The pseudobulb of Oncidium orchid is a storage organ for supplying water, minerals and carbohydrates to the developing inflorescence. Different patterns of mannan, starch and pectin metabolism were observed in the pseudobulb of three developmental stages by histochemical staining and high performance anion exchange chromatographic (HPAEC) analysis. Copious pectin was strongly stained by ruthenium red in young pseudobulbs demonstrating that mannan and pectin were preferentially accumulated in the young pseudobulb sink at inflorescence pre-initiation stage. Concomitant with the emergence of the inflorescence, mannan and pectin decreased gradually and converted to starch. The starch, synthesized at the inflorescence developing stage, was eventually degraded at the floral development stage. A systematic survey on the subtractive EST (expression sequence tag) library of pseudobulb in the inflorescence pre-initiation stage revealed the presence of five groups of gene homologues related to sucrose, mannan, starch, pectin and other carbohydrate metabolism. The transcriptional level of 13 relevant genes related to carbohydrate metabolism was characterized from pseudobulbs of three different developmental stages. The specific activities of the enzymes encoded by these genes were also assayed. The expression profiles of these genes show that the transcriptional levels largely correlated with the enzyme activities, which were associated with the respective carbohydrate pools. These results demonstrated a novel functional profile of polysaccharide mobilization pathway as well as their relevant gene expression in the pseudobulb of Oncidium orchid during the flowering process.