Glucuronic acids in Arabidopsis thaliana xylans exist in 4- O -methylated (MeGlcA) and non-methylated (GlcA) forms at a ratio of about 3:2. The matrix-assisted laser desorption/ionization mass spectrometry analysis of the endoxylanase liberated acidic oligosaccharides from the Arabidopsis inflorescence stem showed that two peaks with GlcA (GlcA-Xyl 4 Ac 1 and GlcA-Xyl 5 Ac 2 ) had abnormally high intensities, as well as different tandem mass spectra, than their 4- O -methylated counterparts. These peaks were interestingly enriched in the xylan biosynthesis mutant irx7 and irx9-1 . Multi-stages fragmentation analysis using negative ion electrospray-ion trap mass spectrometry indicated that this GlcA was further carrying a pentose residue in the glucuronoxylan-derived oligosaccharide from irx9-1 . The structure was also identified in Arabidopsis wild type. The results prove evidence of a new pentose substitution on the GlcA residue of Arabidopsis GX, which is likely present in the primary walls. [ABSTRACT FROM AUTHOR]