTENTS ……………………………………….…………… iii LIST OF TABLES …………………………………………….. v LIST OF FIGURES ……………………………………………. vi ENGLISH ABSTRACT ……………………………………… vii ARABIC ABSTRACT …………………………………………. viii INTRODUCTION ……………………………………………… 1 CHAPTER ONE: LITERATURE REVIEW ….…………… 4 1.1 Lipids ………………………………………………………. 4 1.1.1 Cholesterol ………………………………………... 5 1.1.2 Types of cholesterol ……………………………… 6 1.1.2.1 High density lipoprotein ………………….. 6 1.1.2.2 Low density lipoprotein …………………… 7 1.1.3 Triglycerides ……………………………………… 7 1.2 Development of atherosclerosis and coronary heart disease. 8 1.2.1 Atherosclerosis ……………………………….…… 8 1.2.2 Coronary heart disease …………………………… 8 1.3 Hypolipidemic plants ………….…........................................ 9 1.3.1 Mukul myrrh tree (commiphora mukul)………..…. 9 1.3.2 Fenugreek (Trigonella foenum–graecum) ………... 10 1.3.3 Soy protein ……………………………………..… 10 1.3.4 Hawthron (Crateagus spp) …………………….… 10 1.3.5 Oats ……………………………………………… 11 1.3.6 Vaccinium myrtillus ……………………………… 11 1.3.7 Nigella sativa ……………………………………… 12 1.3.8 Sugar cane ………………………………………… 12 1.3.9 Green tea (Camellia sinensis) …………………… 13 1.3.10 Turmeric root….………………………….………. 13 1.4 Taxonomy of Allium sativum …………………………….. 13 1.4.1 Botanical description ……………………………... 14 1.4.2 Distribution ………………………………............. 14 1.5 Chemistry of garlic bulbs …………………………………. 16

6 Antioxidant action of Allicin ………………………………. 17 1.7 Medical usage of garlic …………………………….……… 19 CHAPTER TWO: MATERIALS AND METHODS……….. 23 2.1 Experimental details ……………………………………….. 23 2.1.1 Experimental animals……………………………… 23 2.1.2 The rat basal diet …................................................ 23 2.2 Cholesterol supplementation in the rat diet...………………. 24 2.3 Preparation of the plant material …………………………... 24 2.4 Equipments used ………………………………………….. 24 2.5 Chemicals …………………………………………………. 25 2.6 Experimental procedure …………………………………… 25 2.7 Blood sampling …………………………………………… 25 2.8 Analytical methods ……………………………………….. 26 2.8.1 The roch diagnostic/Hitachi 902 analyzer ……….. 26 2.8.2 Determination of plasma total cholesterol ………. 26 2.8.3 Determination of plasma low density Lipoprotein 28 2.8.4 Determination of plasma high density lipoprotein 29 2.8.5 Determination of plasma triglycerides ……………. 30 2.8.6 Procedure ………………………………………… 31 2.9 Statistical analysis ………………………………………… 31

CHAPTER THREE: RESULTS……………………………... 33 3.1 The effect of fresh crushed garlic bulbs on plasma total

cholesterol ………………………………………………... 33 3.2 The effect of fresh crushed garlic bulbs on plasma low

density lipoprotein ……………………………………….. 36 3.3 The effect of fresh crushed garlic bulbs on plasma high

density lipoprotein ………………………………………... 38 3.4 The effect of fresh crushed garlic bulbs on plasma

triglycerides ……………………………………………… 40

CHAPTER FOUR: DISCUSSION ………………………… 42 4.1 Plasma total cholesterol …………………………………... 42 4.2 Plasma low density lipoprotein ………………………… 43 4.3 Plasma high density lipoprotein …………………...…… 44 4.4 Plasma Triglycerides …………………………………… 45 CONCLUSION ……………………………………………… 46 REFERENCES ……………………………………………… 47

LIST OF TABLES

Page

Table (1): Garlic products in the market …………………..... 21

Table.(2): The effect of fresh crushed garlic bulbs on plasma

total cholesterol, LDL-C, HDL-C and TG in an

induced hypercholesterolemic Wistar albino rats… 34

LIST OF FIGURES

Page

Figure (1): Garlic plant. …………………………………… 15

Figure (2): Garlic bulbs ………………………..………….. 15

Figure (3): Chemical change in garlic……………………... 18

Figure (4): The roch diagnostic / Hitachi 902 Analyzer…. 27

Figure (5): Hitachi sample cups …………………………... 32

Figure (6): The effect of fresh crushed garlic bulbs on plasma total cholesterol level (mg/dl)…..... 35

Figure (7): The effect of fresh crushed garlic bulbs on plasma LDL-cholesterol level (mg/dl)………. 37

Figure (8): The effect of fresh crushed garlic bulbs on plasma HDL-cholesterol level (mg/dl)……… 39

Figure (9): The effect of fresh crushed garlic bulbs on plasma triglycerides level (mg/dl)…………… 41

ABSTRACT

The aim of this study was to investigate the effect of fresh crushed

garlic bulbs on plasma total cholesterol (TC), low density lipoprotein

(LDL), high density lipoprotein (HDL) and triglycerides (TG) levels in an

induced hypercholesterolemic Wistar albino rats.

Twenty male Wistar albino rats were used in this study. They were

allotted to four groups A, B, C and D. Group A ( control group) received

a basal diet, group B received a basal diet with 2% cholesterol, group C

received a basal diet with 2% cholesterol and 4% garlic, group D received

a basal diet with 2% cholesterol and 8% garlic.

The results showed that, plasma TC, LDL-cholesterol and TG

levels were increased significantly (P< 0.05) in group B compared to the

control group, while HDL-cholesterol level was decreased significantly.

In group C, the plasma levels of TC and LDL-cholesterol were

decreased significantly (P< 0.05), however, HDL-cholesterol was

increased compared to group B, but no significant difference in the level

of TG was observed compared to group B. However group C showed

significantly (P< 0.05) higher levels of TC and TG and non- significant

difference in LDL-cholesterol and HDL-cholesterol levels compared to

the control group.

The plasma levels of TC, LDL-cholesterol and TG in group D were

decreased significantly (P< 0.05) compared to group B, while HDL-

cholesterol was increased. The levels of TC, LDL-cholesterol, HDL-

cholesterol and TG in group D were nonsignificantly different compared

to the control group.

ملخص األطروحة

هدفت هذه األطروحة إلى معرفة أثـر الثـوم الطـازج المهـروس علـى مـستوى

الكوليسترول الكلي والاليبوبروتين منخفض الكثافة والاليبوبروتين عالي الكثافة والجلسريدات

.ضافياً بالكوليسترولالثالثية في دم الجرذان المغذية إ

جرذ ذكر وقُسمت العينة إلى أربعـة مجموعـات 20احتوت عينة الدراسة على عدد

هي عينة التحكم والتي ُأعطيت وجبة ) أ(حيث كانت المجموعة . متساوية هي أ ، ب ، ج ، د

، المجموعـة %2ُأعطيت الوجبة األساسية بإضافة كوليسترول بنسبة ) ب(أساسية، المجموعة

أما المجموعـة % 4وثوم بنسبة % 2ُأعطيت الوجبة األساسية بإضافة كوليسترول بنسبة ) ج(

%.8بنسبة وثوم % 2ُأعطيت الوجبة األساسية بإضافة كوليسترول بنسبة ) د(

أوضحت النتائج زيادة معنوية في مستوى الكوليسترول الكلي والاليبوبروتين منخفض

حـين أن بالمقارنة مع مجموعة التحكم فـي ) ب(المجموعة الكثافة والجلسريدات الثالثية في

.مستوى الاليبوبروتين عالي الكثافة انخفض معنوياً

نجد أن مستويات الكوليسترول الكلي والاليبـوبروتين مـنخفض ) ج(وفي المجموعة

الكثافة قد انخفضا معنوياً، في حين أن الاليبوبروتين عالي الكثافة قـد ازداد بالمقارنـة مـع

على الـرغم . ولكن لم يالحظ اختالفاً معنوياً في مستوى الجلسريدات الثالثية ) ب(المجموعة

اظهرت ارتفاعاً معنوياً في الكوليسترول الكلي والجلسريدات الثالثيـة ) ج (من أن المجموعة

وال يوجد اختالفاً معنوياً في مستويات الاليبوبروتين منخفض الكثافة والاليبـوبروتين عـالي

.الكثافة بالمقارنة مع مجموعة التحكم

ـ ) د(في المجموعة نخفض نجد أن مستويات الكوليسترول الكلـي والاليبـوبروتين م

بينمـا مـستوى ) ب(الكثافة والجلسريدات الثالثية قد انخفضت معنوياً مقارنة مع المجموعة

وأن مـستويات الكوليـسترول الكلـي والاليبـوبروتين . الاليبوبروتين عالي الكثافة قد ازداد

رلم تُظه ) د(منخفض الكثافة والاليبوبروتين عالي الكثافة والجلسريدات الثالثية في المجموعة

.اختالفاً معنوياً مقارنةً مع مجموعة التحكم

INTRODUCTION

Elevated plasma level of cholesterol is linked to the development

of atherosclerosis. This disease results when LDL-cholesterol deposited

in the wall of blood vessels (Gonona, 1997).

Many studies indicated that lowering plasma cholesterol may

prevent, control and even reverse atherosclerosis and coronary heart

disease (Bierman, 1994).

The drugs used for lowering blood cholesterol generally have

undesirable side effects. Hence, a harmless lipid-controlling mediation

would be a welcome development. Medicinal and aromatic plants are the

important renewable source of drugs. In addition, WHO encourages the

inclusion of medicinal plants in programmes of developing countries

because of the great potential these plants represent in combating various

diseases. Recently, the Sudan has produced certain drugs locally using

important or native raw materials (Gabir, 2001).

Garlic has been employed in folk medicine since ancient time for

prophylaxis as well as cure of a variety of diseases (Alie and Thomson,

1995; Merat and Fallahzadeh, 1996).

Extensive clinical and scientific studies support the use of garlic for

treatment of hypercholesterolemia, however, the potency of garlic

product can vary substancially due to mode of preparation (David et al.,

2001). These studies include garlic tablets (Lash et al., 1998), garlic

powder (Issacsohn et al., 1998; Tohidi and Rahbani, 2000), enteric coated

garlic preparations i.e., Garlinase, Garlicin and Garique, essential oil and

oil macerate ( Harunobu et al., 2001), extract of frozen and raw garlic

(Slowing et al., 2001), water extractable fraction (WEF), petroleum

extractable fraction (PEF), methanol extractable fraction (MEF), aged

garlic extract (AGE) (Yu-Yan and Lijuan, 2001), swallowing and

chewing garlic (Abbas et al., 2005). The hypercholesterolemic effect of

the different preparations used was attributed to water soluble sulfur

compounds, especially, S-allyl cysteine (SAC) and ajoene. However,

fresh crushed garlic bulbs has been reported to contain high amount of

allicin (3.7 mg/dl) (Lawson and Hughes, 1992 and Harunobu et al.,

2001).

Allicin has been proposed as the active compound produced by

garlic, which is responsible for its hypocholesterolemic effect. Since no

garlic product on the market contains a detectable amount of allicin

(<1ppm) (Harunobu et al., 2001), and also some of the bioactive

compounds of garlic are affected by heat (Gazzani et al., 1998 and Kim,

2002). Therefore, the objective of this study is to investigate the effect of

fresh crushed garlic bulbs, the form which most people consume it, on the

level of plasma total cholesterol and its fractions (low density lipoproteins

and High density lipoproteins) and triglycerides in an induced hyper-

cholesterolemic Wistar albino rats.

CHAPTER ONE

LITERATURE REVIEW

1.1 Lipids:

Lipids have the common property of being relatively insoluble

in water but soluble in nopolar solvent. Lipids are important dietary

constituents not only because of their high energy value but also because

of the fat-soluble vitamins and the essential fatty acids contained in the

fat of natural food (Robert et al., 1996). All lipids are transported as a

part of lipoprotein complexes, four major lipid classes are presented in

lipoproteins, including triacylglycerols, phosphor-lipids, cholesterol and

cholesterol esters (Brian et al., 2001). The proportions of these vary

considerably between the different lipoprotein classes. Plasma lipids are

classified into two groups, non-esterified fatty acids (NEFA), which are

transported, bound to albumin and the other group is lipoproteins, these

are, in order of increasing density, very low density lipoproteins (VLDL),

low density lipoproteins (LDL) and high density lipoproteins (HDL).

These forms of lipoproteins are all derived from endogenous lipid

components, whereas, the fourth type of lipoprotein, termed

chylomicrons, drive their lipids from dietary sources. IDL is another class

lies on the pathway which VLDL is converted to LDL (Robert et al.,

1996). In general the structures of the different plasma lipoproteins are

very similar, being more or less spherical. The hydrophobic components,

triacylglycerols and cholesterol esters are located in the core of

complexes surrounded by a monolayer of phospholipids and cholesterol

associated with integral and peripheral apoprotein. The biosynthesis and

utilization of the components of these complexes are now being

elucidated and it is appreciated what a central role they play in both

health and disease (Brian et al., 2001).

1.1.1 Cholesterol:

It is a steroid compound , however, it is the precursor of a large

number of important steroids including the bile acids, adrenocortical

hormones, sex hormones and vitamin D. It is also an important

amphipathic lipid, which allows it to play a structural role in membranes

and in the outer layer of lipoproteins. Cholesterol ester is the storage form

of cholesterol found in most tissues. Cholesterol is synthesized in the

body entirely from acetyl-coA via a complex pathway. The greater part of

cholesterol of the body about 1500 – 2000 mg are manufactured each day

by the liver from the fats, protein and carbohydrates we eat, but only

about 200 – 800 mg is provided from our routine diet (Daniel, 1995). On

the other hand excess cholesterol is excreted from the liver in the bile as

cholesterol or bile salts. A large proportion of bile salts returned to the

liver as part of the enterohepatic circulation (Robert et al., 1996).

Cholesterol needs a special carrier in the blood stream, so it is transported

as lipoprotein in the plasma. Various studies indicated that high serum

levels of cholesterol are strongly related to coronary atherosclerosis and

increased risk of coronary heart disease. Clinical studies have shown that

lowering levels of serum total cholesterol by diet or drugs decreased the

incidence of coronary heart disease (Bierman , 1994).

1.1.2 Types of cholesterol:

1.1.2.1 High density lipoprotein (HDL):

High density lipoproteins are one of the major classes of the

plasma lipoproteins. It is a central player in the cholesterol economy of

the body (good form of cholesterol), derived largely from the liver and on

entering the circulation is found to be discoid in shape. The disc consists

of phospholipids, cholesterol bilayer associated with a number of

apolipoproteins and the enzyme lecithin cholesterol acyl- transferase

(LCAT) (Brian et al., 2001). In reverse cholesterol transport, HDL

attaches to the apo A-1 receptors, causing translocation of cholesterol to

the cell membrane, where it is taken up by HDL. A concentration

gradient is maintained by the activity of LCAT, which allow cholesterol

to be esterified and deposited in the core of HDL, then it is taken up by

the liver either directly, or after transport to VLDL, IDL or LDL via the

cholesteryl ester transfer protein (Robert et al., 1996). An inverse

relationship between HDL-cholesterol levels in serum and incidence of

coronary heart disease (CHD) has been demonstrated in a number of

epidemiological studies (Hassan, 2001).

1.1.2.2 Low density lipoprotein (LDL):

Low density lipoprotein is called bad form of cholesterol, it is the

major cholesterol-carrying lipoprotein in plasma. Most LDL-cholesterol

appears to be formed from VLDL via IDL. These remnants are composed

mainly of cholesterol ester and apo-B 100 and it is taken up by the liver

(70%) and other tissues (30%) via the LDL receptors. The number of

LDL receptors on the cell surfaces is regulated by the cholesterol

requirement for membrane, steroids hormones, or bile acid synthesis.

Thus, influx of cholesterol down-regulates the number of LDL receptors

(Robert et al., 1996 and Brian et al., 2001). A positive correlation exists

between the incidence of atherosclerosis and concentration of LDL-

cholesterol (Lalitha, 1996).

1.1.3 Triglycerides:

Triglycerides are the main storage form of fatty acids. They represent

the most important class of dietary fat which constitute more than 90% of

the dietary lipids. These compounds also supply the body with the

essential fatty acids required, in addition to the energy (Murray et al.,

1999). High blood levels of triglycerides were found in many cases, such

as metabolism disorder, diabetes and obesity.

A small portion of triglycerides are found in the blood stream which

alone do not cause atherosclerosis. But lipoproteins that are rich in

triglycerides cause atherosclerosis in many people with high triglycerides

level, so increasing the chance of having heart attack (Mirkin, 2000).

1.2 Development of atherosclerosis and coronary heart disease:

1.2.1 Atherosclerosis:

Atherosclerosis refers to the deposition of cholesterol and

cholesterol ester of lipoproteins containing apo B-100 in the arterial wall

which leads to the formation of plaque and results in endothelial damage

and narrowing of the lumen.

It remains dormant in the body for two or three decades without

showing and outward symptoms of its presence in the form of high blood

pressure, heart disease and cerebral haemorrhage, eventually leading to

coronary heart disease (Basavaraju and Jones, 1998).

1.2.2 Coronary heart disease (CHD):

CHD refers to spectrum of clinical heart disorders resulting from

inadequate blood flow to the heart muscle, compared to its needs. CHD is

caused by atherosclerotic narrowing of coronary arteries. It has been

demonstrated consistently, in epidemiological studies within and between

populations, that there is a continuous increase in risk of death from CHD

with increasing serum cholesterol. A major risk factor for CHD seems to

be a high concentration of LDL-cholesterol, which carries about 70% of

the total circulating cholesterol. In contrast, high levels of HDL-

cholesterol are thought to have a protective effect, and lower the risk of

CHD (Griffin et al., 1998).

1.3 Hypolipidemic plants:

Numerous medicinal plants have shown hypolipidemic activity by

various experimental assay methods. Several of these are available in

commercial preparations, either as a single extract or in combina-tion

products (Winston, 1999).

1.3.1 Mukul myrrh tree (Commiphora mukul):

It has been shown that a 14-27% of the total cholestero and 22-30%

of triglycerides levels were reduced while HDL-cholesterol was increased

by 20% when 25g three times per day for 12 weeks of gugglul gum of the

Mukul myrrh tree were given to patients with elevated serum cholesterol

level (Satyavati, 1988). Analysis of guggul gum showed that the guggul-

sterons are the active ingredients. The mechanism of cholesterol lowering

action appears to be due to its ability to increase the liver's metabolism of

LDL-cholesterol. It was shown that by increasing the break down of

LDL-cholesterol in the liver, it can lower LDL-cholesterol levels by 25-

35% (Verma and Bordia, 1998; Werbach and Murray, 1994).

1.3.2 Fenugreek (Trigonella foenum - graecum):

Subjects with elevated serum cholesterol who consumed 30 g of

powdered fenugreek seeds three times per day showed a significant

reduction in total cholesterol, triglycerides and LDL-cholesterol. The

saponine contained in plant fibers increase the loss of bile acids by fecal

excretion, which then leads to an increased conversion of cholesterol into

bile acids by the liver (Elson et al., 1998 and Hassan, 2001).

1.3.3 Soy protein:

Several studies have shown that when soy protein consumption

increases, serum lipid profile was improved. Daily intake of 27g soy

protein lowered serum total cholesterol and LDL-cholesterol by 9% and

13% respectively and triglycerides by 11% (Anderson, 1995). The precise

mechanism of this effect is unknown, but a possible explanation

including a higher bile acids excretion and saponine in soy can block

cholesterol absorption from the diet (Carrol, 1991 and Potter, 1995).

1.3.4 Hawthron (Crateagus spp):

The leaves and fruits of the hawthorn plant have been used widely

for cholesterol reduction. Studies using rats suggested that the tincture

from crateagus (made from the barriers) may be a powerful agent for the

removal of LDL-cholesterol from the blood stream, the tincture of

hawthorn barriers also reduced the production of cholesterol in the liver

of rats fed a high cholesterol diet (Schmidt, 1994).

1.3.5 Oats:

The effective viscous fibers from oats are β-glucan. (Braaten ,

1994). 3 g of β-glucan per day lowered serum total cholesterol

concentration by 16 mg/dl in hypercholesterolemic subjects (Ripsin,

1992). There were several mechanisms to explain the cholesterol

lowering effect of β-glucan. One hypothesis proposes that β-glucan

increases bile acids synthesis by binding to bile acids in the intestine.

Alternatively, it is possible that β-glucan act as a physical barrier in the

intestine, blocking absorption of bile acids and cholesterol (Bell, 1999).

Other hypothesis has postulated that soluble fibers are bacterially

fermented in the colon, leading to production of short-chain fatty acids,

which may lower hepatic cholesterol synthesis. Soluble fibers lower

insulin level, which is known to lower cholesterol synthesis (Bourbon,

1999).

1.3.6 Vaccinium myrtillus:

Oral administration of hydro-alcoholic extract of Vaccinium

myrtillus leaves significantly decreased plasma triglycerides and

cholesterol levels. Animal studies suggested that the active ingredient

bilberry may prevent the oxidation of LDL-cholesterol (Cignarella et al.,

1993).

1.3.7 Nigella sativa:

Nigella sativa oil was reported to reduce serum total cholesterol,

triglycerides and LDL-cholesterol and significantly increased serum

HDL-cholesterol levels when was given orally to rats at a dose of 800

mg/kg body weight (Bwt) for four weeks (El-Dakhakhny et al., 2000).

The thymoquinone is the active ingredient of Nigella sativa which acts, to

induce the liver cells to increase conversion of cholesterol into bile acids

(Amir et al., 2006).

1.3.8 Sugar cane:

Policocanol is the main ingredient, it is a mixture of alcohol isolated

and refined from sugar cane. Administration of 10 g of Policocanol daily

for eight weeks to patients with hypercholesterolemia had shown a 18%

reduction in total cholesterol, a 25% reduction in LDL-cholesterol and

21% increase in HDL-cholesterol. Policocanol appears to normalize

cholesterol better than cholesterol lowering drugs, without side effects.

Although its exact mechanism is not known, Policocanol appears to act

by blocking the body synthesis of cholesterol and inhibiting the oxidation

of LDL-cholesterol (Arrzazabala, 2000 and Castano et al., 2001).

1.3.9 Green tea (Camellia sinensis):

The green tea has the ability to lower total cholesterol and raise

HDL-cholesterol levels in both animals and humans. Although an animal

study which was conducted to determine how green tea affects these

changes was not conclusive, suggesting that catechins in green tea may

block intestinal absorption of cholesterol and promote its excretion from

the body (Chan et al., 1999 and Kuan et al., 2004).

1.3.10 Turmeric root:

Turmeric contains curcumin and other essential oil. Curcumin

increases the secretion of bile by stimulating the bile duct, and then helps

to treat the high cholesterol level (Soudamini et al., 1999).

1.4 Taxonomy of Allium sativum:

Kingdom : Plantae.

Division : Magnoliophyta.

Class : Liliopsida.

Order : Asparagales.

Family : Alliaceae.

Sup family : Allioidear.

Tribe : Allieae.

Genus : Allium

Species : A sativum.

1.4.1 Botanical description:

Garlic is a perennial plant, belong to the genus allium, which is

comprised of approximately 600 known species. The garlic plant has

long, narrow and flat leaves (Fig. 1). The bulb of garlic consists of

numerous bulbest “cloves” of different size (1– 6 cm tall), with an acute

summit and the base truncate. Each clove is separately enclosed in a

white scale and covered with a pinkish-white skin, and the whole

surrounded by layers of white scale leaves (Fig. 2) (Koch and Lawson,

1996 and Mcgee, 2004).

Flowers are variable in numbers, found on slender pedicles and

sometime absent, they seldom open and may wither in bud. Fruit is a

small capsule, seeds are seldom (Ion, 1999). The domesticated garlic

plant dose not produces seed, but is grown from bulbs.

1.4.2 Distribution:

Garlic is the second most cultivated of alliums spp, after onion

worldwide (Purseglove, 1988).

The crop is grown during late autumn in many countries while it is

grown as a winter crop in Sudan in high altitude parts of Western States

e.g. Jabal Marra or in the sails along the Nile banks in the Northern

Region.

Fig (1): Garlic plant

Fig (2): Garlic bulbs

Production depends upon many factors. Lipinski et al. (1994)

reported increased yields in response to frequent irrigation and nitrogen

application.

1.5 Chemistry of garlic bulbs:

The chemistry of garlic is quite complex. Intact garlic bulbs contain

high amount of γ-glutamylcysteines. These reserve compounds can be

hydrolyzed and oxidized to form alliin, which accumulates naturally

during storage of garlic bulbs. After processing, such as cutting, crushing

or chewing, the vacuolar enzyme, alliinase, rapidly lyses cytosolic

cysteine sulfoxides (alliin) to form the odoriferous alkyl-alkane-

thiosulfinates (allicin). The enzymetic metabolite, allicin, is a highly

reactive substance which is transformed more rapidly or more slowly,

depending on preparation and surrounding media into lipid soluble

compounds, such as diallyl sulfide (DAS), diallyl disulfide (DADS) and

diallyl trisulfide (DATS) or to water soluble compounds such as S-

methylcystein (SMC), S-allyl acetylcysteine (SAAC) and ajoene (Yu-yan

and Lijuan, 2001). At the same time γ-glutamylcysteines are converted to

S-allyl cysteine (SAC) via a pathway other than the alliin/allicin pathway

(Fig. 3) ( Harunobu et al., 2001). Additional constituents of intact garlic

include glycosides, prostaglandins, pectin, essential oil, vitamins,

enzymes, flavonoids, essential amino acids, trace elements, minerals and

others (Liu, 2000; Barenes et al., 2002 and Skidmore, 2003).

1.6 Antioxidant action of allicin:

Oxidation of lipid, notably oxidative modification of LDL-

cholesterol by reactive oxygen species (ROS), is implicated in the

development of cardiovascular disease. Oxidation of polyunsaturated

fatty acids in LDL-cholesterol leads to the release of degradation products

such as peroxides, which react with lysine residues on apoprotein B,

modifying its structure. As a result, it will not be recognized by normal

LDL receptors. It is, however, recognized by scavenger receptors on

macrophages and smooth muscle cells which are capable of taking up

large amount of lipid leading to the formation of foam cells and

consequently formation of plaque and increased risk of heart disease and

stroke (Ide and Lau, 1997 and Brian et al., 2001).

Substances which have been found to protect us from the cellular

damage caused by free radicals include vitamin C, bioflavonoid, vitamin

E and selenium. Garlic is not usually found on standard lists and probably

should be (Brian et al., 2001). Fresh crushed garlic has an abundance of

thiosulfinates (allicin) which is an excellent antioxidant, that may remove

peroxides, thereby, preventing further generation of ROS by enhancing

the cellular antioxidant enzymes such as superoxide dimutase, catalase

and glutathione peroxidase (Java, 2005). Hence allicin reduce the amount

NH2 H O NH2

N S HooC C SH CooH

O CooH

γ-glutamylcysteines alliin

alliinase NH2 O S S CooH S

S-allyl cysteine allicin (SAC)

DAS, DADS, DATS, SMC, SAAC , a joene, etc.

Fig. (3): Chemical change in garlic

of circulatory oxidized LDL-cholesterol and subsequently accumulation

of cholesterol in macrophage, smooth muscle and blood vessels walls,

resulting in an inhibition of atherogenic fatty streak then lower the risk of

cardiovascular disease (Boston, 2001).

1.7 Medical usage of garlic:

Garlic contains characteristic organosulfur compounds that

contribute to its pharmacologic activities. These organosulfur compounds

are converted to other compounds during processing. Processing triggers

the formation of a cascade of compounds that do not exist originally in

garlic cloves, therefore different processing techniques produce different

constituents in the preparations. Thus, different preparations may have

different health benefits (Munday et al., 1999 and Mckenna et al., 2002).

Although there are many garlic supplements commercially available, they

fall into one of five categories, each one have its own characteristics as

shown in table (1) (Harunobu et al., 2001). However, recent publications

suggested that all preparations may not have hypercholesterolemic

activity because, highly unstable thiosulfinate (allicin) disappears during

processing. For this reason no garlic product on the market contains a

detectable amount of allicin (<1 ppm) (Harunobu et al., 2001). A

European trial comparing garlic with commercial lipid lowering drugs

(benzafibrate and a fabric acid derivative) found them to be equally in

decreasing lipid to a statistically significant extent (Holzgartner et al.,

1992). Although the mechanism of cholesterol lowering effect of garlic

is not completely understood, the data from animal study indicated

that the hypocholesterolemic action of garlic may be due to the inhibition

of hepatic cholesterol synthesis by allicin through the formation of sulfide

bridges by the disulfides found in garlic with 3-hydroxy-3-methyl

glutaryl coenzyme A (HMG-COA) reductase (Zhang et al., 2001).

Sulfur containing amino acids such as allyl propyle disulfur isolated

from garlic has hypoglycemia properties (Sheela, 1992). Experiments in

non-diabetic individuals showed significant decrease in fasting serum

glucose (Kiesewetter et al., 1991) by mechanisms including increased

secretion or slowed degradation of insulin, increased glutathione

peroxidase activity and improved liver glycogen storage (Shane, 2001).

Aqueous extracts of garlic have long been established to exhibit

wide spectrum of antimicrobial activity, they are active against several

general of bacteria (Staphylococcus and Salmonella tryhimarium), fungi

(Candida albicans) and worms (Ascaris lumbricodes). The key

substances responsible for the antimicrobial activity of garlic are the

thiosulfinates, particularly allicin (Johnson and Vaughn, 1996).

Table (1): Garlic products in the market

Type of product

Main compounds and characteristics

Garlic essential oil

Garlic oil macerate Garlic powder Aged garlic extract Enteric-coated garlic tablets

Only lipid-soluble compounds. No water-soluble fraction. No allicin. Not well standardized. No safety data. Lipid-soluble sulfur compounds and allin. No allicin. Not well standardized. No safety data. Allin, small amount of lipid-soluble compounds. No allicin. Not well standardized. No safety data (severe effects). Results on cholesterol are not consistent. Mainly water-soluble compounds (SAC, saponine). Standardized with SAC. Small amount of oil-soluble sulfur compounds. Various beneficial effects. Well established safety. Heavily researched (>200 papers). Produce allicin in the intestine, but causes loss of epithelial cell at the top of crypts in the ileum.

Water extract of fresh bulb at concentration of 1.0 mg/ml had an

inhibitory effect on the growth of Aspergillus flavus and its aflatoxin

production (Mohamed, 1997).

Daily sulfide (DAS) increases cellular glutathione in a variety of

cells, including those in normal livers and mammary tissue. The ability of

DAS to increase glutathione peroxidase and other scavenging enzymes is

important, in reducing the risk of radiation and chemically induced cancer

(Lisk, 1995; Lih and Wang, 2004).

CHAPTER TWO

MATERIALS AND METHODS

2.1 Experimental details:

These experiments were designed to investigate the effect of fresh

crushed garlic bulbs supplemented diet on plasma lipids of Wistar albino

rats.

2.1.1 Experimental animals:

Twenty male Wister albino rats weighing between 85 – 100 g,

obtained from Medicinal and Aromatic Plants Research Institute were

used in this study. The rats were housed identically in stainless steel

cages in an air room under a 12-h light: dark cycle. All of the rats were

initially fed a standard laboratory diet for at least 7 days to acclimatize to

our laboratory. Tap water was freely available. They were divided into

four groups of five rats each.

2.1.2 The rat basal diet:

A basal diet which fulfilled rats requirements was obtained from

the National Center for Research, Khartoum.

The diet composition includes the following:

Wheat flour 665 g

Meat powder 167 g

Sodium chloride 3 g

Oil 120 g

2.2 Cholesterol supplementation in the rat diet:

2% cholesterol powder was supplemented to the basal diet of rats so

as to induce hypercholesterolemia according to Ali et al. (2000), in all

groups of rat except the control group.

2.3 Preparation of the plant material:

Garlic cloves were obtained from the local market. They were

cleared of any adhering dried material. The edible parts were crushed

then weighted in different concentrations of, 4% and 8% of the basal diet

which then used in the dietary supplement daily.

2.4 Equipments used:

Centrifuge.

Pipettes (Eppedorf).

Heparinized capillary tubes.

Plain containers.

Heparin containers.

The roch diagnostic / Hitachi 92 analyzer.

2.5 Chemicals:

Cholesterol powder (the lab line Ltd).

Lithium heparin.

2.6 Experimental procedure:

The animals were divided into four groups, A, B, C and D. group

B, C and D received a diet containing 2% cholesterol so as to induce

hypercholesterolemia. Then the crushed fresh garlic bulbs were added to

the diet at a concentration of 4% and 8% to groups C and D, respectively.

Group A served as a control and was given a standard diet. The duration

of the experiment was four weeks.

2.7 Blood sampling:

At the end of the experiment, and after an overnight fast (from 6

pm to 9 am) the blood samples were drawn from the orbital plexus by

means of capillary glass tubes according to the methods of Khanna et

al. (1993). Blood was collected in lithium heparin container to separate

the serum. After 10 minutes of centrifugation of blood at 5000 rpm,

supernatant plasma were immediately separated and frozen at -20oC until

it was analyzed for plasma total cholesterol, LDL-cholesterol, HDL-

cholesterol and triglycerides.

2.8 Analytical methods:

All parameters in plasma were measured by using fully automated

apparatus (Hitachi 902) in Research and Laboratory Unit, Khartoum

Teaching Hospital.

2.8.1 The roch diagnostic / Hitachi 902 analyzer:

It is an analyzer which used to report results on various body fluids

samples for a wide range of analytic (Fig. 4). It is fully automated,

computerized and performs potentiometric and photo-metric assays.

It consists of photometric measuring system, analytical processing

unit, screen and printer.

The analyzer characteristics include: 200 tests photometric tests per

hour and refrigerated storage for 40 reagents containers.

2.8.2 Determination of plasma total cholesterol:

Principle:

Free and esterified cholesterol in the sample originate, by means of

the coupled reactions described below:

Cholesterol ester + H2O Cholesterol esterase cholesterol + fatty acid.

Cholesterol + ½ O2 + H2O Cholesterol oxidase cholesterol-3-one + H2O2.

2H2O2 + 4-aminoantipyrine + phenol peroxidase Quinoneimine (pink

color) + 4 H2O.

Fig. (4): The roch diagnostic / Hitachi 902 Analyzer

Reagents:

- Good's buffer (6 – 7) pH.

- Phenol.

- 4-aminoantipyrine.

- Peroxidase.

- Cholesterol esterase.

- Cholesterol oxidase.

- Sodium cholate.

2.8.3 Determination of plasma low density Lipoprotein (LDL):

Principle:

In this method LDL-cholesterol is selectively protected from

lipoprotein lipase by protecting reagent (R1), then in the second step

LDL-cholesterol is released and selectively determined by releasing

reagent (R2).

Cholesterol ester + H2O cholesterol esterase cholesterol + fatty acid.

Cholesterol +½ O2+ H2O cholesterol oxidase cholestenone + H2O2.

2H2O2 catalase 2H2O + O2.

2H2O2+ 4-aminoantipyrine + phenol Peroxidase + Quinoneimine

(blue color complex).

Reagents:

Reagent (1) consists of: - Good's buffer (pH 6.8).



- Catalase.

Reagent (2) consists of:

- Good's buffer (pH 7.0).


- Peroxidase.

- Sodium azide.

2.8.4 Determination of plasma high density lipoprotein (HDL):

Principle:

It is a direct method consisting of two steps:

1. Elimination of chylomicron, VLDL and LDL-cholesterol by

cholesterol esterase, cholesterol oxidase and subsequently cholesterol

catalase.

Cholesterol ester cholesterol esterase cholesterol + fatty acid.

Cholesterol + O2 cholesterol oxidase cholestenone + H2O2.

2H2O2 catalase 2H2O + O2.

2. Specific measurement of the release of HDL-cholesterol, after the

release of HDL-cholesterol by detergent in reagent (2) the intensity of

quinineimine dye produced is directly proportional to the cholesterol

concentration when measured at 600 nm.

2H2O2+ 4-aminoantipyrine + N-(2-hydroxy-3-sulfopropyl)-3, 5-

dimethoxyaniline Peroxidase Quinoneimine pigment (pink color) +

4 H2O.

Reagents:



- Ascorbate oxidase.

- Catalase.



- Surfactants.


- Good's buffer (pH. 7).

- Sodium azide.


- Peroxidase.

2.8.5 Determination of plasma triglycerides:

Principle:

They were determined after enzymatic splitting with lipo-protein

lipase, the indicator is Quinoneimine which is generated from 4-

aminoantipyirne and 4-chlorophenol by hydrogen peroxide under the

catalytic action of peroxidase.

Triglycerides Lipoprotein lipase glycerol + fatty acid.

Glycerol + ATP Glycerol kinase glycerol-3-phosphate + ADP.

Glycerol-3-phosphat + O2 Glycerol-3-Phosphat oxidase dihydroxyaceton-

phosphate + H2O2.

4-aminoantipyrine + 4-chlorophenol + 2 H2O2 peroxidase Quinoneimine

(blue color) + 4 H2O + HCL.

Reagents:

- ATP.


- 4-chlorophenol.

- Magnesium chloride.

- Peroxidase.

- Glycerol kinase.

- Glycerol-3-phosphat oxidase.

- Lipoprotein lipase.


2.8.6 Procedure:

For the determination of each parameter, 0.5 ml of the plasma was

put in Hitachi sample cups (Fig. 5).

The sample cups were entered in a specific place on Hitachi 902

and then the parameters to be measured were selected and registered in

the screen. Thereafter, the machine was put on and the results were

printed in ten minutes.

2.9 Statistical analysis:

The data were subjected to completely randomized design.

Analysis of variance (ANOVA) and mean separation were conducted to

test significant differences of groups, according to Gomez and Gomex.

(1984) with the aid of SAS computer programme (SAS, 1998).

Fig. (5): Hitachi sample cups

CHAPTER THREE

RESULTS

The present study was carried out to investigate the effect of fresh

crushed garlic bulbs (Allium sativum) on the level of total cholesterol, low

density lipoprotein, high density lipoprotein and triglycerides in an

induced hypercholesterolemic Wistar albino rats.

3.1 The effect of fresh crushed garlic bulbs on plasma total cholesterol level in an induced hypercholesterolemic Wistar albino rats.

The results of plasma cholesterol levels of groups A, B, C and D

are presented in Table (2) and Fig. (6).

There was a significant increase (P< 0.05) in plasma total

cholesterol level in Group B (cholesterol fed untreated group) compared

to the control group (group A).

There was a significant (P< 0.05) decrease in plasma total

cholesterol level in both groups treated with garlic (group C and D) when

compared to Group B. However, the decrease was more in Group D

compared to group C.

The plasma total cholesterol level in group D was nonsignificantly

different compared to group A. while group C showed a significantly (P<

0.05) higher total cholesterol level compared to group A.

Table (2): The effect of fresh crushed garlic bulbs on plasma total cholesterol, LDL-C, HDL-C and TG in an induced hypercholesterolemic Wistar albino rats.

Parameters

Groups

Total cholesterol

(mg/dl)

Low density lipoprotein

(mg/dl)

High density lipoprotein

(mg/dl)

Triglycerides (mg/dl)

A 68 ± 5.89a 18 ± 1.26a 58 ± 2.92a 60 ± 9.91a

B 174 ± 19.76b 120 ± 16.21b 22 ± 3.03b 83 ± 7.01b

C 133 ± 32.38c 36 ± 10.2a 74 ± 17.5a 73 ± 4.49b

D 90 ± 7.60a 18 ± 1.38a 81 ± 13.41a 50 ± 4.24a

Means ± (SE) within the same column having different superscript small letters are significantly different at (P< 0.05) based on t-test.

Group A: fed standard diet. Group B: fed standard diet with 2% cholesterol. Group C: fed standard diet with 2% cholesterol and 4% garlic. Group D: fed standard diet with 2% cholesterol and 8% garlic.

Fig. (6): The effect of fresh crushed garlic bulbs on plasma total cholesterol level in an induced hypercholesterolemic Wistar albino rats.

Bars with the same small letters are non–significantly different. Significant difference (P< 0.05) between a, b and c. Group A: fed standard diet. Group B: fed standard diet with 2% cholesterol. Group C: fed standard diet with 2% cholesterol and 4% garlic. Group D: fed standard diet with 2% cholesterol and 8% garlic.

3.2 The effect of fresh crushed garlic bulbs on plasma low density lipoprotein level in an induced hypercholesterolemic Wistar albino rats:

The results of plasma LDL-cholesterol levels of group A, B, C and

D are presented in Table (2) and Fig. (7).

The levels of LDL-cholesterol are significantly (P< 0.05) higher in

Group B (cholesterol fed untreated group) compared to the control group

(Group A).

There is a significant (P< 0.05) decrease in plasma LDL-

cholesterol levels in the groups treated with garlic (Groups C and D)

compared to Group B. However, there is no significant difference

between the groups treated with garlic (group C and D) and the control

group (group A). Also there were no significant differences between the

treated groups (C and D).

Fig. (7): The effect of fresh crushed garlic bulbs on plasma low density lipoprotein level in an induced hyper-cholesterolemic Wistar albino rats.

Bars with the same small letters are non–significantly different. Significant difference (P< 0.05) between a and b. Group A: fed standard diet. Group B: fed standard diet with 2% cholesterol. Group C: fed standard diet with 2% cholesterol and 4% garlic. Group D: fed standard diet with 2% cholesterol and 8% garlic.

3.3 The effect of fresh crushed garlic bulbs on plasma high density lipoprotein level in an induced hypercholesterolemic Wistar albino rats.

The results of plasma HDL-cholesterol levels of group A, B, C and

D are presented in Table (2) and Fig (8).

The plasma HDL-cholesterol levels are significantly (P< 0.05)

lower in group B (cholesterol fed untreated group) compared to the

control group (Group A).

The levels of HDL-cholesterol in the treated groups (Group C and

D) were significantly (P< 0.05) higher compared to group B.

The levels of HDL-cholesterol in the treated groups (Group C and

D) are nonsignificantly different compared to the control group (group

A), also there is no significant difference between group C and D with

respect to HDL-cholesterol levels.

Fig. (8): The effect of fresh crushed garlic bulbs on plasma high density lipoprotein level in an induced hyper-cholesterolemic Wistar albino rats.


3.4 The effect of fresh crushed garlic bulbs on plasma triglycerides level in an induced hypercholesterolemic Wistar albino rats.

The results of plasma triglycerides levels of groups A, B, C and D

are presented in Table (2) and Fig. (9).

Plasma triglycerides levels are significantly (P< 0.05) increased in

group B (cholesterol fed untreated group) compared to the control group

(Group A).

There is a significantly (P< 0.05) decreased level of triglycerides in

group D when compared to group B, while no significant difference is

found in group C compared to group B.

The plasma triglycerides level in group D is nonsignificantly

different compared to the control group. While group C showed

significantly (P< 0.05) higher triglycerides level compared to group A.

Fig. (9): The effect of fresh crushed garlic bulbs on plasma triglycerides level in an induced hypercholesterolemic Wistar albino rats.


CHAPTER FOUR

DISSCUSION

Common available garlic preparations in the form of garlic oil,

garlic powder, pills and different extractions are widely used for certain

therapeutic purposes, including lowering blood pressure and improving

lipid profile (Stevinson et al., 2000; Abdel-Wahhab and Aly, 2003 and

El-kayam et al., 2003).

Therefore, this study was designed to examine the effect of fresh

crushed garlic bulbs on the level of plasma total cholesterol and its

fractions (low density lipoproteins and High density lipoproteins) and

triglycerides in an induced hypercholesterolemic Wistar albino rats.

4.1 Plasma total cholesterol:

The results showed that plasma total cholesterol level was decreased

significantly following administration of garlic, however, the higher

concentration (8%) possessed the highest activity. These results were in

line with the reports from medical literature which suggests that

administration of garlic to human, rats and cell culture is effective in

decreasing total cholesterol and triglycerides (Heinle and Betz, 1994; Alie

and Thomson, 1995). Others have also demonstrated the hypolipidemic

effect of aged garlic extract when added to diets of rats fed cholesterol

(Banerjee et al., 2002 and Oue et al., 2003). Reported mechanisms

included directly reduction of cholesterol production by the liver by

inhibition of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-COA)

reductase (Yu-Yan and Lijuan, 2001) and increased bile acid excretion

(Stephen et al., 1993),

4.2 Plasma low density lipoprotein:

In the present study garlic supplemented diet caused significant

reduction in plasma LDL-cholesterol levels in the groups treated with

garlic. These results agree with Shela et al. (2005) who reported that,

adding aqueous extract of raw garlic with 1% cholesterol to rabbits,

decreased plasma total cholesterol and LDL-cholesterol significantly in

the groups treated with garlic, also Merat and Fallahzadeh (1996) and

Tohidi and Rahbani (2000) reported that administration of garlic powder

in the diet reduced LDL-cholesterol significantly. But this finding

contradicted with that reported by Abbas et al. (2005) who reported that

chewing of garlic (crushed garlic) did not affect plasma LDL-cholesterol

levels.

This reduction in LDL-cholesterol level may be due to allicin which

reduces the production and release of LDL-cholesterol by the liver and

promote LDL receptors activity in the liver cells, which helps the liver to

clear the circulating LDL-cholesterol (Holzgartner et al., 1992).

4.3 Plasma high density lipoprotein:

In the current study, feeding fresh crushed garlic bulbs to an

induced hypercholesterolemic rats caused a significant increase in HDL-

cholesterol levels. These findings agree with Aouadi et al. (2000), who

reported that, feeding a supplement with 10% fresh crushed garlic and 2%

cholesterol to rats resulted in a significant reduction in LDL-cholesterol

levels, however, HDL-cholesterol increase significantly. But Abbas et al.

(2005) reported that chewing of garlic (crushed garlic) did not affect

plasma HDL-cholesterol levels.

In reverse cholesterol transport by HDL, cholesterol-ester transferred

to VLDL in a process facilitated by apo D which is known as cholesteryl

ester transfer protein (CETP). Therefore, the inhibition of apo D has been

proposed as a strategy to increase HDL-cholesterol level (Schaefer and

Asztalos, 2006).

Brousseau et al. (2004) reported that, the increase in HDL-

cholesterol level is usually attributed to allicin, which significantly altered

the distribution of cholesterol among HDL and LDL subclasses by a

mechanism appears to be due to a reduction of VLDL level, with a

secondary decrease in apo D activity which results in less transfer of

HDL-cholesterol to VLDL acceptors particles.

4.4 Plasma triglycerides:

The results showed that plasma triglycerides decreased significantly

in the group treated with high concentration of garlic (8%), thus this

group showed control like level. These results are in line with Ali et al.

(2000), who suggested that administration of garlic to rats is effective in

decreasing total cholesterol and triglycerides significantly. The

mechanism for triglycerides lowering effect of garlic is not well

understood. However, Yeh and Yeh. (1994) demonstrated that the rate of

acetate incorporation into fatty acid was reduced in hepatocyte cell

culture treated with garlic extract. Thus, the triglycerides lowering effect

of garlic may some how be due to the inhibition of fatty acids synthesis.

CONCLUSION

It is concluded from this study that fresh crushed garlic bulbs

supplemented diet deceased significantly the levels of total cholesterol,

low density lipoprotein-cholesterol and triglycerides in an induced

hypercholesterolemic Wistar albino rats. While the level of high density

lipoprotein-cholesterol is significantly increased, suggesting that this

garlic preparation could prevent diet-induced hypercholesterolemia and

may has a protective role against atherosclerosis.

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