Medlar (Mespilus) is an important fruit and is rich in nutritional and medical properties. Bioactive content and phenolic compounds of leaf, fruit, and seed samples of common medlar (M. germanica) and Stern’s medlar (M. canescens) were studied. The coefficient of variation showed high values among all the characters and was more than 50.00%, ranging from 59.74 to 196.81%. The leaf of M. canescens showed the highest total phenolic content (14.73 mg/g dry weight, DW) and followed by 13.70 mg/g DW in the seed of M. germanica collected from Ilam province, Iran. The highest flavonoid content was observed in fruits of M. germanica collected from the north of Iran (0.90 mg/g DW), and followed by the leaf of M. canescens (0.53 mg/g DW). The phenolic compounds showed strong variation. Principal component analysis showed that four PCs explained 95.07% of the observed total variance. Ward dendrogram indicated two different clusters based on the characters measured, indicating high variation among the accessions. The current investigation clearly showed the potential value of the common medlar (M. germanica) and Stern’s medlar (M. canescens) germplasm, as different medlar organs are significant sources of phenolic compounds and high antioxidant activity. Therefore, these species can be considered suitable sources of natural antioxidants, and may show potential future use in food and nutraceutical supplement formulations.