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Journal of Medicinal Materials, 2014, VoL 19, No. 3 (pp. 150 -154)
ANTI-DIARRHEAL EFFECT OF PSEUDERANTHEMUM PALAT1FERUM(NEES) RADLK. IN MOUSE MODEL
Hoang Le Son *, Giang Thi Nuong, Tran Van Minh
International University, Vietnam National University Ho Chi Minh City
* Corresponding author: [email protected] (Received April 17*. 2014)
Summary
Anti-Diarrheal Effect of Pseuderanthemumpalatiferum (Nees) Radik. in Mouse Model This research aimed to investigate the anti-diarrheal effect of ethanol extract of/*, palatiferum at doses/3.o. of 100, 200, 400, 600 and 800 mg/kg body ^^ eight on castor oil-induced diarrhea, small intestinal transit and intra- luminal fluid accumulation secretion in mice. The results showed that the ethanol extract of P. palatiferum at a dose p.o. of 800 mg/kg body weight significantly exhibits the most effect against diarrhea, reducing the numlier of wet stools (inhibition percentage of 41.53%), inhibiting up to 42.01%, in small intestinal transit and eliminating intra-luminal fluid accumulation up to 76.01%. These findings provide scientific evidence that the ethanol extract of P. palatiferum possesses anti-diarrheal property.
Keywords: Pseuderanthemum palatiferum, Anti-diarrheal, Castor oil, Intestinal transit. Intra-luminal fluid.
1. Introduction synthesized drugs have been approved for the Diarrhea is a common cause of death in treatment of diarrhea, the search for new developing countries and ranks second in die drugs of natural origin that can be used as number of fatal children's diseases in the alternatives for ciuing diarrhea is still of world. According to the survey conducted by interest to scientists. In fact, numerous WHO, diarrhea causes about 1.5 million medicinal plants have been ti-aditionally used children under age five mortality, particularly in the treatment of diarrhea, and much in the rural area [1]. Although many research is now being devoted to scientifically
evaluate the potential of medicinal plants against diarriiea.
Pseuderanthemum palatiferum (Nees) Radlk (P. palatiferum) is a medicinal plant belonging to the Acanthaceae family. P. palatiferum was first found in the Cue Phuong forest in Northern Vietnam and expanded throughout the country including the Mekong Delta region [2]. Phytochemical study revealed that P. palatiferum contains flavonoids, triterpenoid s^xjnins, P-sitosterol, stigmasteroL, kaempferol, apigenin, phytol, palmitic acid, and salicylic [3]. Traditionally, the plant is widely used for therapeutic purposes such as diabetes, anti- inflammation and anti-diarrhea. The toxicity of P. palatiferum was also tested in previous study, resulting in no sign of acute toxicity or mortality when rats were treated with the ethanol extract dose up to 2,000 mg/kg body weight [4]. The aim of this study was to assess the anti-dianheal effect of P. palatiferum in mouse model
2. Materials and methods
Collection of plant material and extraction:
Plants of P. palatiferum were collected from Tay Ninh Province, Vietnam in July, 2013.
The plant was identified by Associate Professor Tran Van Minh. A voucher specimen was deposited in the herbariimi of Applied Biochemistry laboratory. Department of Applied Chemistry, School of Biotechnology, International University, Viet Nam National University Ho Chi Minh City. The fresh plants were harvested, rinsed with tap water to remove any type of contamination and flien soaked in 70" EtOH. Washed plants were then dried in the oven at 50 °C until the weight of plants was unchanged. The dried plant materials were ground into the powder by grinder. The ethanol extracts were prepared by soaking 2 kg of the dried plant powder in 10 L of 90' ethanol at room temperatixre for 24 h. Then the extracts were filtered through
a Whatmann filter paper. The filtrate was concentrated under reduced pressure at 55°C for 30 min using a rotary evaporator. The extracts were then collected, kept m Petri dish and stored in a desiccator at room temperature.
Evaluating the antidiarrheal effect of ethanol extract of P. palatiferum on castor oil-induced diarrhea in mice [5,6].' Mice were divided into seven groups of five mice per group. Group I was given 10 mlAg, p.o, body weight of distilled water and served as negative control; groups of II- VI received ethanol extract of P. palatiferum at doses of 100, 200, 400, 600 and 800 mg/kg, p.o, body weight, respectively; group VII was given 2 mg/kg, p.o, body weight of loperamide hydrochloride (Ankiu- Pharmaceuticals Pvt, Ltd) and served as positive control. The diarrhea was induced by administrating 0.4 mL of castor oil orally to mice 30 min later.
Each mouse was kept for observation on a plastic fimnel, the bottom lined with filter paper and observed for the period of 4 h. The parameters observed were the total weight of wet stools, scored and compared with control groups. The results were expressed in percentage of inhibition.
Evaluating the effect of ethanol extract of P. palatiferum on castor oil-induced small intestinal transit in mice [7]; Mice were divided into five groups of five mice per group. Ciroup I was given 10 wJJkg,p.o, body weight of distilled water and served as negative control; groups of II- IV received ethanol extract of P. palatiferum at doses of 400, 600 and 800 mg/kg, p.o, body weight, respectively; group V (positive control) was intraperitoneally injected 5 mg/kg of atropme sulphate. The diarrhea was induced by administrating 0.4 mL castor oil orally to mice 30 min later. All mice were orally administrated 1 mL of standard charcoal Journal of Medicinal Materials, 2014, VoL 19, No. 3 151
(10% charcoal suspension in 5% gum acacia) Ih after castor oil administration. These mice were sacrificed after 1 h and then removed small intestine. The peristaltic index (PI), which is the distance travelled by charcoal meal to the total length of small intestine expressed in terms of percentage:
PI = LM/LSlxlOO%
% hhibition = (Control - Test)/Contn)l X 100 PI = Peristaltic index (%)
LM = Length of charcoal meal (cm) LSI = Length of small intestine (cm) Evaluating the efTect of ethanol extract of P. palatiferum on castor oil-induced small intestinal secretions by enteropooling assay [7]: Mice were divided into five groups of five mice per group. Group 1 was given 10 mL/kg,/j.o, body weight of distilled water and served as negative control; groups of 11 IV received ethanol extract of P.
palatiferum at doses of 400, 600 and 800 mg/kg, p.o, body weight; group V was intraperitoneally mjected 30 mg/kg, body weight of chlorpromazine and served as positive control. The diarrhea was induced by administrating 0.4 mL of castor oil orally to
mice 30 min later. All mice were sacrificed 30 min later and the small intestine was removed. The difference mean weight of intestine in control and castor oil-treated group was considered as the castor oil - induced accumulation of intestinal fluid.
Intra luminal fluid accumulation was calculated by the following formula:
Intra-limiinal fluid accumulation = Mean weight of test groups-Mean weight of nomial mice
Statistical analysis: Results were expressed as the mean ± standard error of mean.
Statistical analysis was performed by using SPSS version 16.0. P-Values less than 0.05 were considered to be statistically significant.
3. Results and discussion
Effect of ethanol extract of P. palatiferum in castor oil-induced diarrhea: The ethanol extract of P. palatiferum reduced number of wet stools m a dose-dependent manner (Table 1). After 4h of treatment, the group received 800 mg/kg p.o, body weight of P.
palatiferum showed the highest effect with the inhibition percentage of 41.53%, lower than that of positive control (60.47%).
Table I. Effect of ethanol extract of P. palatiferum on castor oil-induced dianiiea in mice Group
I II III IV V VI VII
Treatment Distilled water (10 mL) Ethanol extract of P. palatiferum (100 mg/kg, p.o, body weight) Ethanol extract of P. palatiferum (200 mg/kg, ;j. 0, body weight) Ethanol extract of P. palatiferum (400 mg/kg,p.o, body weight) Ethanol extract of P. palatiferum (600 mg/kg, p.o, body weight) Ethanol extract of P. palatiferum (800 mg/kg, p.o, body weight)
Loperamide hydrochloride (2 mg/kg, p . 0, body weight)
Mean weight of the stools after 4 h (mg)
38.65±1.07 38.16±2.26 38.5fti2.0l 29.51±l.93'
26.08±5.01 22.60t2.95' 15.28±6.3I'
Inhibition (%)
-
0.08 0.23 7.43 32.52 41.53 60.47 Values are the mean ± SEM for S mice per group
* Significant difference with/j < 0.05 of weigiit of stool level compared to that of control
Journal of Medicinal Materials, 2014, VoL 19, No, 3
Effect of ethanol extract of P. palatiferum on castor oil-induced small mtesOntd transit in mice: Since the ethanol extract of P. palatiferum at doses of 100 and 200 mg/kg, p.o, body weight had no anti- diarrheal effect in mouse model (Table 1), this assay was done using the ethanol
extract of P. palatiferum at doses of 400,600 and 800 mg/kg, p.o, body weight As can be seen fix)m Table 2, mice received 400 and 600 mg/kg, p.o, body weight of P.
palatiferum produced the same effect with the inhibition percentage of 27.08% and 27.73%, respectively.
Table 2. Effect of ethanol extract of P. palatiferum on castor oil-induced intestinal transit in mice Group Treatment Peristaltic index (PI) (%)
I II III rv
V
Distilled water (10 mL) Ethanol extract of P. palatiferum (400
mg/kg,p.o, body weight) Ethanol extract of P. palatiferum (600
mg/kg, p.o, body weight) Ethanol extiuct of P. palatiferum (800
mg/kg,p.o, body weight) Atropine sulphate (5 mg/kg, ip, body weight,)
38.73±6.35 28.24±2.89
Inhibition (%)
-
27.08 27.99±2.99 i 27.73 22.'16±2.72«
15.404826'
42.01 (023 Values are the mean ± SEM for 5 mice per group
• Significant difference withp < 0.05 of (PI) level compared to that of control
Effect of ethanol extract of P. palatiferum respectively, which is almost the same effect on small intestinal secretion: Mice received caused by positive control (50.27%).
400 and 600 mg/kg, p.o, body weight of P.
palatiferum significantly reduced intra luminal fluid accumulation with the inhibition percentage of 48.36% and 49.68%,
Mean\^iiile, mice received 800 mg/kg, p.o, body weight of P. palatiferum eliminate intra - luminal fluid acciunulation up to 76.01%
(Table 3).
Table 3. Effect of ethanol extract of P. palatiferum on castor oil induced intra-luminal fluid accumulation in mice
Group
'
11 ill IV V
Treatment Distilled water (10 mL) Ethanol extract of P. palatiferum (400
mg/kg, p.o, body weight) Ethanol extract of P. palatiferum (600
mg/kg, p.o, body weight) Ethanol extract of P. palatiferum (800
mg/kg, p. 0, body weight) Chlorpromazine (30mg/kg, ip, body weight)
Intra-luminal fluid accumulation (mg)
82.11±234
42.40±0.8 4I.32±0.72 I9.70±0J7' 40.82±1.02
Inhibition (%)
48.36 49.68 76.01 50.27 Values are the mean ± SEM for 5 mice per group
• Significant difference with p < 0.05 of weight of small intestine and intra-luminal fluid compared to that of control
Journal of Medicinal Materials, 2014, VoL 19, No. 3 153
It is well documented that castor oil induce diarrhea by mcreasing the volume of intestinal content by prevention of the reabsorption of water. Ricinoleic acid, the most active compound of castor oil, stimulates peristaltic activity in the small intestme, leading to changes in the electrolyte permeability of the intestinal mucosa Castor oil is also reported to stimulate the release of endogenous prostaglandins E and F which cause stomach cramp and diarrhea due to the effect on the smooth muscle and secretion [8]. Anti- diarrheal activity of medicinal plants has been scientifically validated by investigating the biological activity of plant extracts including antispasmodic effects, delaying intestinal transit, inhibition of gastrointestinal motility, stimulation of water absorption, or reduction of intra-luminal fluid accumulation.
In this study, the anti-diarrheal property of ethanol extract of P. palatiferum at doses of 100, 200, 400, 600 and 800 mg/kg, p.o, body weight has been assessed by assaying the effect on castor oil-induced diarrhea, small intestinal transit and intra-luminal fluid accumulation secretion in mice. The results
of the present study showed that the ethanol exti-act of P. palatiferum at a dose of 800 mg/kg, p.o, body weight exhibits the most effective against diarrhea; though this was not comparable to the positive control (loperamide and atropine sulphate) in the test of anti-diarrheal activity and small intestinal transit. However, in the enteropooling assay, the ethanol extract of P. palatiferum at a dose of 800 mg/kg, p . o , body weight significantly reduces the intra-limiinal fluid accimiulation, more effective than that of positive control.
4. Conclusion
The ethanol extract of P. palatiferum at a dose of 800 mg/kg, p.o, body weight possesses significant anti-diarrheal activity due to its effect on the reduction of wel stools, inhibiting in small intestine transit and eliminating the intra-luminal fluid accumulation on the small intestine of mouse model.
Further study is needed to determine the target components playing a key role in the treatment of diarrhea.
Acknowledgement: This research was supported by the International University, Vietnam National University-HCMC.
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