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Unformatted text preview: édéric Faizal Weber, Sadia Sultan, Kalavathy Ramasamy and Anthony L.J. Cole Institute for the Study of Natural Remedies (iKUS) and Faculty of Ph armacy, Universiti Teknologi MARA, Kampus Puncak Alam, 42300 Kuala Selangor, Selangor, Malaysia 2 School of Biological Sciences, University of Canterbury, Christchurch, New Zealand E-mail: [email protected] 1 1 1 2 1 A rapid and robust chromatography method to generate chromatographic profile of extracts plays an important role in the discovery of new chemical entities from natural sources. Fractionation and isolation of fungal metabolites was done by high performance liquid chromatography (HPLC). A qualitative HPLC method was developed for multiple-analyte assays and applied for the analysis of some 150 of different fungal extracts. The chromatographic system includes an HPLC Agilent 1200 series system equipped with a diode-array detector (DAD) and a fraction collector fitted with up to 4 deep-well microtitre plates. All analyses and separations are carried out in a reverse phase mode, using a Synergy ® 4u Hydro-RP 8 0Å column (150 × 4.6 mm, 4 µm particle size, Phenomenex , USA) with a guard column filled with the same material. The column temperature is maintained at 36 °C. The DAD collects the full UV spectrum (190 to 600 nm) and is set to display the absorbance at the following wavelengths: 220, 254, 280 and 360 nm. The mobile phase consists of purified water (s olvent A) and acetonitrile (solvent B). The separations carried out at a flow rate of 1 mL/min with the following elution gradient: 0 min 10 % B, 10 min 46 % B, 14 min 70 % B, and 20 min 100 % B. Aliquots of 10 µL of each extract solution (10 mg/mL) are injected in order to record their chromatographic profiles. Then, 30 µL of aliquot were injected two times consecutively for preparative purpose. Fractions collected every 0.5 minutes for 25 minutes into the 2 mL 96-deep well plates (polypropylene) and then dried with help of an EZ-2 ® centrifugal evaporation system (Genevac , UK). The collected dried fractions are then subjected to antimicrobial assays. This non-line...
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This note was uploaded on 02/17/2011 for the course CHEMISTRY 101 taught by Professor Csr during the Spring '11 term at University of Louisville.
- Spring '11