Pulmonary, Gastrointestinal and Urogenital Pharmacology
β2-Adrenergic activity of 6-methoxykaempferol-3-O-glucoside on rat uterus: In vitro and in silico studies

https://doi.org/10.1016/j.ejphar.2011.05.066Get rights and content

Abstract

6-Methoxykaempferol-3-O-glucoside (6-MKG) was isolated from a Sudanese herb (El-hazha). The pharmacological effects of 6-MKG were tested on isolated non-pregnant or late-pregnant rat uteri in vitro, whilst docking studies were carried out modelling of the binding of 6-MKG to the rat β2-adrenoceptor in silico. In vitro studies revealed that 6-MKG was able to relax both the non-pregnant and the late-pregnant uterine contractility with 50% of the Emax of terbutaline, whilst the EC50 for 6-MKG was at least half than that of terbutaline. The β2-adrenoceptors antagonist 3-(isopropylamino)-1-[(7-methyl-4-indanyl)oxy]butan-2-ol(ICI118,551) competitively antagonised the relaxing effect of 6-MKG. Radioligand binding and cAMP studies confirmed the β2-adrenoceptors agonistic property of the compound. In in silico docking studies, 6-MKG bound to rat β2-adrenoceptors with low ∆Gbind value (− 11.53 ± 0.06 kcal/mol) and it interacted with four residues of the active site (Asp113, Asn312, Cys191and Tyr316). It is concluded that 6-MKG exerts weak β2-adrenoceptor agonistic activity and can be considered a natural compound with potential therapeutic significance in the field of premature pregnant uterine contractions and asthmatic problems.

Research highlights

► 6-methoxykaempferol-3-O-glucoside (6-MKG) was isolated and identified early (2004). ► Its pharmacological profile remains controversial. ► This study aimed to proof and determine the pharmacological profile of 6-MKG by in silico - in vitro techniques using β2-adrenoceptor as main target. ► The present study revealed that 6-MKG possesses weak β2-adrenoceptor agonistic activity. ► 6-MKG was with therapeutic potential for premature labour inhibition and asthma treatment.

Introduction

El-hazha (Haplophyllum tuberculatum) (Forssk.) A. Juss. (Rutaceae) is an herb indigenous to the northern part of Sudan, North Africa and other areas of the Middle East (Boulus, 1983). Named locally in Sudan “a plant of all disease”, it is to be found in every Sudanese home as an emergency drug and is used extensively by the old Sudanese in the rural areas. The herb is utilised in Sudan as an antispasmodic, an antiflatulant, to relieve toothache, and to treat allergic rhinitis (Mohamed et al., 1996), malaria, gynaecological disorders, asthma, inspiration difficulties, renal disorders and others.

This plant is well-known amongst herbalists and is widely used traditionally in other countries, such as Saudi Arabia (Al-Yahya et al., 1991) and Oman, to treat skin discolouration, infections and parasitic diseases (Mossa et al., 1987). Due to its importance, new species were reported recently from Spain (Navarro et al., 2004).

Our previous investigation results revealed that the Methanolic-extract of the plant had significant relaxant activity (Ahmed et al., 2010). This finding stimulated the isolation and pharmacological characterization of its active compounds.

β2-Adrenoceptor agonists are of therapeutic potential due to their use for asthma (D'Urzo et al., 2010) and to inhibit pre-term labour (Giles and Bisits, 2007), which is still medical challenge (Clouse et al., 2007). A few β2-adrenoceptor drugs are of a natural origin. This encouraged us to attempt to isolate and test the pharmacological features of isolated compounds on the β2-adrenergic system.

Although the literature revealed the isolation and identification of 6-methoxykaempferol-3-O-glucoside (6-MKG) (Wei et al., 2004), its pharmacological profile remains undetermined.

In this study we set out to determine the pharmacological profile of 6-MKG on the basis of its uterus-relaxing activity, using β2-adrenoceptors as main target.

Section snippets

Plant material

The aerial parts of H. tuberculatum (El-hazha) were collected in the north of Sudan (Abu-hamad, Nahr El-Neel State) in November 2008 and identified by Dr. Wai'l E. abdalla and Yahia S. Mohamed from MAPRI (Medicinal and Aromatic Plants Research Institute, Khartoum, Sudan). A voucher specimen (No. M23/08) has been deposited at the Herbarium of MAPRI.

Isolation and identification of 6-MKG

6-MKG, 3,4′,5,7-tetrahydroxyflavone-3-O-β-d-glucopyranoside, was isolated from the dried aerial parts of El-hazha, and its structure was determined

Isolated rat uterus

6-MKG and terbutaline were both found to induce a concentration-dependent relaxation on the isolated non-pregnant and the late-pregnant rat uterus pre-contracted with 25 mM KCl (Fig. 1).

On the non-pregnant uterus (Fig. 1A), the Emax of 6-MKG decreased and differed significantly from that of terbutaline, whereas the EC50 values did not differ significantly.

On the late-pregnant uterus (Fig. 1B), both Emax and EC50 of 6-MKG had low difference from the terbutaline values.

There were no signs of

Discussion

El-hazha is a plant that possesses uterus-relaxant activity (Ahmed et al., 2010). We therefore fractionated and isolated 6-MKG from it by bioassay-guided fractionation procedures (Hostettmann et al., 2000) and subjected it to post-isolation in vitro pharmacological investigations and in silico molecular modelling with docking studies.

6-MKG evidently has agonistic features, because it produced approximately 50% of the maximum activity of terbutaline on the isolated rat uterus, with a higher

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