Negative-ion electrospray tandem mass spectrometry (ES-MS/MS) with collision-induced dissociation (CID) is attempted for sequence determination of alginate oligosaccharides, derived from polyanionic alginic acid, polymannuronate, and polyguluronate by partial depolymerization using either alginate lyase or mild acid hydrolysis. Sixteen homo-and hetero-oligomeric fragments were obtained after fractionation by gel-filtration and strong anion exchange high performance liquid chromatography. The product-ion spectra of these alginate oligosaccharides were dominated by intense B-, C-, Y-, and Z-type ions together with 0,2 A-and 2,5 A-ions of lower intensities. Internal mannuronate residues (M) produce weak but specific decarboxylated Z int -ions (Z int Ϫ 44 Da; int: denotes internal), which can be used for distinction of M and a guluronate residue (G) at an internal position. A reducing terminal M or G, although neither gives rise to a specific ion, can be identified by differences in the intensity ratio of fragment ions of the reducing terminal residue [ [2,3]. Infection with the latter opportunistic pathogen seen in cystic fibrosis patients remains a major health concern [4]. Alginate consists of hexuronic acid residues -Dmannuronic acid (M) and ␣-L-guluronic acid (G) with exclusively 1¡4 glycosidic linkages. Along its linear chain, there are homo-oligomeric regions of mannuronate (M-blocks) and of guluronate (G-blocks) as well as hetero-oligomeric regions (MG-blocks) [5,6]. In contrast to algal alginate and alginate of A. vinelandii, the absence of G-blocks or low ratio of G-to M-blocks is characteristic for alginate from most bacteria such as P. aeruginosa. The alginate coat of P. aeruginosa is known to provide protection from immune defence system. Oligomannuronates have been shown to have an inhibitory effect on reactive oxidizing species production [7] from immune cells, whereas oligoguluronates with similar sizes did not exhibit any significant effect [8]. The physical properties of the alginates have also been well documented and can be used as thermally stable cold setting gelling agents in the presence of calcium ions, gelling at far lower concentrations than gelatin. High G content produces strong, brittle gels with good heat stability whereas high M content produces weaker, more-elastic gels with good freeze-thaw behavior. Alginate oligosaccharide fragments and their derivatives have been widely used as releasing agents in the pharmacy [9], as additives in the food industry [10], and as growth-promoters in agriculture [11]. Recently, it has also found important applications in tissue engineering [12].Polyanionic oligosaccharides such as those derived from glycosaminoglycans have attracted considerable interests recently because of the increased awareness of their functional properties. The functional properties of alginates and their oligosaccharide fragments are intimately linked to the composition (M/G ratio) and sequences of the hexuronic acid M and G residues. Methods for identification of oligosacc...