Determining Neu5Ac in infant formula with ultra‐performance liquid chromatography–tandem mass spectrometry

Abstract

The aim of this work was to measure N-acetylneuraminic acid (Neu5Ac) in milk-based infant formulas. The analysis was performed by ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The total Neu5Ac were released using trichloroacetic acid and hydrochloric acid and purified using a HLB column. The linearity from 0.05 to 5.0 lg/mg Neu5Ac was adequate. Sialic acid recoveries ranged from 91.8% to 112.4%. The detection and quantification limits (limit of detection, 0.01 lg Neu5Ac/mg; limit of quantitation, 1.08 lg Neu5Ac/mg) were low enough to determine the sialic acid in infant formulas. The validated method is highly reproducible and sensitive, and it is easy to perform.

Evaluation of Sialic Acid in Infant Feeding: Contents and Bioavailability

“…Neu5Ac represented 95.5–99% of total Sia, the proportion being similar to that found in mature bovine milk. ,,, The Sia values found in our study agree with the wide range in Sia concentration in IFs recorded in previous studies (Table ). ,,,,,− On the other hand, larger Sia concentrations have been reported in bovine mature milk (6–22 mg/100 mL), ,,− which could partially explain the higher Sia values recorded in our study. The highest Sia content described in the literature was attributed to the addition of colostrum to an IF.…”

“…Sialic acids are bioactive compounds present in HM (9–155 mg/100 mL) ,,− and in lesser concentrations in IFs (6.5–32.5 mg/100 mL). ,,,,,,− The differences in Sia concentrations may be related to different factors such as the stage of lactation of HM, ingredients used in the formulation of IFs, or the analytical methodology used (spectrophotometric and HPLC methods) among others. From nutritional and functional points of view, it is interesting to know not only the Sia contents in infant foods and the intakes but also the bioavailability (i.e., the fraction of an ingested nutrient or bioactive compound that is available for use in physiological functions or for storage).…”

Explorations in a galaxy of sialic acids: a review of sensing horizons, motivated by emerging biomedical and nutritional relevance

“…448 The ultra-HPLC coupled with positive-ion mode ESI source and selective reaction monitor tandem quadrupole MS/MS detector were also used to determine Neu5Ac in the milk-based commercial infant formulas with LDR 0.05-5.0 μg mg −1 . 449 However, the formation of Na + /K + /NH 4 + adduct with the COO − functional group of Sia sometimes limits the direct HPLC-MS assay. Fernando et al protected the carboxyl group of Neu5Ac with fluorophore 1,2-diamine-4,5-dimethoxyl benzene dihydrochloride (excitation and luminescence maxima at 369 and 453 nm, respectively) and carried out the HPLC-MS assay coupled with scanning fluorescence detector, positive ion mode ESI source and quadrupole mass spectrometer with an internal standard α-keto glutaric acid.…”

“…448 The ultra-HPLC coupled with positive-ion mode ESI source and selective reaction monitor tandem quadrupole MS/MS detector were also used to determine Neu5Ac in the milk-based commercial infant formulas with LDR 0.05–5.0 μg mg −1 . 449 However, the formation of Na + /K + /NH 4 + adduct with the COO − functional group of Sia sometimes limits the direct HPLC-MS assay.…”

A novel, sensitive and convenient method for determination of sialic acids in human serum utilizing ultrasonic-assisted closed in-syringe hydrolysis and derivatization prior to high performance liquid chromatography

“…The critical step in sialic acid analysis is their liberation from human serum, which entails isolation of sialic acids from the parental glycoconjugate. The blood composition is more complex; multi-step manual operations such as solid-phase extraction (SPE) 31 are usually required to remove impurities from the sample. However, multi-step operations are tedious and time-consuming, and more seriously, tend to cause the loss of analytes and high reagent-consumption, which will pose a potential threat to experimenters and the environment.…”