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Nigerian Journal of Physiological Sciences
Physiological Society of Nigeria
ISSN: 0794-859X
Vol. 24, Num. 1, 2009, pp. 79-83
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Nigerian Journal Of Physiological Sciences, Vol. 24, No. 1, 2009, pp. 79-83
Phytochemistry And Some
Haematological Changes Following Oral Administration Of Ethanolic Root Extract
Of Gonglonema latifolium In Rats
A. B. Antai, O. E. Ofem, D. E.
Ikpi, S. Ukafia1, And E. A. Agiang2
Department
of Physiology, College of Medical Sciences, University of Calabar-Nigeria. 1Department
of Anatomy, Faculty of Health Sciences, University of Uyo, Uyo, Nigeria. 2Department of Animal Sciences, Faculty of Agriculture, University of Calabar, Calabar, Nigeria. E-mail: eduanwana@yahoo.com
Code Number: np09013
Summary
Gonglonema latifolium is a climbing perennial
plant that belongs to the family of asclepidaceae. The medicinal uses of the
leaves and stem bark extract of Gonglonema latifolium has been reported
in many scientific literatures to include anti-hyperglycaemic, anti-ulcerative
etc. There is paucity in scientific reports on the medicinal properties of the
root extract. This study was therefore designed to elucidate the phytochemical
constituents and effect of ethanolic root extract of Gonglonema latifolium on some hematological parameters. Eighteen (18) albino Wistar rats were
randomly assigned into 3 groups of 6 rats each. Group 1 (control) was fed on
normal rat chow + drinking water, the test groups (group 2 and 3) in addition
received extract treatment (p.o) at a dose of 100mg/kg and 200mg/kg
respectively. The feeding regimens lasted for 14 days. Results show that the
extract contains polyphenols, glycosides, reducing sugars and alkaloids. The
mean RBC count for control was 4.44 ± 0.15 x106cells/mm3,
this did not differ significantly from the test groups. The total WBC counts
were 3.79 ± 0.15 x103cells/mm3, 3.37 ± 0.19 x103cells/mm3 and 5.54 ± 0.39 x103cells/mm3 for groups 1, 2, and 3 respectively.
WBC was significantly (P<0.01) higher in the high dose (group 3) compared
with control. Mean PCV (%) were 33.60 ± 0.81, 37.20 ± 0.80 and 34.80 ± 1.35 for
groups 1, 2, and 3 respectively. The PCV was significantly (P<0.05) higher
in low dose extract recipients (group 2) than in control. Hb content, MCV and
MCH did not differ significantly among the groups, but MCHC was significantly
lower in the test groups compared with control group. Neutrophils were
significantly (P<0.001) reduced in the test groups, while the eosinophils
and monocytes count were significantly increased in the test groups than in
controls. In conclusion, the extract has little or no effect on RBC count, PCV,
MCV and MCH. But low doses of the extract significantly increased the Hb count
while high doses significantly increased WBC count, probably due to increase in
eosinophil and monocyte counts. MCHC was dose-dependently reduced. The extract
was also observed to contain polyphenols, alkaloids, glycosides and reducing
sugars.
Key words: Gonglonema latifolium, root extract, Phytochemistry, blood.
Introduction
Gonglonema latifolium is a climbing perennial plant that belongs to the family
of asclepidaceae (Okafor and Ejiofor, 1996; Eleyinmi et al, 2006). It is
a rainforest plant which has been traditionally used in the south-eastern part
of Nigeria for the management of diseases such as diabetes, high blood pressure
etc. (Ugochukwu et al, 2003). Gonglonema latifolium is also used
in the western African sub-region for a number of medicinal and nutritional
purposes such as spice and vegetable (Dalziel, 1937). It is commonly called
Utazi and Arokeke in the south-eastern part of Nigeria. The plant is
traditionally used in the control of weight gain in lactating women and
promotes fertility in women (Schneider et al, 2003). It is also used to
treat malaria, stomach ache, worm infestations, cough, ulcers, and cancers. The
hypoglycaemic and anti-hyperglycaemic properties of the ethanolic stem extract
of Gonglonema latifolium have been reported by Farombi (2003).
Gonglonema latifolium leaves extract is rich in proteins (27.2%DM) which
compared well with values reported for chickpea (24.0%DM) and other protein
rich plants. Phytochemical analysis of leaves extract of Gonglonema
latifolium reveals the presence of essential oil, saponins (asterglycosides),
alkaloids, minerals like calcium, phosphorus, magnesium, copper and potassium
(Schneider et al, 2003; Eleyinmi and Bressler, 2007).
Blood is a tissue which consists of fluid plasma in
which are suspended a number of formed elements (erythrocyte, leucocyte and
thrombocytes). Its primary function is to provide a link between the various
organs and cells of the body, and to maintain a constant cellular environment
by circulating through every tissue delivering nutrient to them and removing
waste products (Bowman and Rand, 1980). The blood cells (erythrocytes,
leucocytes and thrombocytes) exist at fairly constant levels, suggesting the
existence of feedback mechanism for the cells (Guyton and Hall, 2006). The
effects of ethanolic stem extract of Gonglonema latifolium have been
widely reported but there is paucity in scientific literature on the effect of
the root extract, especially on hematological parameters. It is therefore the
aim of this study to investigate the effect of ethanolic root extract of Gonglonema
latifolium on some blood parameters and its biochemical constituents.
Materials
and Methods
Experimental
animals
Albino Wistar rats (weighing between 180-210g) were
used for this study. They were obtained from the animal house of the Department
of Physiology, University of Calabar-Nigeria and housed at room temperature (28
+ 2°C).
Experimental
plant
Fresh mature roots of Gonglonema latifolium
were obtained from the botanical garden of University of Calabar-Nigeria and
were identified by the Chief Herbarium Officer of Botany Department, University
of Calabar-Nigeria.
Preparation
of plant extract
The roots were washed to free debris, sun dried. They
were later oven (Astell Hearson, England) dried at 35°C-40°C.
The dried roots were ground to fine powder and 500g powder was percolated in
300mls of ethanol (80%v/v, BDH) for 24 hours. It was thereafter filtered with
Whatman No. 1 filter paper. The filtrate was then dried to paste at 45°C.
This resulted in 56g (yielding 11.20%) of the crude ethanolic extract. A stock
solution of 1g/ml was prepared for the experiments.
Experimental
protocol
Eighteen (18) albino Wistar rats were randomly
assigned into 3 groups of 6 rats each. Group 1 was the control group while
groups 2 and 3 were the test groups. They were all fed on normal rat chow and
drinking water. Groups 2 and 3 in addition received 100mg/kg (Low dose) and 200mg/kg
(high dose) of ethanolic extract of Gonglonema latifolium extract
(orally, once daily) respectively. The feeding regimens lasted for 14 days.
Collection
of blood samples
Rats were sacrificed using chloroform anaesthesia.
Blood samples were collected by cardiac puncture into EDTA capped bottles with
the aid of a 5ml syringe. The blood samples were then used for the experiments.
Phytochemical
screening of Gonglonema latifolium
The different phytochemical tests were carried out
using standard laboratory techniques. Alkaloids, glycosides (Salkowski test) and
saponins (Frothing test) were identified following the method of Sofowora, (1984).
The presence phlobatanins, anthraquinones, flavonoids and tannins were tested
using the method of Trease and Evans, (1989). Identification of polyphenols and
reducing compounds was by the method of Gullei, (1982).
Determination
of hematological parameter
Red blood cell (RBC) count was done using the
conventional method of Dacie and Lewis (2001). Blood was diluted to 1:200 with
Hayems fluid which preserved the corpuscles and then counted with a Neubauer
counting chamber under a light microscope. The counting of total white blood
cells was done after the method of Brown (1974) using a diluting fluid (Turks
fluid) in a ratio of 1:20. Differential white blood cell count was carried out
using Leishmans stain. The conventional method (using Sahlis
haemoglobinometer) was employed for estimation of hemoglobin (Hb) content of
the blood while packed cell volume (PCV) was done using the macrohaematocrit
method (Dacie and Lewis, 2001).
Calculation
of absolute values
The different absolute values: mean corpuscular volume
(MCV), mean corpuscular hemoglobin (MCH) and mean corpuscular hemoglobin
concentration (MCHC) were calculated from values of RBC, PCV and Hb as follows:
MCV (millimicron) = PCV% x 10 / RBC
count (x million per mm3); MCH (picogram) =Hb
g/dl x 10 / RBC count (x million per mm3) and
MCHC (picogram) = Hb g/dl x 100 / PCV %
Statistical
analysis
All data are presented as mean + SEM. The one
way ANOVA was used to analyze the data, followed by a post-hoc test (LSD). The
results were considered significant at p values of less than 0.05.
RESULTS
Phytochemical
screening of Gonglonema latifolium extract
The result of phytochemical screening of ethanolic
root extract of Gonglonema latifolium is shown in Table 1. The results
show that the root extract of Gonglonema latifolium contains polyphenols
in abundance. Alkaloids, glycosides and reducing sugars were present in
moderate amounts while saponins, tannins, flavonoids, phlobatanins,
anthraquinones and hydroxymethyl anthraquinones were absent.
Hematological
parameter in control and test groups
As summarized in table 2, the mean RBC counts were
4.44 ± 0.15, 4.90 ± 0.26 and 4.63 ± 0.12 x103cells/mm6 for
the control, low and high dose extract fed groups respectively. Although,
increases in RBC counts were observed in the low and high dose group, these
were not statistically different from the control groups.
The mean total WBC count in
control group was 3.79 ± 0.15 x103 cells/mm3. It was
significantly (P<0.01) higher in the high dose recipients (5.54 ± 0.39 x103cells/mm3)compared with control and low dose recipients which had a total WBC of
3.37 ± 0.19 x103cells/mm3.
The mean PCV was significantly higher in the low dose
recipients (37.20 ± 0.80 %) than in control (33.60 ± 0.81 %). The mean PCV of
the high dose group (34.80 ± 1.35 %) was not significantly different to that of
the controls, (table 2).
Table
1: Phytochemical screening of Gonglonema latifolium extract
SN |
Chemical constituents |
Score indication |
1
2
3
4
5
6
7
8
9
10
|
Alkaloids
Glycosides
Saponins
Tannins
Flavonoids
Reducing sugars
Polyphenols
Phlobatanins
Anthraquinones
Hydroxymethyl-anthraquinones |
+
+
-
-
-
+
++
-
-
- |
Table
2: Hematological parameters in the different experimental groups of rats.
Parameters |
Group 1
(Control) |
Group 2
(Low dose) |
Group 3
(High dose) |
RBC (x106/mm3) |
4.44 ± 0.15 |
4.90 ± 0.26 |
4.63 ± 0.12 |
WBC
(x103/mm3) |
3.79 ± 0.15 |
3.37 ± 0.19 |
5.54 ± 0.39**, b |
PCV
(%) |
33.60 ± 0.81 |
37.20 ± 0.80* |
34.80 ± 1.35 |
Hb
(g/dl) |
14.06 ± 1.30 |
14.45 ± 0.16 |
13.38 ± 0.43* |
MCV
(fl) |
76.04 ± 3.58 |
76.56 ± 3.39 |
75.65 ± 4.58 |
MCH
(pg) |
31.80 ± 1.28 |
29.81 ± 1.58 |
29.09 ± 1.63 |
MCHC
(%) |
41.90 ± 0.71 |
38.88 ± 0.57* |
38.51 ± 0.53** |
*P<0.05,
**P<0.01 vs control. b = P<0.01 vs low dose. Values are mean ± SEM, n =
6.
Table
3: Differential white blood cell count in the different experimental groups of
rats.
Parameters |
Group 1
(Control) |
Group 2
(Low dose) |
Group 3
(High dose) |
Neutrophils
(%) |
23.20 ± 1.28 |
13.60 ± 0.40*** |
15.60 ± 0.51***, a |
Lymphocytes
(%) |
64.00 ± 0.71 |
68.40 ± 0.40*** |
62.20 ± 1.02c |
Basophils
(%) |
0.60 ± 0.30 |
0.40 ± 0.30 |
0.40 ± 0.30 |
Monocytes
(%) |
8.40 ± 0.93 |
7.80 ± 0.70 |
14.20 ± 0.70***, c |
Eosinophils
(%) |
4.80 ± 0.37 |
9.80 ± 0.58*** |
7.60 ± 0.60***, a |
***P<0.01
vs control. a = P<0.05, c = P<0.001 vs low dose. Values are mean ± SEM,
n = 6.
The mean control Hb concentration was 14.06 ± 1.30
g/dl. The Hb concentration of the high dose group was significantly (P<0.05)
lower compared with controls. The increase in Hb observed in low dose group was
not significantly different from controls.
Also, shown in table 2, are the mean values of MCV for
the control, low and high dose groups which were 76.04 ± 3.58, 76.56 ± 3.39 and
75.65 ± 4.58 fl respectively. These values were not significantly different
from each other. The mean values of MCH were also not significantly different
among the groups. But the MCHC was significantly higher in the low (38.88 ±
0.57%, P<0.05) and high (38.51 ± 0.53, P<0.01) dose groups compared with
controls.
Differential
white blood cell counts in control and tests groups
The mean values of neutrophils in low and high dose
groups (13.60 ± 0.40 % and 15.60 ± 0.51% respectively) were significantly
(P<0.001) low compared with controls (23.20 + 1.28%). The high dose
group in turn had a significantly (P<0.05) higher neutrophils count than the
low dose group, (table 3).
The mean value of lymphocytes in the low dose group
(68.40 ± 0.40 %) was significantly (P<0.001) higher compared with values
obtained for control (64.00 ± 0.71 %) and high dose (62.20 ± 1.02 %) groups,
(table 3).
Eosinophils were not significantly different among the
groups. Monocyte counts was significantly (P<0.001) higher in the high dose
extract recipient group than in control and low dose groups. The test groups
had significantly (P<0.001) higher basophils than controls; the high dose
group in turn had a significantly (P<0.05) lower counts than the low dose
group, (table 3).
Discussion
The phytochemical constituents and effect of ethanolic
root extract of Gonglonema latifolium on some hematological parameter in
rats were studied. The results obtained showed that the extract contains an
agent with profound effects on blood parameters. The results show that the root
extract of Gonglonema latifolium contains polyphenols in abundance with
moderate amounts of alkaloids, glycosides and reducing sugars while saponins,
tannins, flavonoids, phlobatanins, anthraquinones and hydroxymethyl
anthraquinones were absent compared with the leaves extract which was reported
by Schenider et al (2003) to contain essential oil, saponins
(asterglycosides) and alkaloids.
From the results obtained, the control values of red
blood cells obtained agreed with what was reported elsewhere (Eastham et al,
1993; 1983). However, red blood cells were not significantly affected by Gonglonema
latifolium root extract in the doses given in this study.
The picture for PCV was not clear as there was no
difference between the high dose and control groups but significant increase
occurred between the control and the low dose group. The reason for this is not
quite clear.
Hemoglobin concentration in this study agreed with
report of Dacie and Lewis, 2001 conducted in rats. Paradoxically, there was a
slight but significant decrease between rats that received high and low doses
of Gonglonema latifolium root extract, even though the former did not
differ from controls.
White blood cell count in the three groups studied
fell within the reference range as reported for rats (Constable, 1963). The
extract appeared to increase WBC count in a dose dependent manner, although,
the increase in the low dose compared to controls was not significant. The
effect of the extract on the total WBC count could be due to the presence of
glycosides. This compound has an anti-inflammatory property and so has vital
effect on inflammatory processes of some pathological states such as bacterial
infection, malaria and liver diseases (Ugochukwu, 2002).
The differential WBC count shows variations in the
different parameters. The results obtained for neutrophils in the extract
treated group agreed with the report that leucopenia occurs in some
pathological states (Baker and Silverton, 1998). The lymphocytes in the three
experimental groups studied fell within the reference range in rats (Constable,
1963). There was significant increase in the low dose treated group compared to
the control and high dose groups. This is not dose dependent. Results obtained
in this study for monocytes and eosinophils agree with values stated by Guyton
and Hall (2006). They were significantly higher in the test groups compared to
control. MCV and MCH did not differ significantly among the groups. MCHC in
the test groups decreased significantly than in controls.
In conclusion, Gonglonema latifolium ethanolic root
extract has little or no effect on the RBC count and haemoglobin content of
blood but causes an increase in total WBC count probably by increasing
monocytes and eosinophils in the blood. The extract was also observed to
contain abundant polyphenols and moderate amounts of alkaloids, glycosides and
reducing sugars which would have contributed to the observed effect of the
extract on the blood parameters.
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