Phytoohemioal and Biological Studies on Solanum dubium seeds and Sunnut honey Combination as Potential Anti-asthmatic

Abstract

ABSTRACT Asthma is an allergic disease resulting in inflammation of the airways leading to bronchoconstriction and wheezing. Anti asthmatic agents are of substantial adverse effect, the present study attempted to evaluate scientifically a herbal remedy composed of Sunnut honey and ground seeds of Solanum dubium which is used in Sudanese traditional medicine for control of bronchial asthma _ The effect of the crude extract of combination, seeds and honey of S. dubium known in Sudan as Gubbain on carageenan-induced acute inflammation was evaluated by the rat hind paw oedema method. The result showed a significant inhibitory effect on the carrageenan-induced oedema in rats at all the doses tested in rats in all groups, when compared to the normal saline control and standard Diclofenac Sodium. The activity resides more at the dose of 4ml/kg of combination with 91.16% inhibition and then the percentage (%) inhibition of paw edema in 2ml/kg of combination, Zml/kg of S. dubium seeds and 2m]/kg honey, with 81.05, 72.07 and 66.75% respectively after 4 hours of extract administration, when compared with standard drug 74. 34%. The isolated guinea pigs tracheal strip a pattern of allergens were used to investigate the bronchodilating effect of crude extracts of combination, S. dubium seeds and honey, S. dubium seeds and Surmut honey. Changes in the reactivity of the tracheal smooth muscle are measured as the reaction on a single dose of bronchoconstrictor mediator histamine. The results showed, both seed aqueous extract and the combination relaxed effectively the guinea pigs tracheal strip isolated and their relaxant effect was shown to be Br Adrenoceptor stimulates like action since it was completely blocked by Propranolol and antagonized the contracting effect of histamine on the guinea pigs tracheal strip. Acute and Sub-chronic toxicity studies of the crude extracts of the combination, honey and the seeds on rats, the LD50 percentage was detennined. The toxicity study revealed that there is no acute or sub-chronic toxicity in different concentrations of the extracts were found to be safe up to 4000mg/kg body weight. Phytochemical studies revealed variable concentrations of alkaloids, sterols, triterpene, flavonoids, coumarins, saponins and tannins. The aqueous extract, 96% ethanolic extract and methanolic extract of the three samples, were tested against four standard bacteria: Bacillus subtilis; Staphylococcus aureus, Escherichia coli; Salmonella typhi, and two standard fungi: Aspergillus niger and Candida albicans, using the agar plate diffusion method. All groups tested exerted

significant antimicrobial activity of concentration of 100mg/ml ranging from 20 to 35mm zone of inhibition and intermediate to significant antifungal activity ranging from 14 to 25mm zone of inhibition. The prepared extracts were also tested for their free radical scavenging properties using 2, 2- diphyenyl-l- picrylhydrazyl (DPPH) and propyl gallate assays. The aqueous extract of the seeds and honey combination showed the highest radical scavenging activity (RSA) 91i0.04%, according to the DPPH assay and the control propyl gallate gave RSA 93i0.01%. Attempts to separate active compounds from the methanolic extract of S. dubium seeds was carried out using bioassay-guided coltunn chromatography. Structure of the isolated compormds of the methanolic extract of S. dubium seeds was achieved by different spectroscopic methods: nuclear magnetic resonance (NMR), mass spectrometry (MS), ultraviolet (UV) and infrared (IR) spectrometry. The spectroscopic analysis of fraction 10 indicated the presence of phenolic and steroidal alkaloid compounds, identified by ms- spectrometry as mixtures of Gallic and ellagic acids and the steroidal alkaloid solasodine. A simple and rapid high-performance thin-layer chromatographic method (HPTLC) was developed and validated for quantitative estimation of gallic acid, ellagic acid and solasodine from the seed extracts, using a CAMAG Linomat V and were controlled by WinCATS software CAMAG. The seeds were found to contain 1.1% w/W of ellagic acid, 2.1% w/w of gallic acid and 1.1% of solasodine. The method was validated for precision, recovery and repeatability as per the International Conference on Harmonization guidelines for gallic acid, ellagic acid and solasodine. Statistical analysis of data showed that, the method was precise, accurate, reproducible and selective for the analysis of gallic acid, ellagic acid and solasodine, and sensitive HPTLC procedure, that could be successfully applied for the estimation of solasodine, ellagic acid and gallic acid in Solanaceous plants and solasodine, ellagic acid and gallic acid production and can be usefill for quality control of herbal extracts or herbal drugs prepared from S. dubium. Isolation of gallic acid, ellagic acid and solasodine from enriched fractions using preparative HPTLC, of the dried methanolic extract was carried out on the CAMAG HPTLC System and characterization was achieved by different spectroscopic methods: nuclear magnetic resonance (NMR), mass spectrometry (MS), ultraviolet (UV) and infrared (IR) spectrometry.