Physicochemical Characterization of Low Molecular Weight Heparin

Atha, Donald H.; Coxon, Bruce; Reipa, Vytas; Gaigalas, Adolfas K.

doi:10.1002/jps.2600840318

Cited by 15 publications

(12 citation statements)

References 18 publications

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“…16 Although we have not used Raman spectoscopy to examine modified heparins with varying levels of sulfation, we have examined low molecular weight heparins prepared by alternative methods. 6 In this previous study we found distinct differences in the proportion of N-sulfate in two preparations of low molecular weight heparin which were consistent with their different methods of preparation. Previous studies have assigned sulfate resonant frequencies consistent with these measurements and have shown for a number of heparins and N-desulfated heparin that the signal amplitude is a linear function of the percentage content of sulfate groups.…”

Section: Resultssupporting

confidence: 76%

“…Although the intensities of the N-and O-sulfate peaks differ due to a strong influence by environmental factors, their relative intensities are constant and can be used to determine the relative proportions of the N-and O-sulfates in equivalent samples. 6 We have accurately determined the position and proportion of these peaks by deconvolution as shown in Figure 1B. The low intensity peak at about 1000 cm -1 in heparin is in a region of the spectrum attributed to the C-N stretch.…”

Section: Resultsmentioning

confidence: 99%

“…[4][5][6] Raman spectroscopy, which is complementary to NMR, can provide additional structural information such as the proportions of N-sulfated and O-sulfated residues. [6][7][8] In this regard, we wanted to determine whether Raman spectroscopy would be sufficiently sensitive to discriminate between the N-and O-sulfated residues at different ring positions in the heparin disaccharide. Sensitivity to such structural features would make Raman spectroscopy particularly useful in the characterization of various heparin preparations.…”

Section: Introductionmentioning

confidence: 99%

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Structural Analysis of Heparin by Raman Spectroscopy

Atha

Gaigalas

Reipa

1996

Journal of Pharmaceutical Sciences

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The Raman spectra of commercially available heparin disaccharide standards exhibit bands associated with the N-sulfate and the 6-O-sulfate groups of the glucosamine and the 2-O-sulfate of the iduronic acid. The N-sulfate has a strong band at 1039 cm-1. The 6-O-sulfate and the 2-O-sulfate exhibit bands at 1055 and 1065 cm-1, respectively. The pattern of these modes, which are assigned to the symmetric SO3 vibrations, was supported by semiempirical quantum mechanical calculations. The above bands were identified in the Raman spectrum of a commercial preparation of porcine mucosal heparin and were used to determine the relative proportion of the N-sulfate, 6-O-sulfate, and 2-O-sulfate groups in the heparin molecule. This information, which is complementary to that obtained by NMR spectroscopy, is of particular importance in relation to biological activity. This study also extends the usefulness of Raman spectroscopy to include structural details required for the quality assurance of pharmaceutical preparations of heparin.

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Section: Resultssupporting

confidence: 76%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Structural Analysis of Heparin by Raman Spectroscopy

Atha

Gaigalas

Reipa

1996

Journal of Pharmaceutical Sciences

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“…Due to its high sensitivity to even minor structural variations, 1 H NMR spectroscopy has been used previously to detect variations in the chemical composition of heparin [25, 55, 56], low molecular weight heparins [57], heparin-derived oligosaccharides [58, 59] and as a screening tool for GAG impurities [25, 60]. As 1 H NMR has also been recommended by the FDA as one of the analytical techniques for rapidly screening OSCS, intense efforts are being made to improve the NMR-based characterization of heparin API.…”

Section: Nmr Spectroscopymentioning

confidence: 99%

Analysis and characterization of heparin impurities

2010

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This review discusses recent developments in analytical methods available for the sensitive separation, detection and structural characterization of heparin contaminants. The adulteration of raw heparin with oversulfated chondroitin sulfate (OSCS) in 2007–2008 spawned a global crisis resulting in extensive revisions to the pharmacopeia monographs on heparin and prompting the FDA to recommend the development of additional physicochemical methods for the analysis of heparin purity. The analytical chemistry community quickly responded to this challenge, developing a wide variety of innovative approaches, several of which are reported in this special issue. This review provides an overview of methods of heparin isolation and digestion, discusses known heparin contaminants, including OSCS, and summarizes recent publications on heparin impurity analysis using sensors, near-IR, Raman, and NMR spectroscopy, as well as electrophoretic and chromatographic separations.FigureSchematic illustrating the process for heparin impurity characterization

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“…Therefore, to date, MS has been used for the characterization of short oligosaccharides fragments released by chemical or enzymatic depolymerization previously isolated by chromatographic or electrophoretic steps [20,21,22], and no information regarding the origin of the products was extracted. 1 H nuclear magnetic resonance (NMR) spectroscopy has become a successful technique for characterizing the chemical composition of intact heparin [1,23,24,25] and heparin-derived oligosaccharides [26,27], and for checking the impurities from other GAGs [23,24,28]. In other instances, NMR spectroscopy is being used in food research in the authentication of the quality of products such as wines, fruit juices and jams [29,30,31,32].…”

Section: Introductionmentioning

confidence: 99%

Estimation of the composition of heparin mixtures from various origins using proton nuclear magnetic resonance and multivariate calibration methods

et al. 2002

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A multivariate calibration method for the characterization of heparin samples based on the analysis of (1)H nuclear magnetic resonance (NMR) spectral data is proposed. Heparin samples under study consisted of two-component or four-component mixtures of heparins from porcine, ovine and bovine mucosae and bovine lung. Although the (1)H NMR spectra of all heparin types were highly overlapping, each origin showed some particular features that could be advantageously used for the quantification of the components. These features mainly concerned the anomeric H, which appeared in the range 5.0-5.7 ppm and the peaks of acetamidomethyl protons at 2.0-2.1 ppm. The determination of the percentage of each heparin class depended on these differences and was carried out using partial least squares regression (PLS) as a calibration method. Prior to the PLS analysis, the spectral data were standardized using the internal standard peak (sodium 4,4-dimethyl-4-silapentanoate- 2,2,3,3- d (4), TSP) as the reference. The quantification of each heparin type in the samples using PLS models built with 4 or 5 components was satisfactory, with an overall prediction error ranging from 3% to 10%.

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Physicochemical Characterization of Low Molecular Weight Heparin

Cited by 15 publications

References 18 publications

Structural Analysis of Heparin by Raman Spectroscopy

Structural Analysis of Heparin by Raman Spectroscopy

Analysis and characterization of heparin impurities

Estimation of the composition of heparin mixtures from various origins using proton nuclear magnetic resonance and multivariate calibration methods

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