Abstract
The structural diversity of human milk oligosaccharides (HMOs) strongly depends on the Lewis (Le) blood group status of the donor which allows a classification of these glycans into three different groups. Starting from 50 μL of human milk, a new high-throughput, standardized, and widely automated mass spectrometric approach has been established which can be used for correlation of HMO structures with the respective Lewis blood groups on the basis of mass profiles of the entire mixture of glycans together with selected fragment ion spectra. For this purpose, the relative abundance of diagnostically relevant compositional species, such as Hex2Fuc2 and Hex3HexNAc1Fuc2, as well as the relative intensities of characteristic fragment ions obtained thereof are of key importance. For each Lewis blood group, i.e., Le(a − b+), Le(a + b−), and Le(a − b−), specific mass profile and fragment ion patterns could be thus verified. The described statistically proven classification of the derived glycan patterns may be a valuable tool for analysis and comparison of large sets of milk samples in metabolic studies. Furthermore, the outlined protocol may be used for rapid screening in clinical studies and quality control of milk samples donated to milk banks.
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Abbreviations
- ACN:
-
Acetonitrile
- ATT:
-
6-Aza-2-thiothymine
- DMB:
-
2-Diamino-4,5-methylenedioxybenzene
- Fuc:
-
Fucose
- FucT:
-
Fucosyltransferase
- Hex:
-
Hexose
- HexNAc:
-
N-acetylhexosamine
- HMO:
-
Human milk oligosaccharide
- LID:
-
Laser-induced dissociation
- LNFP I:
-
Lacto-N-fucopentaose I (Fuc(α1-2)Gal(β1-3)GlcNAc(β1-3)Gal(β1-4)Glc)
- LNFP II:
-
Lacto-N-fucopentaose II (Gal(β1-3)[Fuc(α1-4)]GlcNAc(β1-3)Gal(β1-4)Glc)
- LNnT:
-
Lacto-N-neotetraose
- LNT:
-
Lacto-N-tetraose
- MALDI-TOF-MS:
-
Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry
- MS/MS:
-
Tandem mass spectrometry
- PGC:
-
Porous graphitic carbon
- Sia:
-
Sialic acid
- SPE:
-
Solid-phase extraction
- TFA:
-
Trifluoroacetic acid
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Acknowledgments
We thank Professor G. Bein and Professor H. Jomaa (Institute of Immunology and Transfusion Medicine, University Hospital Giessen-Marburg, Germany) for performing the Lewis blood group determination of blood samples as well as Dr. P. Gilbert (St. Josef’s Hospital Giessen) for the collection of milk and blood samples.
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Dennis Blank, Sabine Gebhardt, and Kai Maass contributed equally to this work.
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Blank, D., Gebhardt, S., Maass, K. et al. High-throughput mass finger printing and Lewis blood group assignment of human milk oligosaccharides. Anal Bioanal Chem 401, 2495–2510 (2011). https://doi.org/10.1007/s00216-011-5349-9
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DOI: https://doi.org/10.1007/s00216-011-5349-9