Development and validation of a high-performance liquid chromatography method for standardization of the bioactive ethyl acetate fraction of Alstonia scholaris (Linn.) R. Br. growing in Egypt.
Bio-guided fractionation of the ethanolic extract of the leaves of Alstonia scholaris (Apocynaceae) growing in Egypt was carried out to evaluate its antihyperglycemic activity in alloxan-induced diabetic rats and its hepatoprotective activity against CCl4-induced hepatotoxicity in rats. The ethyl acetate fraction of the ethanolic extract showed the highest antihyperglycemic [(133.6 +/- 4.2) mg/mL, relative to metformin with (92.3 +/- 2.7) mg/mL] and hepatoprotective [(37.9 +/- 1.4) U/L, relative to silymarin with (29.7 +/- 0.8) U/L] activities. Four compounds were isolated from this fraction, and identified by spectroscopic techniques and by comparison with reported data: caffeic acid and isoquercitrin for the first time from this plant, in addition to quercetin 3-O-beta-D-xylopyranosyl (1''' --> 2")-beta-D-galactopyranoside (major compound) and chlorogenic acid. A validated reversed phase-high-performance liquid chromatography (RP-HPLC) method was developed for the standardization of the bioactive ethyl acetate fraction. The calibration curve showed good linearity (r2 > 0.999) within tested ranges. The relative standard deviation of the method was less than 3% for intra- (0.4-2.0%) and inter-day (1.9-2.8%) assays. Mean recovery of the method was within the range of 98.5-102.5%. The minimum detectable concentration of the analyte (LOD) was found to be 0.04 microg/mL. This developed HPLC method was shown to be simple, rapid, precise, reproducible, robust, specific, and accurate for quality assessment of the bioactive fraction.
Department of Pharmacognosy, Faculty of Pharmacy, Cairo University, Cairo 11562, Egypt. helaskary@hotmail.com
Hesham I
HI
El-Askary
Department of Pharmaceutical Biology, Faculty of Pharmacy & Biotechnology, German University in Cairo, Cairo 11835, Egypt.
Mahmoud M
MM
El-Olemy
Department of Pharmacognosy, Faculty of Pharmacy, Cairo University, Cairo 11562, Egypt.
Maha M
MM
Salama
Department of Pharmaceutical Biology, Faculty of Pharmacy & Biotechnology, German University in Cairo, Cairo 11835, Egypt.
Mahetab H
MH
Amer
D000085:Acetates / Q000032:analysis*; D031051:Alstonia / Q000737:chemistry*; D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004534:Egypt; D005260:Female; D009682:Magnetic Resonance Spectroscopy; D051381:Rats; D017207:Rats, Sprague-Dawley
D000085:Acetates; C007650:ethyl acetate