In Vivo Trypanocidal activity of Hydroethanolic extract of Hymenocardia acida stem bark in Rats 1 2

The in vivo trypanocidal efficacy of Hydroethanolic extract of Hymenocardia acida stem bark was evaluated in Wistar rats. Three groups of rats were treated orally with the extract at doses of 100, 200 and 400 mg/kg body weight for 6 days. Two other groups received the vehicle and Diminazene accurate at 3.5 mg/kg to serve as negative and positive control respectively. The mean survival period of infected animals, daily level of parasitaemia, packed cell volume, total and differential leukocyte counts were evaluated. Oral administration of the extract did not significantly (P > 0.05) affect the packed cell volume. However, the extract reduced the level of parasitaemia and prolonged the life span of infected rats. This study shows in vivo potential of hydroethanolic extract of H. acida in the treatment of African trypanosomosis.


Introduction
been in use for well over 40 years.Thus, the search for medicinal plants with trypanocidal activities is of In developing countries, the use of products of research interest (Hoet et al., 2004).plant origin by persons with limited access to modern Several reports indicate antitrypanosomal medicines to treat human and animal diseases is activity exits in some medicinal plants (Ibrahim et al., widespread.Herbalists and herdsmen in North Central 2008 and Shuaibu et al., 2008).Also there have been Nigeria have claims to the use of medicinal plants in previous reports of in vitro antitrypanosomal efficacy the traditional management of African trypanosoof leaf (Hoet et al., 2004) and root bark (Atindehou et mosis (Atawodi et al., 2002;Abu et al., 2009) The stem bark of Hymenocardia drugs (Amaechi, 2001).The poor prospect for a acida was collected within the premises of University vaccine due to antigenic variation of the parasite of Agriculture, Makurdi and authenticated by Mr. (Nantulya and Moloo, 1989) is further compounded Patrick Ekwuno of College of Forestry, University of by unwillingness of the pharmaceutical industry to Agriculture Makurdi, Nigeria.Voucher specimen was develop new compounds because of uncertain and deposited at the College herbarium.unprofitable market or perhaps the localized nature of Preparation of crude extract: The stem bark was the disease.The few commercial trypanocides have washed, air dried at room temperature for one week, pulverized and stored in air-tight container until 1985) and was approved by the Departmental Ethics Committee.required.One hundred grams of powdered material Phytochemical screening: Analysis of major was soaked in 500 ml of 70% ethanol and stirred phytoconstituents was carried out qualitatively using intermittently for 48 hours at room temperature.The standard procedures (Odebiyi and Sofowora, 1978).material was filtered using sterile cotton wool and Trypanocidal activity: The animals were randomly Whatman (No.-1) filter paper; the residue was divided into five groups of five rats each and infected resuspended in the same amount of solvent and then 5 filtered three more times.The pooled filtrates obtained intraperitoneally with T. brucei brucei (1×10 ).The were dried at room temperature under the electric fan.hydroethanolic extract was administered by galvage to o The extracts were stored in air-tight containers at 4 C Groups II, II and IV at daily doses of 100, 200 and 400 until needed.mg/kg body weight respectively for 6 days.Groups I Animals: Twenty five white albino rats of weighing and V received distilled water (vehicle) and 3.5 mg/kg 120 g-150 g were obtained from the College of Health Diminazene aceturate respectively.Parasitaemia was Sciences, Benue State University Makurdi, Nigeria.monitored daily in blood obtained from the tail of the The animals were kept in polypropylene cages under rats.A wet film and haematocrit buffy coat methods room temperature, with 12-hour light and 12-hour (Murray et al., 1983) were used for the initial detection dark cycle and were allowed to acclimatize for two of parasitaemia.The degree of parasitaemia was weeks.The animals were provided commercial feed estimated as previously described (Herbert and (Grand Cereals and Oil Mills Ltd, Bukuru, Jos, Lumsden, 1976).Packed cell volume and other Nigeria) and clean water ad libitum.Protocols for this haematological parameters were determined experiment was in accordance with the guidelines on according to the standard method (Schalm et al., the care and well being of research animals (NIH, 1975).The mean survival periods of the animals were  parasitaemia and concluded that high parasite load Parasite: Stabilates of T. brucei brucei (federe strain) could mask the efficacy of crude extract (Ekanem et were obtained from the National Institute for al., 2008).Efficacy of crude extracts might also Trypanosomosis Research, Vom, Jos, Nigeria.The require administration via parenteral route or in parasites were maintained in the laboratory by passage exceedingly high doses which sometimes causes in mice.Blood harvested from a donor animal at peak toxicity and mortality.In a pilot study, administration parasitaemia was used for in vivo trypanocidal assay.
of the extract at 20 mg/kg body weight intraperi-Statistical analysis: The results are expressed as toneally (i.p) had no effect on the parasites.But at mean ± standard error of mean (SEM) using Graph higher doses i.p. the rats died within 24 hours of Prism statistical package.
administration of the extract.Also the reduced efficacy of crude extract of H. acida could be due to  ,1993).Trypanosome In vivo trypanocidal activity-The parasitaemia of generated reactive oxygen species can also attack red infected, untreated and infected but treated groups are blood cells' membranes, induce oxidation and as shown in Figure 1.In all the groups, parasites were subsequently hemolysis (Karori et al., 2008).Thus, first sighted 4 or 5 days post infection.In Group 1 scavenging the trypanosome associated free radicals (infected, untreated), there were progressive increases may ameliorate anemia.Anemia assessed by a drop in in parasitaemia until the rats died 5 to 6 days post packed cell volume is a consistent feature of infection (0% survival).Results showed that parasites trypanosomosis.in the blood stream of rats treated with Diminazene

enzymatic inactivation of active compounds or
The near stable packed cell volume of rats given aceturate (positive control) were completely H. acida extract orally is suggestive of its potential to eliminated on day 5.The rats remained aparasitaemic ameliorate anaemia (Mpiana et al., 2007).The free and survived beyond the 14 -day observation period radical scavenging activity of H. acida is well (100% survival).The extract did not affect the onset of established (Sofidiya et al., 2006).Leucocytosis due to parasitaemia, but was able to reduce its level and lymphocytosis observed at the outset and leucopenia prolong the lifespan of the treated rats.Oral adminiat the terminal stage of the infection is a phenomenon stration of the extract did not significantly (P > 0.05) associated with waves of parasitaemia (Losos and affect the packed cell volume.The initial increases in Ikede, 1972) and the wax and wear syndrome on the total leukocyte counts were followed by decreases host's immune system (Anosa, 1988).Hymenocardia towards the terminal stage of the infection.There were acida stem bark extract did not induce leucocytosis, no marked differences in lymphocyte, neutrophil, but extended the life span of rats in the treatment monocyte and eosinophil counts respectively.groups compared with the negative control group.The Discussion prolongation of life span in infected groups following oral administration of the crude extract agrees with The prepatent period of 4 to 5 days observed in previous reports (Asuzu and Chineme, 1990; the present study is consistent with earlier findings Abubakar et al., 2005).Some of the infected rats which (Umar et al., 2007).These investigators reported that were treated with the extract still died even when they an acute infection associated with sudden death and had low parasitaemia.Death of rats at low rapid rise in parasitaemia is a feature of this strain of parasitaemia was attributed to the release of extra trypanosome.Trypanosomes have the capacity to cellular factors other than the direct effects of the divide very rapidly resulting in their large population parasites.in the blood stream of host animal within short time.
The trypanocidal property of the extract might The hydroethanolic extract of Hymenocardia be due to the action of one or more constituents.The acida stem bark did not completely eliminate the present study did not involve detailed characterization parasites from the blood stream of infected rats, but and isolation of different compounds that could be only reduced the level of parasitaemia.Several responsible for the observed activity.However, s, researchers made similar observations on reduction in

Table - 1: Phytochemical screening of hydroethanolic extract of H. acida for alkaloids, glycosides, flavonoids, saponins, tannins, terpenoids, anthraquinones and phlobatannins.
, glycosides, flavonoids, 2009).Many natural products exhibit trypanocidal saponins, tannins and terpenoids as shown in Table 1.efficacy by generating free radicals which interfere with the redox balance of the parasite (Atawodi et al., Anthraquinones and phlobatannins were not detected 2003) or by binding on kinetoplast DNA of the in the extract.trypanosomes (Nok et al.