title: Qualification of HSQC methods for quantitative composition of heparin and low molecular weight heparins.
abstract: An NMR HSQC method has recently been proposed for the quantitative determination of the mono- and disaccharide subunits of heparin and low molecular weight heparins (LMWH). The focus of the current study was the validation of this procedure to make the 2D-NMR method suitable for pharmaceutical quality control applications. Pre-validation work investigated the effects of several experimental parameters to assess robustness and to optimize critical factors. Important experimental parameters were pulse sequence selection, equilibration interval between pulse trains and temperature. These observations were needed so that the NMR method was sufficiently understood to enable continuous improvement. A standard validation study on heparin then examined linearity, repeatability, intermediate precision and limits of detection and quantitation; selected validation parameters were also determined for LMWH.
authors.affiliation: Institute for Chemical and Biochemical Research G. Ronzoni, via G. Colombo 81, 20133 Milan, Italy.
authors.forename: Lucio
authors.initials: L
authors.lastname: Mauri
authors.affiliation: Institute for Chemical and Biochemical Research G. Ronzoni, via G. Colombo 81, 20133 Milan, Italy.
authors.forename: Giovanni
authors.initials: G
authors.lastname: Boccardi
authors.affiliation: Institute for Chemical and Biochemical Research G. Ronzoni, via G. Colombo 81, 20133 Milan, Italy.
authors.forename: Giangiacomo
authors.initials: G
authors.lastname: Torri
authors.affiliation: Division of Pharmaceutical Analysis, Office of Testing and Research, Center for Drug Evaluation and Research, U.S. Food and Drug Administration, 645 S. Newstead Ave., St. Louis, MO 63110, USA.
authors.forename: Michael
authors.initials: M
authors.lastname: Karfunkle
authors.affiliation: Institute for Chemical and Biochemical Research G. Ronzoni, via G. Colombo 81, 20133 Milan, Italy.
authors.forename: Eleonora
authors.initials: E
authors.lastname: Macchi
authors.affiliation: Institute for Chemical and Biochemical Research G. Ronzoni, via G. Colombo 81, 20133 Milan, Italy.
authors.forename: Laura
authors.initials: L
authors.lastname: Muzi
authors.affiliation: Division of Pharmaceutical Analysis, Office of Testing and Research, Center for Drug Evaluation and Research, U.S. Food and Drug Administration, 645 S. Newstead Ave., St. Louis, MO 63110, USA. Electronic address: David.Keire@fda.hhs.gov.
authors.forename: David
authors.initials: D
authors.lastname: Keire
authors.affiliation: Institute for Chemical and Biochemical Research G. Ronzoni, via G. Colombo 81, 20133 Milan, Italy. Electronic address: guerrini@ronzoni.it.
authors.forename: Marco
authors.initials: M
authors.lastname: Guerrini
mesh_terms: D004187:Disaccharides / Q000032:analysis*; D006493:Heparin / Q000032:analysis*; D006495:Heparin, Low-Molecular-Weight / Q000032:analysis*; D057230:Limit of Detection; D015394:Molecular Structure; D008970:Molecular Weight; D009005:Monosaccharides / Q000032:analysis*; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D015203:Reproducibility of Results
chemical_list: D004187:Disaccharides; D006495:Heparin, Low-Molecular-Weight; D009005:Monosaccharides; D006493:Heparin
keywords: Heparin; Heteronuclear Single Quantum Correlation (HSQC); Low molecular weight heparins; Method qualification; Quantitative NMR; Robustness