Psidium guajava fruit is a subtropical fruit, functional food and traditional medicine for the adjuvant treatment of diabetes mellitus in China. To investigate the active components responsible for its health benefits, a novel heteropolysaccharide GP70-3 was purified by water extraction, ethanol precipitation and column chromatography. Structural characterization of GP70-3 was elucidated for the first time by monosaccharide composition assay, Fourier transform-infrared spectroscopy (FT-IR), methylation analysis, gas chromatography coupled with mass spectrometry (GC-MS) and nuclear magnetic resonance spectroscopy (NMR). The data revealed that GP70-3 contained a backbone of 1→3,6)-linked β-d-Galp, 1→5)-linked α-l-Araf, 1→6)-linked β-d-Galp and 1→3)-linked β-d-Galp, branched with 1→2,3,5)-linked α-l-Araf, 1→3)-linked α-l-Araf, 1→3)-linked α-l-Rhap, 1→3)-linked β-d-GlcpA, 1→3)-linked β-d-GalpA and terminated with →1)-linked β-d-Galp. Advanced structure studies showed GP70-3 consisted of irregular flakes with rounded-spherical pores. Moreover, GP70-3 exhibited outstanding α-glucosidase inhibitory activity in vitro, with an IC50 value of 2.539 ± 0.144 μM, which was 1867 times higher than that of the positive control acarbose (IC50 value of 4.744 ± 0.026 mM). Therefore, consumption of guava polysaccharides may be beneficial as an α-glucosidase inhibitor for reducing the postprandial blood glucose level and treating type II diabetes.