Journal of Ethnopharmacology 95 (2004) 363366

Diuretic effect of the aqueous extract of Bidens odorata in the rat

Ma. Estela Mel endez Camargo , Blanca Berdeja, Guadalupe Miranda
Departamento de Farmacia, Escuela Nacional de Ciencias Biol ogicas, Instituto Polit ecnico Nacional, Prolongaci on de Carpio y Plan de Ayala S/N, Apartado 42-186, CP 11340, M exico, DF, Mexico Received 27 November 2003; received in revised form 12 August 2004; accepted 12 August 2004 Available online 29 September 2004

Abstract The aerial part of Bidens odorata Cav., are used in Mexican Folk Medicine to treat renal diseases. The aim of this study is to measure the diuretic response to aqueous extract of this plant at doses of 41 and 166 mg/kg and to compare it with that induced by furosemide at 2 mg/kg. Aqueous extracts, furosemide or vehicle were administered orally to adult rats and the effects on sodium, potassium and water balance were assessed. Aqueous extracts at both doses and furosemide produced important and signicant increments in urinary excretion of sodium, potassium and water in rats with respect to control group. This increase was dose dependent and aqueous extract at the highest dose induced a more marked sodium and water excretion than furosemide, potassium excretion increased but it was less than the one induced by furosemide. These results suggest that the aqueous extract of Bidens odorata induce diuretic response. 2004 Elsevier Ireland Ltd. All rights reserved.
Keywords: Water excretion; Saluretic effect; Bidens odorata; Compositae

1. Introduction The use of plants to heal is a common practice among the Mexican population, as well as countries such as Guatemala, specially because of the geographic situation which provides a varied diversity of them and the traditional culture inherited from the Mayas and other ethnic groups (Rzedowski and Rzedowski, 1979; Caceres et al., 1987; Mart nez, 1996). There are a great number of species with potent possibility to produce some therapeutic effect, among them is Bidens odorata Cav. (Compositae). This plant has Cosmos tenellus Kunth, Bidens caucifolia DC. and Bidens rosea Sch. Bip as synonyms (Argueta et al., 1994). Bidens odorata a specie used in Folk Medicine which grows in Mexico. It is locally known as mozote blanco, aceitilla and it is used by the people in several communities for its benecial effects on lungs, renal function, and hypoglycemic action (Mart nez,
Corresponding author. Tel.: +52 55 5729 6300x62341; fax: +52 55 5729 6300x62341. E-mail addresses: mcamargo@ipn.mx, emelendezc@hotmail.com (Ma.E. Mel endez Camargo).

1996; Hern andez, 1984). In Folk Medicine these leaves are often used as an infusion given in the treatment of renal disorders. Due to its popular use, in this study, the diuretic activity on aqueous infusion of Bidens odorata in rats, by means of their water and electrolyte balance, was evaluated. In addition to creatinine clearance, the acute toxicity was determined. The oral route was used because this is the way the people use it.

2. Material and methods 2.1. Plant material The plant material was collected in September in Malinalco, Estado de Mexico. Bidens odorata (CAV), mozote blanco, aceitilla (Compositae) was botanically authenticated by Ma. Eugenia Ordorica (Department of Botany, ENCB). A voucher specimen was deposited in the herbarium of the National School of Biological Sciences, IPN (documentation specimen FLORES 151). The material was shade-dried and powdered.

0378-8741/$ see front matter 2004 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.jep.2004.08.005

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2.2. Preparation of the extract The dried powdered aerial part of Bidens odorata (500 g) was extracted with 1000 ml of distilled water for 5 min. Following ltration and concentration under vacuum, a brown sticky residue was obtained (yield: 17.92%). Phytochemical analysis of this extract was made by the conventional tests (Dom nguez, 1978). 2.3. Animals Adult female Wistar rats were used. They were housed and maintained in the animal house at room temperature (2224 C) and 5055% relative humidity, with day/night cycles of 12 12 h. They were fed with standard rodent diet and water ad libitum. Care and handling of the animals were in agreement with internationally accepted procedures and approved by our institutional committee following the recommendation indicated in the Mexican Technical Specications for the Production, Care, and Use of Laboratory Animals (Secretar a de Agricultura, Ganader a, Desarrollo Rural, Pesca y Alimentaci on, 1999). 2.4. Diuretic activity 2.4.1. Effects of Bidens odorata on water balance in the rat The experiments were carried out in unanesthetized adult female Wistar rats. These were distributed by means of a table of random numbers into four groups: control group received vehicle (distilled water, 1 ml/kg body weight (b.wt.)), furosemide-treated group (positive-control group, 2 mg/kg b.wt.) and two groups treated with the aqueous extract from Bidens odorata at the doses of 41 and 166 mg/kg b.wt. In all groups, the administration was oral and there were at least eight animals per group. At the beginning of the experiment, the urinary bladder was emptied by gentle compression of the abdomen (t = 0), as previously described (Mel endez et al., 1990). In this paper adequacy of this method to obtain complete emptiness of urinary bladder was veried by opening the abdomen and direct inspection of the bladder. Once the bladder was emptied, either the vehicle, furosemide or aqueous extracts were administered. The animals were kept during 6 h in metabolic cages in a quiet and warm environment (2123 C) under uid and food deprivation. At the end of this period, the sample of urine was obtained (t = 6 h) and collected into calibrated pipettes (Dade, Miami, FL), also by abdomen compression to ensure complete emptiness of the bladder. The sample was added to the spontaneously voided urine that was collected under mineral oil and ice-cooled to prevent evaporation. The animals were decapitated and blood samples were obtained into microhematocrit capillaries that were centrifuged al 11,000 rpm (IEC MB centrifuge, Damon/IEC Division, Needham Heights, MA) for 5 min to separate blood cells from plasma.

Osmolalities of plasma and urine were measured in a vapor pressure osmometer (Wescor, Logan, UT) in triplicate. Urine volume was also measured and then osmolal and free water clearances were calculated. 2.4.2. Effect of aqueous extract on electrolyte balance in the adult rat To study the effect of aqueous extract from Bidens odorata, sodium and potassium concentrations were measured in the urine and plasma samples (Flamephotometer Corning 400, Corning Medical and Scientic, England) from rats treated with two doses of aqueous extract. Urinary excretions and clearances of sodium and potassium were calculated. All data were normalized per 100 g of body weight. 2.4.3. Effect of aqueous extract on glomerular ltration rate Glomerular ltration rate (GFR) was estimated by the clearance of endogenous creatinine. To avoid the error due to tubular secretion of creatinine, only female animals, in which no secretion occurs, were used (Harvey and Malvin, 1966). Plasma and urinary creatinine were determined by the Jaffe alkaline picrate method (Mel endez et al., 1991). The following parameters were determined: creatinine clearance corrected for 1 g of kidney, fractional excretion of sodium (FENa ) and potassium (FEK ) were calculated using the conventional equation: FENa = CNa 100 Ccreat

where CNa = UNa V/PNa , and Ccreat = Ucreat V/Pcreat . Filtered sodium and potassium loads were also calculated. 2.4.4. The amount of sodium and potassium in plant extracts The content of Na+ and K+ were determined in both extracts of the plant as described previously. 2.5. Acute toxicity Six groups of NIH female mice of 25 5 g of body weight were formed, a control group (vehicle) and ve groups treated with different doses of aqueous extract (from 350 to 5000 mg/kg of body weight). The animals had access to food and water ad libitum and were observed for clinical signs during 24 h following per os administration. Lethality was assessed using death within 7 days as an index of toxicity (Litcheld and Wilcoxon, 1949). 2.6. Statistical analysis The variance analysis was used to compare the results obtained in the different groups. A difference was considered signicant when p 0.05.

Ma.E. Mel endez Camargo et al. / Journal of Ethnopharmacology 95 (2004) 363366 Table 1 Effect of aqueous extract of Bidens odorata on osmolal and free water clearances, FENa and FEK Control (vehicle) Osmolal clearance (l/min/100 g b.wt.) Free water clearance (l/min/100 g b.wt.) FENa (%) FEK (%) 9.2 0.5 (10) 8.9 0.5 (10) 2.5 0.7 (10) 5.8 1.5 (10) Furosemide (2 mg/kg) (8) 3.6 0.6** (8) 6.9 0.7** (8) 22.2 2.8** (8) 11.9 0.4* Aqueous extract (41 mg/kg) (10) 4.8 0.7* (10) 3.5 0.8* (10) 9.9 1.3* (10) 11.8 0.6*

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Aqueous extract (166 mg/kg) 12.9 0.7* (9) 4.3 0.3* (9) 8.4 0.5** (9) 12.1 0.8** (9)

Mean S.E.M. are shown. Figures in parenthesis indicate number of animals. p < 0.05. p < 0.01.

3. Results The phytochemical analysis showed the presence of alkaloids, reducing sugars and tannins. 3.1. Effect of aqueous extract of Bidens odorata on the water balance in the rat An increase in the urinary volume was observed in treatedextract groups and in furosemide-treated group (p < 0.05). Osmolal clearance increased in the treated groups either with furosemide or aqueous extract at both doses (Table 1, p < 0.05). Free water clearance decreased in both extract-treated groups and in the furosemide-treated group (Table 1), the response in this last group was higher than in the groups treated with aqueous extract. 3.2. Effects of aqueous extract of Bidens odorata on the electrolyte balance in the rat Sodium clearance increased in the groups treated with either aqueous extracts or furosemide. The increment was dependent on the aqueous extracts doses and it was signicantly different to control values (Fig. 1A). The extract increases the potassium clearance in both treated groups, in contrast to furosemide, the plant does not affect the excretion of potassium in the same degree (Fig. 1B), the extract induce a lower effect than furosemide. The basal values of plasma electrolytes were similar in all the groups. There were no signicant changes in the plasma osmolality and hematocrit among the groups (data not shown). 3.3. Effect of Bidens odorata on the GFR and ltered sodium and potassium loads In this study the GFR remained unchanged in all treatedgroups. Changes in the ltered sodium load were produced by increments in GFR at 166 mg/kg of aqueous extract of Bidens odorata, these were not signicant. In the ltered potassium load similar results was obtained (data no shown). 3.4. Sodium and potassium fractional excretions (FENa and FEK ) Signicant increments in FENa and FEK were seen in the three experimental groups, the increments observed in FENa for the treated-group with the aqueous extract of Bidens odor-

ata at dose of 166 mg/kg was higher than furosemide, this was not observed for FEK where the increment was less than furosemide (Table 1). 3.5. The amount of sodium and potassium in extracts of Bidens odorata In the aqueous extracts of Bidens odorata aerial part at the dose of 41 mg/kg, the potassium content was 5 mmol/l, the second dose tested (166 mg/kg) was 21 mmol/l. The content of sodium was 7.2 and 29.4 mmol/l for the dose levels of 41 and 166 mg/kg, respectively. 3.6. Acute toxicity None of the doses of aqueous extract produced mortality or any behavioral disorders.

Fig. 1. Sodium and potassium clearances in rats treated with aqueous extract of Bidens odorata. In panel (A) furosemide and extract of Bidens odorata at both doses elicited signicant increments in the sodium clearance. Mean S.E. are depicted by the heights of the bars and brackets. ** p < 0.01; * p < 0.05. (B) The changes in potassium clearances were measured, furosemide and aqueous extract of Bidens odorata increment potassium excretion, the extract induces a lower effect than furosemide. Mean S.E. are shown. ** p < 0.01; * p < 0.05.

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4. Discussion and conclusions Diuretics are drugs that increase the rate of urine ow and sodium excretion and are used to adjust the volume and/or composition of body uids in a variety of clinical situations, including hypertension, heart failure, nephritic syndrome and cirrhosis. Loop diuretics such as furosemide can increase the urinary ow rate, also they are strongly saluretic inasmuch as they increase urinary sodium and chloride excretion (Jackson, 2001), that is why in this study furosemide was used as positive control. On the other side, the herbal diuretics produce very little acute toxicity and in general they can be considered as mild and good drugs, in comparison to other diuretics used nowadays in therapeutic. The most important thing is that a large number of these plants are rich in potassium, which would not lead to potassium depletion, thus giving the benets of potassium sparing diuretic such as spironolactone, triamterene, etc. (Horisberger and Giebisch, 1987; Jackson, 2001). Through this work, the diuretic activity of the aqueous extract of the Bidens odorata, as described in Traditional Medicine, for the treatment of renal and urinary tract diseases was demonstrated. Bidens odorata induced an increment in the urine output and in the electrolyte excretion at both doses of the extract. These results reveal that the aqueous extract of Bidens odorata is more potent as a diuretic than furosemide, the water excretion is higher in the presence of this extract and the response is dose-dependent. Aqueous extract induces an increment in sodium clearance major to the one obtained with positive control. In contrast to furosemide, extract does not affect in the same degree the potassium clearance, this response is lower in FEK that in FENa , this potassium-sparing effect should be viewed as a favorable feature of the extract in regards to electrolyte excretion (Naik et al., 1981). It is possible that Bidens odorata aqueous extract exerted its diuretic effect by inhibiting tubular reabsorption of water and electrolytes as such action has been suggested for some other plants. The possibility of direct action of potassium content of Bidens odorata extract on diuretic effect is not considered since the K+ content of the extract was very low in comparison with the salt concentration obtained from other plants (Sripanidkulchai et al., 2001). These results are similar to the ones obtain with other species, among them Boerhaavia verticillata (Caceres et al., 1987; Bajpai and Ojha, 2000), these have shown diuretic effect. In the toxicological evaluation, this plant did not present toxic effects at the doses evaluated in this study, due to the results obtained in comparison with the data reported on toxicity of the agents, it is possible to consider that the aqueous extract Bidens odorata is not toxic (Ecobichon, 1992). These experiments justify the use of Bidens odorata as a diuretic agent in Traditional Medicine (Mart nez, 1996; Argueta et al., 1994) as well, its no toxicity was demonstrated.

In conclusion, our results demonstrate that Bidens odorata Cav. (Compositae) administered at both doses induces signicant effects on urinary excretion of water and electrolytes and support the claims of diuretic efcacy extract in Mexican Traditional Medicine.

Acknowledgement This work was partially supported by the Coordinaci on General de Posgrado e Investigaci on del IPN. References
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