Anomeric discrimination and rapid analysis of underivatized lactose, maltose, and sucrose in vegetable matrices by U‐HPLC–ESI‐MS/MS using porous graphitic carbon

Abstract

Lactose intolerance is a common condition caused by intestinal lactase deficiency, and a lactose-free diet represents the simplest way to avoid gastrointestinal symptoms. The emerging use of dietary supplements requires analytical tools that are capable of assessing these analytes, particularly for those based on dry herbal extracts that contain lactose together with maltose and sucrose, because of cross-contamination and/or deliberate addition as excipient. Electrospray ionization mass spectrometry (ESI-MS) and MS/MS are valuable detection methods for underivatized disaccharides; however, the absence of distinctive ions and collision-induced dissociation (CID) fragmentation patterns does not allow discrimination of stereoisomers without good chromatographic resolution. We developed an ultrahigh performance liquid chromatography-ESI (U-HPLC-ESI) approach, based on porous graphitic carbon (PGC) columns, working at 5 °C to separate and detect the disaccharides in their anomeric forms as formate adducts obtained directly in-column by eluting with formate buffer/acetonitrile gradient mixtures. Using a Paul trap, we monitored the adducts [M + HCOO](-) at m/z 387 in ESI negative mode (MS(1)) as well as the CID fragment ion [M - H](-) at m/z 341 (MS(2)) and used MS(3) fragment ions at m/z 178 and 161 to confirm disaccharides identity in complex vegetable matrices. Complete resolution of lactose α- and β-anomers, maltose α- and β-anomers, and sucrose was obtained with R ≥ 2.0 for all peaks and selectivity α = 1.2 between α- and β-anomers of lactose. The limits of detection were in the range of 3-7 µg/l (ppb) for the target disaccharides. Because of the rapidity and good anomeric discrimination, the described method represents an alternative tool to investigate the mutarotation phenomenon for reducing disaccharides.

Absolute Quantitation of Human Milk Oligosaccharides Reveals Phenotypic Variations during Lactation

“…For the analysis of this compound, the 2 peaks were summed. The separation of anomers at the reducing end is present in other separation media such as PGC (37). For this reason, oligosaccharides often were reduced to produce a single alditol at the reducing end.…”

Quantification of lactose content in human and cow's milk using UPLC–tandem mass spectrometry

“…Mulroney et al demonstrated that measurement of ␤-linked glucose disaccharides in the negative ion mode provided more structural information than in the positive ion mode because deprotonation of the anomeric hydroxyl causes fragmentation [12]. Recently, lactose, maltose, and sucrose in vegetable matrices were detected by U-HPLC-MS n in the negative SRM mode using only mother ions for quantification [20]. Moreover, HPLC combined with corona charged aerosol detector (CAD) was presented in the PITTCON conference as a new lactose determination method [18].…”

CH/π Interactions in Carbohydrate Recognition

“…There are many other examples of artificial systems indicating an important role of carbohydrate–aromatic interactions in molecular recognition. For example, porous graphitic carbon columns can be successfully applied in the chromatography of carbohydrates, despite their high polarity [ 26 ]. Carbohydrates were also successfully used in the solubilisation of carbon-based nanomaterials such as carbon nanotubes [ 27 , 28 ] ( Figure 4 B).…”