Improved determination of milk oligosaccharides using a single derivatization with anthranilic acid and separation by reversed-phase high-performance liquid chromatography (original) (raw)
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Variations of neutral oligosaccharides and lactose in human milk during the feeding
Zeitschrift für Ernährungswissenschaft, 1993
There exist only few data concerning the variation of oligosaccharides in human milk. In this study the variations of neutral oligosaccharides and of lactose in human milk during the feeding were determined from five women at day 8 and at day 57 post partum. The milk of the investigated feedings was divided in four parts of equal volumes during sampling; the concentrations of neutral oligosaccharide fractions were determined by gel permeation chromatography on Fractogel TSK HW 40 (S) columns. No significant differences in the concentrations of the neutral oligosaccharide groups monofucosyllactoses, difucosyllactose, lacto-N-tetraoses, monofucosyllacto-N-tetraoses and difucosyUacto-Ntetraoses and of lactose were found in the four milk parts. The results of this study favor the use of socalled mid-feed samples, a simple and convenient sampling method for analytical studies of human milk. Mid-feed samples are representative of the whole feeding as concerned for neutral oligosaccharides.
ELECTROPHORESIS, 2014
Human milk is a unique fluid in glycobiology due to the presence of many free structurally complex oligosaccharides emerging as important dietary factors during early life and having many biological and protective functions. Methods that allow accurate profiling of oligosaccharide mixtures in this complex biological fluid with quantification of the four known genetically determined groups are welcomed. A high-voltage CE separation and detection at 254 nm of 17 neutral and acidic human milk oligosaccharide (HMO) standard along with lactose derivatized with 2-aminoacridone, using a BGE containing 20% methanol as an organic modifier and borate, able to form on-capillary anionic boratepolyol complexes, is reported. This CE approach was able to separate both neutral HMOs and acidic HMOs, with the sialic acid residue, also in the presence of lactose in high content. This method was applied to the four secretory groups individually extracted by a rapid and simple preparative step. LODs were found ranging from ß50 to 700 fmol. We were able to measure HMO content also in the presence of excess fluorophore, or interference from proteins, peptides, salts, and other impurities normally present in this complex biological fluid. Overall, CE equipped with a UV detector is a common analytical approach and this simple CE separation offers high resolution and sensitivity for the differentiation of human milk samples related to genetic groups and days of lactation by considering that important changes in HMO content are a reflection of the lactation day.
Milk Oligosaccharide Profiles by Reversed-Phase HPLC of Their Perbenzoylated Derivatives
Analytical Biochemistry, 1997
and pathogens involved in diseases of infants (2). These Human milk is rich in oligosaccharides, some of oligosaccharides are the third largest solid component in which inhibit toxins and pathogens involved in dismilk after lactose and triglycerides and are believed to eases of infants. To investigate qualitative and quantibe synthesized by the same glycosyltransferases that are tative individual variation of human milk oligosacchainvolved in the biosynthesis of glycoproteins and other rides, a sensitive method for routine identification and glycoconjugates. As a consequence, milk oligosaccharides quantification of intact milk oligosaccharides was decan have terminal carbohydrate structures similar to veloped and applied to milk samples from 50 donors. those present on glycoconjugates, some of which act as The isolated, reduced neutral oligosaccharide fraccell surface receptors for pathogens. Thus, some of these tions were perbenzoylated, resolved by reversedoligosaccharides could act as analogs or homologs to phase HPLC, and detected at 229 nm. This method rethose receptors, thereby providing protection to the nurssolves most structural isomers and does not require ing infant against enteric pathogens. Recent studies on stringent removal of lactose. Peaks were detected at complex sugar structures (3) have revealed the existence the low nanogram (pmol) level and peak areas were of a far greater number of oligosaccharides than what linear from 1 to 1000 mg for a standard oligosaccharide.
Journal of Food Science, 2020
In recent years, production of biosynthesized human milk oligosaccharides (HMOs) has become scalable to industrial standards. As a result, infant formula fortified with 2'-fucosyllactose (2'-FL), the most abundant HMO in human breast milk, is now commercially available. 2'-FL and 3-fucosyllactose (3-FL), another abundant HMO, are thought to be beneficial for infant health and development. Products containing HMOs are projected to expand in the future, showing the need for robust, easily applicable analytical methods for the quantitative assessment of HMOs in different food applications. We present here a validated high-performance liquid chromatography method for the quantification of 2'-FL and 3-FL in whole milk, infant formula, and cereal bars. The sample preparation was simple dispersion and extraction of the sample. The samples were analyzed by hydrophilic interaction liquid chromatography with refractive index detection and a runtime of 19 min. The method had a high degree of linearity (R 2 > 0.9995) in the range 0.2 to 12 mg/mL. The recovery for 2'-FL was 88% to 105% and for 3-FL 94% to 112%. The limit of detection (LOD) for whole milk was 0.1 mg/mL for 2'-FL and 0.2 mg/mL for 3-FL. In infant formula and cereal bars, the LOD was 0.6 mg/g for both 2'-FL and 3-FL. To show the practical application of this method, it was successfully utilized in stability studies of 2'-FL and 3-FL in whole milk, UHT milk, and yoghurt. The method provides a means of simultaneous and robust quantification of 2'-FL and 3-FL in various food matrices with high accuracy and high reproducibility.
Recent advances in the analysis of human milk oligosaccharides by liquid phase separation methods
Journal of Chromatography B, 2021
Human milk is a complex, dynamically changing biological fluid, which contains a large amount of nonconjugated carbohydrates, referred to as human milk oligosaccharides (HMOs). These HMOs are very important for the infants as they play important roles in the formation of the gut microbiome, the immune system and support brain development. HMOs show highly complex structural diversity due to numerous linkage possibilities of the building monosaccharides. In order to elucidate their structure-function relationship and to develop more effective infant formulas, cutting-edge analytical technologies are in great demand. In this paper, we review the current strategies for HMO analysis based on liquid phase separation methods. High performance liquid chromatography, capillary electrophoresis and their hyphenation with mass spectrometry are critically reviewed, emphasizing their advantages and disadvantages from practical point of views. Recent advances of the methods are categorized according to their application fields.
Analysis, structural characterization, and bioactivity of oligosaccharides derived from lactose
ELECTROPHORESIS, 2014
The increasing interest for prebiotic carbohydrates as functional food ingredients has promoted the synthesis of galacto-oligosaccharides and new lactose derivatives. This review provides a comprehensive overview on the chromatographic analysis, structural characterization and bioactivity studies of lactose-derived oligosaccharides. The most common chromatographic techniques used for the separation and structural characterization of this type of oligosaccharides, including GC and HPLC in different operational modes, coupled to various detectors are discussed. Insights on oligosaccharide MS fragmentation patterns using different ionization sources and mass analyzers, as well as data on structural analysis by NMR spectroscopy are also described. Finally, this article deals with the bioactive effects of galactooligosaccharides and oligosaccharides derived from lactulose on the gastrointestinal and immune systems, which support their consumption to provide significant health benefits.
A comparative study of free oligosaccharides in the milk of domestic animals
The British journal of nutrition, 2014
The present study was conducted to obtain a comprehensive overview of oligosaccharides present in the milk of a variety of important domestic animals including cows, goats, sheep, pigs, horses and dromedary camels. Using an analytical workflow that included ultra-performance liquid chromatography-hydrophilic interaction liquid chromatography with fluorescence detection coupled to quadrupole time-of-flight MS, detailed oligosaccharide libraries were established. The partial or full characterisation of the neutral/fucosylated, phosphorylated and sialylated structures was facilitated by sequencing with linkage- and sugar-specific exoglycosidases. Relative peak quantification of the 2-aminobenzamide-labelled oligosaccharides provided additional information. Milk from domestic animals contained a much larger variety of complex oligosaccharides than was previously assumed, and thirteen of these structures have been identified previously in human milk. The direct comparison of the oligosac...
Oligosaccharides in human milk, achievements in analysis: a review
Research for Rural Development 2022 : annual 28th International scientific conference proceedings
Human milk oligosaccharides (HMOs) comprise about 20% of the total carbohydrates of human milk. There is currently a growing interest in HMOs as many researchers have recognized the importance of their benefits to infant health. Accumulated evidence suggests that HMOs are anti-adhesive antimicrobials that serve as soluble bait receptors, prevent pathogens from attaching to infant mucous membranes, and reduce the risk of viral, bacterial, and protozoan parasites. It also provides functionality including anti-adhesion and immunomodulators. Even though the composition of human milk in Latvia has been studied in detail, there are no studies on oligosaccharides in human milk. The aim of the study is to find out recent advances in the analysis of HMOs. Semi-systematic method was used to analyze the latest information about the recent advances in the analysis of HMOs by liquid phase separation methods, to investigate any known associations between HMOs composition and maternal nutrition an...