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Detection of milk oligosaccharides in plasma of infants

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Abstract

Human milk oligosaccharides (HMO) are one of the major components of human milk. HMO are non-digestible by the human gut, where they are known to play important functions as prebiotics and decoys for binding pathogens. Moreover, it has been proposed that HMO may provide sialic acids to the infant that are important in brain development, however this would require absorption of HMO into the bloodstream. HMO have consistently been found in the urine of humans and other mammals, suggesting systemic absorption. Here, we present a procedure for the profiling of milk oligosaccharides (MO) in plasma samples obtained from 13 term infants hospitalized for surgery for congenital heart disease. The method comprises protein denaturation, oligosaccharide reduction, and porous graphitized carbon solid phase extraction for purification followed by analysis using nHPLC-PGC-chip-TOF-MS. Approximately 15 free MO were typically observed in the plasma of human infants, including LNT, LDFP, LNFT, 3′SL, 6′SL, 3′SLN, and 6′SLN, of which the presence was confirmed using fragmentation studies. A novel third isomer of SLN, not found in human or bovine milk was also consistently detected. Differences in the free MO profiles were observed between infants that were totally formula-fed and infants that received at least some part breast milk. Our results indicate that free MO similar in structure to those found in human milk and urine are present in the blood of infants. The method and results presented here will facilitate further research toward the possible roles of free MO in the development of the infant.

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Abbreviations

FA:

Formic acid

FL:

Fucosyllactose

Fuc:

Fucose

Gal:

Galactose

Glc:

Glucose

GlcNAc:

N-acetylglucosamine

HMO:

Human milk oligosaccharide

LDFP:

Lacto-difucosyl-pentaose

LNFP:

Lacto-N-fucopentaose

LNnT:

Lacto-N-neotetraose

LNT:

Lacto-N-tetraose

MO:

Milk oligosaccharide

Neu5Ac:

N-acetylneuraminic acid

PGC:

Porous graphitized carbon

RSD:

Relative standard deviation

SL:

Sialyllactose

SLN:

Sialyllactosamine

TFA:

Trifluoroacetic acid

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Acknowledgments

Funding was provided by the NIH (R01 HD059127 and UL1 TR000002); the authors declare no conflict of interest.

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Authors and Affiliations

  1. Department of Chemistry, University of California Davis, One Shields Avenue, Davis, CA, 95616, USA

    L. Renee Ruhaak, Carol Stroble & Carlito B. Lebrilla

  2. Department of Pediatrics School of Medicine, University of California Davis, 2516 Stockton Blvd, Sacramento, CA, 95817, USA

    Mark A. Underwood

Authors
  1. L. Renee Ruhaak
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  2. Carol Stroble
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  3. Mark A. Underwood
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  4. Carlito B. Lebrilla
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Correspondence to L. Renee Ruhaak.

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Ruhaak, L.R., Stroble, C., Underwood, M.A. et al. Detection of milk oligosaccharides in plasma of infants. Anal Bioanal Chem 406, 5775–5784 (2014). https://doi.org/10.1007/s00216-014-8025-z

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  • DOI: https://doi.org/10.1007/s00216-014-8025-z

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