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|Title:||Determination of Glucosamine by Ultra-high Pressure LC in Shrimp By-Products|
Soto Valdez, H.
|Publisher:||Instituto Nacional de Saúde Doutor Ricardo Jorge, IP|
|Abstract:||Glucosamine is one of the most abundant monosacharides and is part of the structure of the chitosan and chitin, which compose the exoskeletons of crustaceans, other arthropods and the fungi cell walls. Shrimp by-products are a good source of glucosamine because about 45% of the animal is composed of inedible cephalothorax and exoskeleton. The project ‘Preparation of active packaging with antioxidant and antimicrobial activity based on astaxanthin and chitosan’ aims to develop a methodology for the incorporation of compounds obtained from shrimp waste in plastic matrices for the development of an active packaging with antimicrobial and antioxidant properties. In the frame of this project, shrimp by-products were fermented and insoluble chitin was purified by a sequence of steps: depigmentation, deproteinization, demineralization and blanching. The aim of the present work was to optimize a method to determine glucosamine by Ultra-high Pressure Liquid chromatography (Ultra Performance Liquid Chromatography, UPLC) with diode array detection (DAD) from shrimp by-products. First, an acid hydrolysis of the chitin of shrimp by-products is carried out. Afterwards, a derivatization is done because glucosamine does not contain a chromophore. The selected derivatization reagent was 9-fluorenylmethyl-chloroformate (Fmoc-Cl). The chromatographic separation is achieved using a vanguard pre-column (Acquity UPLCÒ HSS T3, 1.8 µm particle size) and a column (Acquity UPLCÒ HSS T3, 2.1 x 50 mm, 1.7 µm particle size) at 38 °C. Quantification of glucosamine was carried out at 265 nm. The mobile phase is a gradient of phase A [water (pH 6.5)/ methanol, 85:15 (v/v)] and phase B (acetonitrile) with a flow rate of 0.3 mL/min. The optimized UPLC method allows an excellent separation in just 10 min of the two anomers of glucosamine, of the Fmoc-Cl and of its corresponding alcohol, which is the hydrolysis product of the spontaneous reaction between Fmoc-Cl with water. Several samples have been analysed, some are commercial samples (chitosan with low molecular weight; chitosan with medium molecular weight; chitosan from shrimp shells; chitosan from crab shells), and three other samples produced from shrimp by-products in the frame of the project. Commercial samples presented a higher amount of glucosamine content than in-house produced samples.|
|Appears in Collections:||DAN - Posters/abstracts em congressos internacionais|
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