Dragendorff's reagent- This reagent is constituted of potassium bismuth

iodide. Alkaloids give reddish brown color

Mayers reagent- This reagent is constituted of potassium mercuric iodide.
Alkaloids give cream color with the mayers reagent.
0.5g of (MECC)+ 5ml of 1% aqueous HCl

(Deliorman et al.,2003)
Twenty four hours later, (Rage et al.,1989; Dongare et. al.,2013)
overnight fasting rabbits were deprived of food but allowed for free
access of drinking water. Animal were anaesthesia with ether and
blood was collected by direct cardiac puncture into tubes for
biochemical analysis.
3.6 Biochemical tests
Blood samples was expressed into the sterilized tubes without the
anticoagulant to reduce the risk of hemolysis after removing of the
needles from syringes (Venkatesan R et al., 2006) and allowed for
30 minutes to clot at room temperature. After the clotting of the
blood, it was centrifuged at 4000 rpm (revolutions per minute) for
ten minutes with Hettich Mikro 220R centrifuge (DJB lab care, UK) at
25C for separation of serum. The supernatant was stored in the
freezer at -21oC until analysis (Prochezian and Ansari, 2005)
Serum levels of alanine transaminase (ALT), aspartate transaminase
(AST), alkaline phosphatase (ALP), total proteins, creatinine, urea
and glucose were determined kinetically using UVVIS Spectrophotometer 1601 (Shimadzu, Tokyo, Japan) and commercially available kits.
3.7 Histopathology
After the animals have been sacrificed by cervical dislocation,
the liver was excised, trimmed of fat and other connective tissues,
blotted dry and weighed on a balance. Sections of the liver from the
autopsy samples were stored in 10% formalin and taken to the

Department of Pathology, Science and technology Hospital for the

histopathological studies. The following the standard procedure
described by (Raghuramulu et al ., 1983).
Results
4.3 Effect of MECC against CCL4-intoxication on liver enzymes.
The observed results from serum biochemical parameters of MECC with respect to
CCL4-induced hepatotoxicity will be discussed as following:
The administration of a single dose of CCL4 significantly increased the level of
(SGOT) when compared to control animals. On one hand, there was a significant
reduction in serum (SGOT) values of animals Pre-treated with MECC+CCL4. This
decrease was dose-dependent in manner when compared to CCL4-treated rabbits.
(Figure 4.1)
The activity of SGPT was significantly increased in CCL4 treated rabbits compared
with control animals. In same vein, MECC + CCl4 significantly and dose
dependently reduce serum SGPT values. (Figure 4.2)
The serum of ALP was also significantly increased by the CCL4 administration with
respect to the control rabbits. Also different doses of MECC (500mg /kg b.wt +
CCL4, 1000mg/ kg b.wt + CCL4 and 1500mg/ kg b.wt + CCL4) significantly
decreased serum ALP in dose-dependent manner with regard to CCL4-treated rabbits.
(Figure 4.3)
Together, the activities of SGOT, SGPT and ALP were significantly increased in
rabbits pretreated with MECC + CCL4 in regarded to control. However, the values
did not arrive to the normal. Similar results were also obtained with liv-52 + CCL4

which considered as a standard group. These results indicating hepatocellular damage

in CCL4 group as compared to animals pretreated with MECC and MECC + CCL4 or
those that pre-treated with 5ml/kg b.wt. of Liv-52. There was no significant alteration
in control rabbits treated solely with MECC.
4.4 Effect of MECC against CCL4-intoxication on total protein.
There was a significant reduction in serum total protein of animals
treated with CCL4 compared with control group. But, there was
significantly increasing in total protein of rabbits that received
500mg, 1000mg, 1500mg/kg b.wt of MECC + CCL4 compared with
CCL4 group. These increases were dose-dependently in manner.
(Figure 4.4)
The total protein of rabbits pretreated with liv-52 + CCl4 have been
increased significantly in regarded to toxic control. This protective
effect almost close to the normal group
4.5 Effect of MECC against CCL4-intoxication on urea and
creatinine levels.
The results exhibited significant increase in blood urea nitrogen
(BUN) in animals received CCL4 compared with control (Figure
4.5). Also, MECC + CCL4 treatment caused Significantly decreased
in serum BUN in dose-dependently manner. In addition, serum BUN
was significantly declined in Liv-52+CCL4 model when compared to
toxic group.

Treatment of animals with CCL4 alone induced a significant increase

in the serum creatinine compared to normal group (Figure 4.6).
Similarly, there was increase in the serum creatinine values of
animals orally pretreated with MECC at doses 500mg and 1000mg/
kg b.wt + CCL4 or MECC alone compared with normal and CCL4 but
this

increase was

significantly,

not significantly,

while

this

increase was

in rabbits that received 1500mg/kg b.wt of MECC

when compared to normal control

4.6 Effect of MECC against CCL4-intoxication on blood sugar level.

The administration of CCL4 alone induced a significant increase in glucose compared
to control group (Figure 4.7). on other side, orally pretreatment of animals with
MECC+CCL4 and Liv-52+CCL4 reduced the level of glucose significantly in these
groups with respect to rabbits intoxicated with CCL4. Additionally, the level of
glucose in the animals that pretreated with MECC, MECC + CCL4 and Liv-52 was
significantly decreased compared to normal group
.7 Histopathology findings
Liver

morphological

changes

were

scored,

described

and

summarized in Control, ME CC treated, CCl4 treated, CCl4 + ME CC

treated & Liv-52 treated groups (Table 4.3). liver sections of control
group showed normal cellular architecture with distinct hepatic cells,
sinusoidal spaces, and central vein (Figure A).In addition, There
were no abnormalities or histological changes in the livers of normal
rabbits pretreated with methanolic extract of Caralluma Cicatricosa

(Figure B). However, CCl4 application constituted histopathological

changes in the liver of rabbits. These changes were characterized by
dilation of central vein with noticeable hemorrhage. Moreover,
hepatocyte

necrosis,

inflammatory

cells

infiltration,

fatty

degeneration, and hydropic degeneration were found in rabbits 24

hours after CCl4 treatment (Figure C&D). However, The liver
sections of the rabbits received methanolic extract of CC and Liv552,

as

oral

pretreatment

followed

by

Carbon

tetrachloride

intoxication showed less histopathological changes in a dose

dependent manner when compared to CCl4 group (Figure E, F,
J,H).Results from the histological studies were in agreement with
the measured activities of serum enzymes.
a

Livers were scored for hepatic injury via light microscopy with

score 0 = no visible cell damage; score1 = focal hepatocyte

damage on less than 25 % of the tissue; score 2 = focal hepatocyte
damage on25-50 % of the tissue; score 3 = extensive, but focal,
hepatocyte lesions; score 4 = global hepatocyte necrosis