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Increased Clinical Sensitivity and Specificity of Plasma Protein N-Glycan Profiling for Diagnosing Congenital Disorders of Glycosylation by Use of Flow Injection-Electrospray Ionization-Quadrupole Time-of-Flight Mass Spectrometry.

ABSTRACT: BACKGROUND:Congenital disorders of glycosylation (CDG) represent 1 of the largest groups of metabolic disorders with >130 subtypes identified to date. The majority of CDG subtypes are disorders of N-linked glycosylation, in which carbohydrate residues, namely, N-glycans, are posttranslationally linked to asparagine molecules in peptides. To improve the diagnostic capability for CDG, we developed and validated a plasma N-glycan assay using flow injection-electrospray ionization-quadrupole time-of-flight mass spectrometry. METHODS:After PNGase F digestion of plasma glycoproteins, N-glycans were linked to a quinolone using a transient amine group at the reducing end, isolated by a hydrophilic interaction chromatography column, and then identified by accurate mass and quantified using a stable isotope-labeled glycopeptide as the internal standard. RESULTS:This assay differed from other N-glycan profiling methods because it was free of any contamination from circulating free glycans and was semiquantitative. The low end of the detection range tested was at 63 nmol/L for disialo-biantennary N-glycan. The majority of N-glycans in normal plasma had <1% abundance. Abnormal N-glycan profiles from 19 patients with known diagnoses of 11 different CDG subtypes were generated, some of which had previously been reported to have normal N-linked protein glycosylation by carbohydrate-deficient transferrin analysis. CONCLUSIONS:The clinical specificity and sensitivity of N-glycan analysis was much improved with this method. Additional CDGs can be diagnosed that would be missed by carbohydrate-deficient transferrin analysis. The assay provides novel biomarkers with diagnostic and potentially therapeutic significance.

SUBMITTER: Chen J

PROVIDER: S-EPMC6886244 | biostudies-literature | 2019 May

REPOSITORIES: biostudies-literature

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Increased Clinical Sensitivity and Specificity of Plasma Protein <i>N</i>-Glycan Profiling for Diagnosing Congenital Disorders of Glycosylation by Use of Flow Injection-Electrospray Ionization-Quadrupole Time-of-Flight Mass Spectrometry.

Chen Jie J Li Xueli X Edmondson Andrew A Meyers Gail Ditewig GD Izumi Kosuke K Ackermann Amanda M AM Morava Eva E Ficicioglu Can C Bennett Michael J MJ He Miao M

Clinical chemistry 20190215 5

<h4>Background</h4>Congenital disorders of glycosylation (CDG) represent 1 of the largest groups of metabolic disorders with >130 subtypes identified to date. The majority of CDG subtypes are disorders of <i>N</i>-linked glycosylation, in which carbohydrate residues, namely, <i>N</i>-glycans, are posttranslationally linked to asparagine molecules in peptides. To improve the diagnostic capability for CDG, we developed and validated a plasma <i>N</i>-glycan assay using flow injection-electrospray ...[more]

PMID: 30770376

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Project description:IntroductionWheat (Triticum aestivum) it is one of the most important staple food crops worldwide and represents an important resource for human nutrition. Besides starch, proteins and micronutrients wheat grains accumulate a highly diverse set of phytochemicals.ObjectivesThis work aimed at the development and validation of an analytical workflow for comprehensive profiling of semi-polar phytochemicals in whole wheat grains.MethodReversed-phase ultra-high performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight mass spectrometry (UHPLC/ESI-QTOFMS) was used as analytical platform. For annotation of metabolites accurate mass collision-induced dissociation mass spectra were acquired and interpreted in conjunction with literature data, database queries and analyses of reference compounds.ResultsBased on reversed-phase UHPLC/ESI-QTOFMS an analytical workflow for comprehensive profiling of semi-polar phytochemicals in whole wheat grains was developed. For method development the extraction procedure and the chromatographic separation were optimized. Using whole grains of eight wheat cultivars a total of 248 metabolites were annotated and characterized by chromatographic and tandem mass spectral data. Annotated metabolites comprise hydroquinones, hydroxycinnamic acid amides, flavonoids, benzoxazinoids, lignans and other phenolics as well as numerous primary metabolites such as nucleosides, amino acids and derivatives, organic acids, saccharides and B vitamin derivatives. For method validation, recovery rates and matrix effects were determined for ten exogenous model compounds. Repeatability and linearity were assessed for 39 representative endogenous metabolites. In addition, the accuracy of relative quantification was evaluated for six exogenous model compounds.ConclusionsIn conjunction with non-targeted and targeted data analysis strategies the developed analytical workflow was successfully applied to discern differences in the profiles of semi-polar phytochemicals accumulating in whole grains of eight wheat cultivars.

Dataset Information

Increased Clinical Sensitivity and Specificity of Plasma Protein N-Glycan Profiling for Diagnosing Congenital Disorders of Glycosylation by Use of Flow Injection-Electrospray Ionization-Quadrupole Time-of-Flight Mass Spectrometry.

Publications

Increased Clinical Sensitivity and Specificity of Plasma Protein <i>N</i>-Glycan Profiling for Diagnosing Congenital Disorders of Glycosylation by Use of Flow Injection-Electrospray Ionization-Quadrupole Time-of-Flight Mass Spectrometry.

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