α-Cyano-3-aminocinnamic acid: A novel reactive matrix for qualitative and quantitative analysis of plant N-glycans by MALDI-MS.

N-glycans have a diversity of crucial biological roles in organisms. Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) has become an indispensable analytical instrument for biomolecules. However, due to the inherent low abundance, high structural heterogeneity, and poor ionization efficiency of N-glycans, as well as the extremely inhomogeneous co-crystal property using traditional matrices, the qualitation and quantitation of N-glycans by MALDI-MS remains challenging. In the present study, α-cyano-3-aminocinnamic acid (3-CACA) was reasonably designed and synthesized as a novel reactive matrix for N-glycan analysis. Combining with traditional matrix α-cyano-4-hydroxycinnamic acid (CHCA) as an acidic catalyst, a combinational matrix 3-CACA/CHCA was obtained with homogeneous co-crystallization and high derivatization efficiency, achieving the sensitive qualitation with the limits of detection low to femtomole and reproducible quantitation with good linearity (R2 > 0.998). As a result, the established method was successfully applied to the on-target derivatization and high-throughput quantification of N-glycans in eight varieties of the peach complex system, indicating that N-glycan has the potential to become a new biomarker for food allergy, and elucidating the prospective correlation between N-glycan epitopes and allergic reactions.

2-Hydrazinoterephthalic Acid as a Novel Negative-Ion Matrix-Assisted Laser Desorption/Ionization Matrix for Qualitative and Quantitative Matrix-Assisted Laser Desorption/Ionization-Mass Spectrometry Analysis of N-Glycans in Peach Allergy Research.

Glycans recently attracted considerable attention as the proposal of cross-reactive carbohydrate determinants for food allergy. Matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-MS) is powerful in analyzing biomolecules, while its applications in glycans are still challenging. Herein, a novel reactive matrix-assisted laser desorption/ionization (MALDI) matrix, 2-hydrazinoterephthalic acid, was rationally designed and synthesized. It provides uniform co-crystallization with glycans and only produces deprotonated ions with high intensities in the negative-ion mode. In combination with sinapic acid, a rapid and high-throughput method was established for on-target analysis of glycans with a superior limit of detection at the femtomole level and a good linearity (R2 > 0.999). Furthermore, the established method was successfully applied to quantify N-glycans in different cultivars and tissues of peach [Prunus persica (L.) Batsch]. Our work suggests the potential role of N-glycans as biomarkers for food-borne allergy and lays a methodological foundation for the elucidation of the possible relationship between carbohydrate epitopes and food allergy.

Rapid quality control of medicine and food dual purpose plant polysaccharides by matrix assisted laser desorption/ionization mass spectrometry.

With their multiple biological activities and health benefit effects, polysaccharides from medicine and food dual purpose plants (MFDPPPs) have been extensively applied in many fields, including in medical treatments, stock farming, and cosmetics. However, to date, quality issues of MFDPPPs and technologies for the analysis of polysaccharides have posed challenges to chemists. Reported herein is a rapid and high-throughput quality control method for analyzing MFDPPPs, based on matrix assisted laser desorption/ionization mass spectrometry (MALDI-MS). For the analysis of illegally added and doped substances, ferroferric oxide nanoparticles were employed as the MALDI matrix to avoid small molecule interference. Qualitatively, high sensitivity was obtained for both illegal drugs and glucose. Quantitatively, the best linear response (R2 > 0.99) was attained in the concentration range from 0.005 to 1 mg mL-1 for glucose. For the analysis of polysaccharides, 2,5-dihydroxybenzoic acid/N-methylaniline was employed as the MALDI matrix to increase the detection sensitivity and mass range coverage. Furthermore, the established method was successfully applied to the analysis of supplements from Astragalus polysaccharides and Lentinan real samples, showing its potential in quality control for MFDPPPs.

Parallel On-Target Derivatization for Mass Calibration and Rapid Profiling of N-Glycans by MALDI-TOF MS.

Glycosylation is an important post-translational modification of proteins, and abnormal glycosylation is involved in a variety of diseases. Accurate and rapid profiling of N-glycans by matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) is still technically challenging and hampered mainly by mass drift of instrument, manual identification of spectrum peaks, and poor cocrystallization with traditional matrices besides low ionization efficiency of analytes. In the present study, a parallel on-target derivatization strategy (POTDS), on the basis of two rationally combined matrices, i.e., 3-hydrazinobenzoic acid plus DHB (DHB/3HBA) and quinoline-3-carbohydrazide plus DHB (DHB/Q3CH), was proposed for mass calibration and rapid detection of reducing N-glycans. Both DHB/3HBA and DHB/Q3CH show high derivatization efficiency and can improve the ionization efficiency of reducing N-glycans significantly. For mass calibration, in combination with dextrans, DHB/3HBA and DHB/Q3CH prove to be highly sensitive matrices facilitating both MS and MS2 calibration for N-glycans in dual polarities. For rapid identification, the regular mass difference observed for each N-glycan labeled with Q3CH and 3HBA respectively can eliminate the occurrence of false positives and promote automated identification of N-glycans in complex samples. For relative quantitation, the acid-base pair of DHB/Q3CH generates a concentrated cocrystallization of glycan-matrix mixtures at the edge of the droplet uniformly, exhibiting good linearity (R2 > 0.998) and accuracy (RSD ≤ 10%). Furthermore, the established POTDS was successfully utilized to assess N-glycans of serum from HCC patients, revealing potential for biomarker discovery in clinical practice.