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Development and validation of a bioanalytical method of analyzing 3 ‘- and 6 ‘-sialyllactose using liquid chromatography-tandem mass spectrometry in minipig plasma and its application in a pharmacokinetic study
Sialyllactose (SL) is an acidic oligosaccharide, consisting of a combination of sialic acid and lactose. It is found in human milk. It has immune-protective effects against pathogens in newborns and helps with the development of the immune system and intestinal microorganisms. We developed and validated a method by which 3′-SL and 6′-SL levels were simultaneously analyzed via liquid chromatography-tandem mass spectrometry (LC-MS/MS), and evaluated the pharmacokinetics of the materials after systemic delivery to minipigs. To improve chromatographic selectivity, several types of columns (C18, amide, and HILIC phase) were used to separate the peaks of 3′-SL and 6′-SL. Ultimately the HILIC phase column was selected, as it had a good peak shape and quick resolution. The mobile phase comprised ammonium acetate buffer and acetonitrile with gradient elution. MS was performed in the negative ion and multiple reaction monitoring modes. Plasma samples were prepared using the protein precipitation method with methanol. A surrogate matrix was used for quantification because SLs are endogenous plasma compounds. The method developed was validated according to U.S. Food and Drug Administration guidance. A pharmacokinetic study was performed with intravenous administration of 3′-SL and 6′-SL in minipigs (Sus scrofa/Yucatan). The concentrations of 3′-SL and 6’-SL were readily measurable in the plasma samples, which suggests that the method adequately determined systemic exposure in minipigs. (C) 2020 Elsevier B.V. All rights reserved.
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