Abstract: In this paper, comparisons of the the molecular weight, monosaccharide composition, infrared spectrum were made between Dendrobium officinale polysaccharides from three different provenances, namely Yunnan, Danxia and Zhejiang. Together with the evaluation of preliminary pharmacological activity, the differences of Dendrobium candidum from different areas were discussed. High performance gel permeation chromatography (HPGPC) was used to determine the molecular weight of the purified polysaccharide fragments, high performance liquid chromatography (HPLC) was used to determine the composition of monosaccharides, infrared spectroscopy (IR) was used to determine the structure of polysaccharides, and MTT method was used to observe the effect of polysaccharide fragments on the proliferation of two kinds of tumor cells (Hela cells and HT-29 cells). The results showed that the molecular weight of the polysaccharide fragment of Dendrobium officinale in Yunnan was 757623 u, the molecular weight of polysaccharide fragment of Dendrobium officinale in Zhejiang was 605958 u, and the molecular weight of Dendrobium officinale polysaccharide fragment in Danxia was 663240 u. The monosaccharide composition of Dendrobium officinale polysaccharide was mainly mannose and glucose. In terms of the ratio of mannose to glucose, Yunnan species>Danxia species>Zhejiang species. Moreover, the ratio of polysaccharides would further increase after purification. Infrared spectrum absorption and characteristic peaks suggested that Dendrobium officinale polysaccharides mainly contained mannose and glucose, and the sugars contained were β-type. Purification of Dendrobium officinale polysaccharides could change the composition ratio of monosaccharides and increase the content of mannose. There were certain differences in molecular weight, monosaccharide composition and infrared spectroscopy of Dendrobium officinale polysaccharides from different provenances, and they had certain inhibitory effect on HT-29 cells. Particularly, the polysaccharide fragments of Dendrobium officinale from Danxia species had a relatively good inhibitory effect on the proliferation of the two tumor cells under the 48 h experimental determination. The specific correlation needs to be further studied.