Lactic acid bacterium isolated from fermented durian (tempoyak) was investigated for its potentials as a probiotic strain. Bacterial tolerance toward gastrointestinal environment, adhesion, and cytotoxic activity in human colon adenocarcinoma cell line HT‐29 was evaluated. 16S rRNA sequencing identified the lactic acid bacterium as Lactobacillus plantarum . The bacterium demonstrated good tolerance toward gastrointestinal pH 2.0 and 0.3% bile salts. It showed strong adhesive capacity in human intestinal cell line, HT‐29, with an adhesion index of 159 ± 10. Cytotoxicity of L. plantarum was investigated using both live bacterial cells (BC) and cell‐free supernatant (CFS). Findings showed that both BC and CFS of L. plantarum reduced proliferation of HT‐29 colon adenocarcinoma cells using MTT assay. The results imply potential probiotic properties of L. plantarum isolated from tempoyak.