Qualitative and Quantitative determinations of active principles having positive effect on mamory deficit in Longan seed extract

Abstract

Several studies of chemicals in longan (Dimocarpus longan Lour.) seed have been suggested to have positive effect on deficit in learning and memory. In this study, the extraction methods from longan seeds found that ethyl acetate extracts showed the positive effect on learning and memory impairment after bilateral carotid arteries (2-VO) occlusion in mice. Using the fractionation of the ethyl acetate part by conventional chromatographic technique together with the screening activities, we obtained four major compounds; quercetin, gallic acid, propyl gallate and rutin. Rutin showed to be a one of the most active principle compounds at dose 100mg/kg and used to be a chemical marker for the standardized herbal extract. All the compounds were identified by physical and spectroscopic properties. Furthermore, we prepared relatively stable standardized herbal extract from longan seeds by removing the problematic phenolic portion. High performance liquid chromatography technique (HPLC technique) for determination of rutin in longan seed extract was developed and validated. This method was validated according to ICH guideline. The rutin in longan seed extract was determined using reverse phase column as stationary phase, an isocratic as mobile phase and hesperidin as internal standard. The method shows linearity range of 0.31-1.54µg/ml of rutin (r² = 0.9940).The percentage recovery (%R) was in the range of 99.30-101.98 and the relative standard determination (%RSD) was not more than 2. The limit of detection and limit of quantification of rutin were 0.13 and 0.38 µg/ml, respectively. A thin-layer chromatographic (TLC) method was developed to determine rutin by using silica gel plate GF[254] as stationary phase and butanol: glacial acetic acid (9:3) as developing solvent, hesperidin as internal standard. The detection of the TLC spot was developed by spraying with anthrone in ethanol. This method was validated according to ICH guideline. The method shows linearity range of 0.82-2.83mg/ml of rutin (r²= 0.9950) .The percentage recovery was in the range of and percentage of 98.41-100.46 .RSD was not more than 2. The limit of detection and limit of quantification of rutin were 0.21 and 0.62mg/ml, respectively.Proper validation of two analytical methods and according to ICH guidelines.