Fitoterapia 78 (2007) 182–185www.elsevier.com/locate/fitote

Effect of Mitragyna speciosa aqueous extract on ethanolwithdrawal symptoms in mice

Ekkasit Kumarnsit a,⁎, Niwat Keawpradub b, Watcharin Nuankaew c

a Department of Physiology, Faculty of Science, Prince of Songkla University (PSU) Hat-Yai, Songkhla 90112, Thailandb Department of Pharmacognosy and Pharmaceutical Botany, Faculty of Pharmaceutical Sciences, PSU, Hat-Yai, Songkhla 90112, Thailand

c Southern Botanical Garden in Classical Thai Literature, Chalung, Hat-Yai, Songkhla 90110, Thailand

Received 27 October 2005; accepted 6 November 2006Available online 6 February 2007

Abstract

Administration of the aqueous extract of Mitragyna speciosa at a dose of 300 mg/kg significantly inhibited ethanol withdrawal-induced behaviors that included rearing, displacement and head weaving. The results also showed that at doses of 100, 300 and500 mg/kg M. speciosa showed antidepressant activity without effect on the spontaneous motor activity.© 2007 Elsevier B.V. All rights reserved.

Keywords: Mitragyna speciosa; Ethanol withdrawal; Antidepressant

1. Introduction

Mitragyna speciosa has been used in traditional medicine for decades in many Southeast Asian countries, especiallysouthern Thailand. Originally, local people used it to alleviate pain, coughing or diarrhea. It was also used to counterfatigue [1]. Recently, it has gained much attention because of the claim that it may able to inhibit the withdrawalsymptoms that follow cessation from long-term ethanol consumption and prevent ethanol patients from relapsing.

Ethanol has inhibitory effects on the central nervous system (CNS) by enhancement of gamma-aminobutyric acid-A(GABAA) receptor function [2]. The abrupt cessation of chronic ethanol consumption leads to hyperexcitability of theCNS [3]. Depression, anxiety and associated seizures are commonly observed during ethanol withdrawal syndrome[4–6]. Classical antidepressant and antianxiety drugs have been consistently demonstrated to alleviate the severity ofethanol withdrawal symptoms [7,8]. There have been many studies that have searched for effective natural productsthat will provide alternative methods for the treatment of alcoholism.

An activity on CNS of M. speciosa involving the serotonergic and noradrenergic systems in the MS effect wasdemonstrated [9].

This study was designed to determine the effect of an aqueous extract of M. speciosa on ethanol withdrawalsymptoms.

⁎ Corresponding author. Tel.: +66 74 288216; fax: +66 74446680.E-mail address: ekumarnsit@yahoo.com (E. Kumarnsit).

0367-326X/$ - see front matter © 2007 Elsevier B.V. All rights reserved.doi:10.1016/j.fitote.2006.11.012

183E. Kumarnsit et al. / Fitoterapia 78 (2007) 182–185

2. Experimental

2.1. Plant

Leaves ofM. speciosa Korth (Rubiaceae) were collected from a natural source in Songkhla province, Thailand. Theplants were identified by Dr. Niwat Keawpradub, Department of Pharmacognosy and Pharmaceutical Botany, Facultyof Pharmaceutical Sciences, PSU. The use of the plant material was approved by the Ministry of Agriculture ofThailand and was restricted to research purpose only.

2.2. Animals

Male Swiss albino mice (30–35 g) were maintained under a 12/12 dark/light cycle and controlled temperature(22 °C). Standard commercial food pellets and filtered tap water were available ad libitum. The experimental protocolsdescribed in the present study were approved by the Animals Ethical Committee of the Prince of Songkla University forcare and use of experimental animals.

2.3. Preparation of the aqueous extract

Young leaves of M. speciosa were dried in sunlight and powdered. The aqueous extract was obtained by boiling,then filtered and freeze-dried. The powder (yield: 8.5% w/w) was stored at 4 °C until used.

For all experiments, mice were administered intragastrically with either distilled water or the extract, at the rate of5 ml/kg b.w., 45 min before testing. The behaviors of the animals were recorded by using a digital video camera.

2.4. Tail suspension test (TST)

The tail suspension test was performed according to Steru et al. [10] with minimal modification. Animals weresuspended 50 cm above the floor by using adhesive tape placed approximately 2 cm from the tip of the tail. Thebehavior was recorded for a period of 6 min. The immobile time during the suspension period was measured. Theanimals were considered immobile when they hung motionless or just passively swung without intentional movement.Animals were treated with 100–300 and 500 mg/kg of M. speciosa (MS) extract 45 min before testing. The behaviorsof the animals were recorded by using a digital video camera.

2.5. Induction of ethanol withdrawal and treatment

The animals were administered with ethanol in a modified liquid diet (MLD) by modifying the origi-nal method described by Uzbay and Kayaalp [11]. The MLD contained low fat cow's milk (925 ml), ethanol(25–75 ml), vitamin A (5000 IU) and sucrose (17 g). This solution contained 1000.7 kcal/l. The schedule startedwith all animals receiving control MLD for 7 days. Then, the ethanol-treated animals received an MLDcontaining ethanol at 2.4% (3 days), 4.8% (4 days) and 7.2% (30 days) with different amounts of sucrose addedto make up 1000.7 kcal/l. Then the ethanol-containing MLD was replaced with ethanol-free MLD to induceethanol withdrawal.

Ethanol-dependent animals were divided into 3 groups (non-withdrawal, withdrawal-vehicle and withdrawal-MS).They received orally either vehicle or MS extract at 300 mg/kg 45 min before recording their behaviors. The animalswere individually put into an observable cylinder (15 cm diameter and 50 cm height). The recording was performed fora period of 5 min at 6 and 10 h following the cessation of ethanol.

2.6. Evaluation of spontaneous motor activity

The effect of the MS aqueous extract (100, 300 and 500 mg/kg p.o.) on spontaneous motor activity of mice wasexamined by using the observable cylinder (45 cm in diameter and 50 cm in height). Its floor was divided into 9roughly equal squares. Animals were individually placed into the center of the cylinder and videotaped for 5 min from atop view. Spontaneous motor activity was assessed from the number of squares crossed.

Fig. 1. Effects ofM. speciosa aqueous extract (MS) on the duration of immobility in the tail suspension test in mice. Data were expressed asmean±S.E.M.N=10. *Pb0.05 different from the control group.

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2.7. Statistical analysis

Data are expressed as mean±S.E.M. The effects of the MS aqueous extract on the duration of immobility andspontaneous motor activity were assessed by one-way analysis of variance (ANOVA) and followed by Dunn's method.The effect of the MS extract on ethanol withdrawal was examined by independent t-test for individual behavior.Pb0.05 was considered statistically significant.

3. Results and discussion

Intragastric administration of the MS aqueous extract had a significant effect in the TST [Pb0.05, Fig. 1]. All dosesof the extract (100, 300 and 500 mg/kg) produced a similar and significant approximately 25% reduction of theduration of immobility compared to the vehicle control.

Fig. 2. Effects ofM. speciosa aqueous extract (MS) on ethanol withdrawal behaviors. Withdrawal behaviors were evaluated for a period of 5min at 6 and 10 h(AandB respectively) following the cessation of ethanol consumption.Datawere expressed asmean±S.E.M.N=10. *Pb0.05 different from the control group.

Fig. 3. Effects of M. speciosa aqueous extract (MS) on spontaneous motor activity in mice. Data were expressed as mean±S.E.M. N=10. ⁎Pb0.05different from the control group.

185E. Kumarnsit et al. / Fitoterapia 78 (2007) 182–185

In ethanol-dependent animals, the non-withdrawal group was used as a control. Six hours after ethanol waswithheld, rearing, displacement and head weaving of the withdrawal-vehicle group were obviously induced to morethan two-fold compared to the control group, whereas grooming, lip smacking and scratching were not (Fig. 2A).Interestingly, the withdrawal-MS group showed significantly lower levels of these three behavior patterns than those ofthe withdrawal-vehicle group.

Ten hours after withdrawal, the patterns of the induced behaviors and the suppressive effects of the extract werealmost the same as those observed at 6 h (Fig. 2B).

On the contrary, MS aqueous extract had no significant effect on the spontaneous motor activity, compared to thevehicle control group (Fig. 3). The differences between the effects of different doses of the MS extract were also notsignificant. In the present preliminary study, we investigated its basic effects on the CNS. The results stronglyconfirmed that it has an antidepressant-like activity. The mechanisms that underlie this effect were not examined. Itcould work through serotonin or noradrenaline neurotransmissions as found for most of other antidepressant andneuroleptic drugs [12,13]. The aqueous extract of MS has a mild antidepressant-like activity. The aqueous extract at300 mg/kg exhibited a reducing effect on the ethanol withdrawal symptoms probably due to its antidepressant activity.This indicates that the young leaves of M. speciosa have properties that promise to be useful for clinical purposes.

Acknowledgements

We would like to thank Dr. Brian Hodgson for reviewing this manuscript. This work was financially supported by agrant from the Thailand Research Fund (code number: MRG46 80162).

References

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