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Identifier 000355526
Title Απομόνωση και χαρακτηρισμός πρωτεΐνης από τον φυτικό οργανισμό Pancratium Maritimum
Alternative Title Isolation and identification protein from plant organism pancratium maritimum
Author Παπαδάκη, Αριάδνη Ανδρέα
Thesis advisor Γανωτάκης, Δημήτριος
Abstract The aim of this work was to study the plant Pancratium Maritimum, which belongs in the family Amaryllidaceae, is a bulbous, perennial, uncultivated plant that lives and completes its biological cycle in the coasts of Mediterranean and Iberian Peninsula. Initially the isolation of proteins from the bulbs of plant was performed with use of cation exchange chromatography S-Sepharose, with this procedure the proteins were separated from the mix of proteins. Two of these proteins that were isolated are of interest to us, of which the molecular weight is approximately 12kDa which was calculated based on proteins of known molecular weight after their electrophoresis in polyacrylamide gel (SDS-PAGE). Followed by precipitation of proteins with various ways, such as using trichloroacetic acid (TCA) and organic solvent acetone. Then, for the identification of these proteins, mass spectroscopy was applied, with matrix assisted laser desorption ionization (MALDI) and time of flight masses analyser (TOF), at which point with the help of bioinformatics certain sequence of peptides were recognized, from homologous proteins of plant organisms, as Vitis vinifera, Oryza sativa, Zea mays. Most homologous proteins that resulted from MS/MS spectra of the main tops are hypothetical, nevertheless we know certain operations they perform in the cell, such as participation in metabolic processes, binding proteins of other macromolecules or ions. Also the plant Pancratium Maritimum as all plants of family Amaryllidaceae is considered a pharmaceutical plant, because of an abundance of properties, which are owed to their content of the organic chemical compounds alkaloids. Alkaloids in small quantities have anticancer, antivirus, antibiotic, analgetic and anticholinesterase activities as well as phytohormonic attributes. However in large quantities they are considered toxic or even poisonous for the human organism. Thus isolation of the alkaloid compounds from the bulbs of plant was performed, in methanol extract. Then according to Molisch test it was realized that the methanol extract contains glycosides alkaloids and possibly free sugars. With the use of thin layer chromatography (TLC) separation of the mixture was performed, and with comparison to the bibliographic data we acquired a first picture for the constitution of mixture. Then followed separation of mix with the use of reversed phase high performance liquid chromatography (PR-HPLC). In the main two fractions that were received from HPLC, spectroscopy MS was performed, with electron spray ionization (ESI) and as a masses analyser, a trap of ions. From the MS spectra we realized that the first fraction contained a diglucoside of a substituted molecule of alkaloid Tazettine, of hydroxyl-O-methylotazettine and in the second fraction, a diglucoside of a substituted molecule of alkaloid Lycorine, of acetyllycorine. Our next objective was to check cytotoxic and at extension anticancer action of the methanolic solution that contains a mix of alkaloid, as well as the two alkaloids that were received after separation using of HPLC
Language Greek
Subject Amarllidaceae
pancratium maritimum
Φασματοσκοπία μάζας
Issue date 2009-11-27
Collection   Faculty/Department--Faculty of Sciences and Engineering--Department of Chemistry--Post-graduate theses
  Type of Work--Post-graduate theses
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