Yams make a significant contribution to food security and medicinal importance in developing countries. In Ethiopia, there is insufficient scientific study on biochemical composition of Ethiopian yams. In order to fill the knowledge gap, this study was conducted to assess the biochemical composition of yams collected from Southwest Ethiopia. Flour from storage tuber of 36 yam landraces collected and the samples run in duplicates. Data on 14 biochemical traits were collected and subjected to various data analysis. Results of the analysis of variance indicated significant variation (p<0.01) among the landraces on organic matter, total nitrogen, protein, fat, carbohydrate, total phosphorus, total energy, tannin and saponin contents. The flour moisture contents ranged from 17.75 to 27.47% with a mean of 22.03%. The ranges of dry matter (15.80 to 27.28%), organic matter (21.38 to 43.56%), ash (1.13 to 3.56%), organic carbon (0.63 to 1.98 g), crude fiber (0.41 to 2.05%), total nitrogen (1.00 to 1.32%), protein (6.25 to 8.28%), fat (0.09 to 0.65%), carbohydrate (12.71 to 33.94%), total phosphorus (23.7 to 53.0 mg/100 g), total energy (92.66 to 173.30 kcal/100 g DM), tannin (19.80 to 181.0 mg/100 g) and saponin (2.31 to 13.94 mg/100 g) contents. The cluster and distance analysis of biochemical traits showed the existence of eight divergent groups. The maximum inter cluster distance was found between clusters VI and VII (133.59), followed by clusters V and VI (109.19), clusters II and VI (105.22), clusters I and VI (100.42), and clusters III and VI (89.25) in order of magnitudes. Maximum genetic divergence between the clusters points out the fact that hybridization among the landraces included with them would produce potential and meaningful hybrids and desirable segregants. Besides, investigation of the existed yam landraces based on molecular marker analysis is vital for better assessment of genetic diversity of yams in Ethiopia.