Short communication: Quantification of carbohydrates in whey permeate products using high-performance anion-exchange chromatography with pulsed amperometric detection

Abstract

A method was developed for the characterization and quantification of the disaccharide lactose and 3 major bovine milk oligosaccharides (BMO) in dairy streams. Based on high-performance anion-exchange chromatography-pulsed amperometric detection (HPAE-PAD), this method is advantageous because it requires minimal sample preparation and achieves good chromatographic separation of oligosaccharide isomers within 30 min. The linear dynamic range and limit of detection were 0.1 to 10 mg/L and 0.03 to 0.22 mg/L, respectively. Mean recoveries of the BMO were excellent and ranged from 98.4 to 100.4%. Without complicated sample preparation procedures, this HPAE-PAD method measured BMO [3′-sialyllactose (3′SL), 6′-sialyllactose (6′SL), and 6′-sialyllactosamine (6′SLN)] and lactose using a single instrument, therefore increasing the accuracy of the measurement and applicability for the dairy industry. In colostrum whey permeate, 3′SL, 6′SL, and 6′SLN were 94, 29, and 46 mg/L, respectively. This work is the first to demonstrate that some commercial products, currently marketed for supporting a healthy immune system, contain significant amounts of bioactive BMO and therefore, carry additional bioactivities.

Indole-3-lactic acid associated with Bifidobacterium-dominated microbiota significantly decreases inflammation in intestinal epithelial cells

“…To obtain lactose-free HMOs, a purification approach of pooled donor milk based on lactose hydrolysis, yeast consumption of the resulting monosaccharides and further clean-up and concentration by sequential membrane filtration was used [ 65 ]. The freeze-dried powder was characterized by mass spectrometry and analyzed by high-performance anion-exchange chromatography with pulsed amperometric detection using a previously published method tailored for oligosaccharide quantification in milk [ 66 ] (Supplemental Table 1 ). B. infantis was incubated at 37 °C until early stationary phase under anaerobic conditions (5% carbon dioxide, 5% hydrogen, and 90% nitrogen) in a vinyl anaerobic chamber (Coy Laboratory Products).…”

Comparative Genome Analysis of Bifidobacterium longum subsp. infantis Strains Reveals Variation in Human Milk Oligosaccharide Utilization Genes among Commercial Probiotics

“…Supernatant was collected and diluted 10,000-fold. Dilutions were filtered through a 0.22 μm cellulose acetate membrane (VWR International, Radnor, PA, USA) and 25 µL of these supernatants were injected into a High-Performance Anion-Exchange Chromatograph Coupled with Pulsed Amperometric Detection instrument (HPAE-PAD ICS-5000, Thermo Scientific, Sunnyvale, CA, USA) according to methods from [ 43 ] with some modifications. Briefly, chromatographic separation was carried out on a CarboPac PA1 analytical column (4 × 250 mm, DionexTM, ThermoFisher Scientific, Waltham, MA, USA) and CarboPac PA1 guard column (4 × 50 mm, Dionex) with an isocratic gradient, 0–30 min 73.5% A, 25% B, 1.5% C, at a 1.0 mL/min flow rate, where solvent A was deionized water, solvent B 100 mM NaOH and solvent C was 500 mM NaOAc in 100 mM NaOH.…”

Role of pH in the recovery of bovine milk oligosaccharides from colostrum whey permeate by nanofiltration

“…Major BMO 3’-sialyllactose (3’-SL), 6’-sialyllactose (6’-SL), and 6’-sialyllactosamine (6’-SLN) as well as the simple sugars lactose, glucose, and galactose were quantified by high performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD; Thermo Scientific HPAEC-PAD ICS-5000, Sunnyvale, CA, USA) (Lee, de MeloSilva, Liu, & Barile, 2015). Samples were diluted between 10 and 1000 times with nanopure water as appropriate and filtered through a 0.2 µm syringe filter (Acrodisc 13mm PES, Pall Life Sciences, Port Washington, NY, USA).…”