• During metabolism, some oxygen molecule escape and become highly
unstable and reactive particles called free radicals.
• Free radical are essential for your survival. They perform many
functions including controlling your blood flow, fighting infection, even
killing cancer cells.
• Fortunately, the body maintains a sophisticated system of chemical
and biochemical defenses to control and neutralize free radicals.
Chemical antioxidants scavenge free radicals, that is, they stabilize
the unstable free radicals by giving them the electron they need
9. Oxidative stress
• Oxidative stress, arising as a result of an imbalance
between free radical production and antioxidant
defenses, is associated with damage to a wide range
of molecular species including lipids, proteins, and
• Hyperlipidemia is a term that refers to high level of
lipids (fats, cholesterol and triglycerides) circulating in
the blood. These lipids can enter the walls of arteries
and increase the risk of atherosclerosis
• Atherosclerosis is a disease process leading to
hardening and narrowing of arteries. The buildup of
fat, cholesterol, and other substances creates plaques
inside arteries, which can lead to many problems
including heart attack, stroke, and death
15. Aim of the study
• the aim of this study was to determine
the antioxidant activity of the two
extracts(methanol and acetone)from
olive leaves (OLE) , also to investigate
the hypolipidemic effect of the
methanol extract on experiment animals
• Plant material
The leaves of olive, were collected from Al-Jabal Al
Akhdar area in Libya during 2017. The leaves were
dried in laboratory and powder in a mixture grinder
eighty four healthy adult male albino mice weighing
between 20-40 g were used for this study
18. Olive leaves
Olive leaf is the leaf of the tree (olea europaea)
- Elenolic acid
• The benefities of olive leaves have been used in
traditional medicine practices as folk remedies for the
treatment of variouse illnessess.
• Oleuropein a polyphenol with unique health-
• These extracts have been used in traditional medicine
for centuries to improve age related disease
• Sample preparation (extraction
The extract was obtained by macerating 50 g of the
dried leaves from olive leaves separately in
acetone(500 ml) and methanol (500 ml) for 3days.
The resultant extract was filtered, concentrated to
dryness in avacumme under reduced pressure at 40
°C, and then stored at -8 °C until use
23. Extract analysis
• Extract analysis
• GC-MS analysis.
• Measurement of the antioxidant activity of the methanol
extract by estimate:
-Total Phenolic Content.
-Total flavonoid content.
-Reducing Power assay.
-Scavenging Activity by DPPH-Radical
- Determination of lethal dose LD50
24. Induction of hyperlipidemia
• Hyperlipidemia was induced in male Swiss albino
mice by using cholesterol/cholic acid mixture (3:1)
and mixed with the synthetic diet in a dose calculated
on the basis that each mouse was received 0.5g of
this mixture/kg b.w daily for 2 weeks.
27. The prophylactic effect of
extracts against hyperlipidemia
• To study the protective effect of the
oils against hyperlipaemia, a total of
36 mice were used and the experiment
lasted for 2 weeks. Animals were
divided randomly into three groups
(12 mice each) as follows:
28. Group 1: mice were fed on the standard synthetic diet
(S.d) and served as negative control group (-ve).
Group 2: mice were daily attained to the hyperlipidemic
diet (H.L.D) and served as positive control group (+ve).
Group 3: mice were daily administered methanol extract a dose
of 500 mg/kg b.w oral (added to the H.L.D).
29. The curative effect of extracts on
• In this experiment, a total of 48 mice were used. 12 mice were
fed on the standard synthetic diet and served as negative
control(- ve)" Group1 "
• .The other mice were subjected to the induction of
experimental hyperlipaemia for 2 weeks as described before.
• The hyperlipidaemic mice (
) were divided randomly into
equal 4 sub-groups (12 mice each) as follows:
Group 2:mice were served as hyperlipidaemic animals(+ve)
Group 3:mice were daily received methanol extract at a dose of
(500 mg/kg b. w) (oral+ S.d).
Group 4:mice were daily received of Atorvastatin as a standard
hypolipidaemic agent 0.9 mg/kg b.w. (oral)
30. Blood sampling
• In the first experiment, blood samples were collected
before treatment and then after 2 weeks. In the second
experiment, blood samples were collected before and after
induction of hyperlipidemia and then again 2 weeks after
the administration of the different treatments.
31. Biochemical analysis
Determination of serum total cholesterol.
Determination of serum HDL-Cholesterol.
Determination of serum LDL-Cholesterol and VLDL-
Determination of serum triglycerides.
Determination of S. Alanine aminotransferase (ALT).
Determination of S. Aspartate aminotransferase (AST).
Determination of serum alkaline phosphatase (ALP).
Determination of serum total protein (TP).
Determination of serum Gamma-Glutamyltransferase (GGT).
32. Determination of superoxide dismutase (SOD)
Determination of Glutathione Reductase (GR).
Determination of Glutathione Peroxidase (GPx).
Determination of Catalase (CAT).
Determination of Malondialdehyde (MDE)
33. At the end of experiments, animals
in all groups were
dissected for postmortem
histopathological studies. Liver,
heart were removed and fixed in
10% neutral formalin.
42. The DPPH radical scavenging
• The results of the DPPH radical scavenging activity
of acetone and methanol extracts are shown in the
figure, these results compared with the well-known
antioxidant ascorbic acid where the percent of the
inhibition is 96.7% at 500 µg/ml of the vitamin C ;
98% at 500 µg/ml of the methanol extract and 95%
at 500µg/ml of acetone extract.
44. the chemical composition of the acetone
extracted from olive leaves.
As can be seen from GC.MS compounds
representing about 99.78%. The major
components are as follows: α–pinene
(34.70), Oleuropein (24.70), 2,6-
67. The liver tissue of control
mice , showing normal
The liver tissue of positive control
mice,showing micro vesicular fatty
change distributed throughout the
liver lobules accompanying with
68. The liver tissue of mice
treated the methanol
extract of olive leaves
(500mg/kg b. w ),
showing normal histological
change with reduction in fat
deposits in liver tissues
69. The heart tissue of control
mice for 2 weeks , showing
The heart tissue of The
positive control mice for 2
weeks , showing mild
groups of fatty cell
70. The heart tissue of group treated with the treated the
methanol extract of olive leaves (500mg/kg b. w ) for 2
weeks, showing vacuolar changes in myocardial
muscles and improves compared to the positive group
71. The kidney tissues of control
group showing normal renal
structure with regulatednuclear
arrangement of uriniferous
The kidney tissue of positive control
mice degenerative changes in
epithelial lining of convoluted tubules
72. The kidney tissue of mice treated the
methanol extract of olive leaves (500mg/kg b. w
), showing significantly improved renal
morphology compared to the positive group
91. Hepatic tissue suffering hyperlipidemia
showing fatty liver tissue with disrupted
celvacuolated cytoplasm and necrosis
with disrupted hepatic strands (arrows)
Hepatic tissues of control group
showing hepatic strands of cells
92. Hepatic tissues pretreated
from olive leaves shows be
restoring the normal
hepatic strands (arrow).
Hepatic tissues pretreated
Atorvastatin (0.9mg/kg b.
w)shows no necrotic
hepatic tissues with normal
93. • Heart tissues of control group
showing normal structure
of cardiac muscles consist of
94. The heart tissue of methanol
extract of olive leaves (500 mg/kg
.b.w.) for 2 weeks, showing
vacuolar degenerative changes
in myocardial muscles
Representing the heart
of positive control mice for
2 weeks and showing
mild groups of fatty cells
The heart of Atorvastatin treated
(0.9mg/kg.bw) for2 weeks, showing
dilated and congested Blood
vessels (arrow) .
95. Renal tissues of control group
showing normal renal structure
The kidney of positive control mice
revealing focal areas of vacuolar
degenerative changes in epithelial
lining of convoluted tubules (arrow).
aphotomicrograph of a kidney
section of methanol extract of olive
leaves treated (500mg/kg. b.w.),
showing few aggregates of
inflammatory cells (arrow) in
perivascular channels and hyaline
casts (arrow head) in the Lumen of
96. The kidney of atorvastatin treated (0.9mg/kg.b.w.) for 2
weeks, showing few calcium depositis (arrow) in some
number of collecting tubules
• The results obtained revealed that the methanol
extracted from olive leaves has an antioxidant activity,
in addition to prophylactic and curative effect
against hyperlipidemia compared with the reference
• In general, to use these extracted as safe prophylactic
and curative agents, more studies should be carried
out to know all the active / inactive components and
their mechanism of actions