title: Electrophoretic mobility as a tool to separate immune adjuvant saponins from Quillaja saponaria Molina.
abstract: Quillaja saponins are used as adjuvants in animal vaccines but their application in human vaccination is still under investigation. Isolation and characterization of adjuvant saponins is very tedious. Furthermore, standardization of Quillaja saponins is critical pertaining to its application in humans. In this study, a convenient method based on agarose gel electrophoresis was developed for the separation of Quillaja saponins. Six different commercial Quillaja saponins were segregated by size/charge into numerous fractions. Each of the fractions was characterized by ESI-TOF-MS spectroscopy and thin layer chromatography. Real-time impedance-based monitoring and red blood cell lysis assay were used to evaluate cytotoxicity and hemolytic activities respectively. Two specific regions in the agarose gel (delimited by specific relative electrophoretic mobility values) were identified and characterized by exclusive migration of acylated saponins known to possess immune adjuvant properties (0.18-0.58), and cytotoxic and hemolytic saponins (0.18-0.94). In vivo experiments in mice with the isolated fractions for evaluation of adjuvant activity also correlated with the relative electrophoretic mobility. In addition to the separation of specific Quillaja saponins with adjuvant effects as a pre-purification step to HPLC, agarose gel electrophoresis stands out as a new method for rapid screening, separation and quality control of saponins.
authors.affiliation: Institut für Laboratoriumsmedizin, Klinische Chemie und Pathobiochemie, Charité-Universitätsmedizin Berlin, Campus Virchow-Klinikum, Augustenburger Platz 1, D-13353 Berlin, Germany.
authors.forename: Roger
authors.initials: R
authors.lastname: Gilabert-Oriol
authors.affiliation: Institut für Laboratoriumsmedizin, Klinische Chemie und Pathobiochemie, Charité-Universitätsmedizin Berlin, Campus Virchow-Klinikum, Augustenburger Platz 1, D-13353 Berlin, Germany.
authors.forename: Alexander
authors.initials: A
authors.lastname: Weng
authors.affiliation: Institut für Laboratoriumsmedizin, Klinische Chemie und Pathobiochemie, Charité-Universitätsmedizin Berlin, Campus Virchow-Klinikum, Augustenburger Platz 1, D-13353 Berlin, Germany.
authors.forename: Benedicta
authors.initials: B
authors.lastname: von Mallinckrodt
authors.affiliation: Institut für Laboratoriumsmedizin, Klinische Chemie und Pathobiochemie, Charité-Universitätsmedizin Berlin, Campus Virchow-Klinikum, Augustenburger Platz 1, D-13353 Berlin, Germany.
authors.forename: Anja
authors.initials: A
authors.lastname: Stöshel
authors.affiliation: Institut für Laboratoriumsmedizin, Klinische Chemie und Pathobiochemie, Charité-Universitätsmedizin Berlin, Campus Virchow-Klinikum, Augustenburger Platz 1, D-13353 Berlin, Germany.
authors.forename: Linda
authors.initials: L
authors.lastname: Nissi
authors.affiliation: Institut für Pharmazie-Pharmazeutische Biologie, Freie Universität Berlin, Königin-Luise-Straße 2+4, D-14195 Berlin, Germany.
authors.forename: Matthias F
authors.initials: MF
authors.lastname: Melzig
authors.affiliation: Institut für Laboratoriumsmedizin, Klinische Chemie und Pathobiochemie, Charité-Universitätsmedizin Berlin, Campus Virchow-Klinikum, Augustenburger Platz 1, D-13353 Berlin, Germany.
authors.forename: Hendrik
authors.initials: H
authors.lastname: Fuchs
authors.affiliation: Institut für Laboratoriumsmedizin, Klinische Chemie und Pathobiochemie, Charité-Universitätsmedizin Berlin, Campus Virchow-Klinikum, Augustenburger Platz 1, D-13353 Berlin, Germany. Electronic address: mayank.thakur@charite.de.
authors.forename: Mayank
authors.initials: M
authors.lastname: Thakur
mesh_terms: D000276:Adjuvants, Immunologic / Q000737:chemistry / Q000302:isolation & purification*; D000818:Animals; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival; D024202:Electrophoretic Mobility Shift Assay; D006461:Hemolysis / Q000187:drug effects; D006801:Humans; D066298:In Vitro Techniques; D051379:Mice; D008807:Mice, Inbred BALB C; D031990:Quillaja / Q000737:chemistry; D062605:Quillaja Saponins / Q000737:chemistry / Q000302:isolation & purification*; D013154:Spleen / Q000166:cytology / Q000187:drug effects
chemical_list: D000276:Adjuvants, Immunologic; D062605:Quillaja Saponins
keywords: Cytotoxicity; Electrophoresis; Hemolysis; Immune adjuvant; Quillaja saponins