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In this study, low-carbon hypoeutectoid steels with different ferrite-pearlite microstructures were fabricated by varying transformation temperature. The microstructural factors such as pearlite fraction and interlamellar spacing, and cementite thickness were quantitatively measured and then Charpy impact tests conducted on the specimens in order to investigate the correlation of the microstructural factors with impact toughness and ductile-brittle transition temperature. The microstructural analysis results showed that the pearlite interlamellar spacing and cementite thickness decreases while the pearlite fraction increases as the transformation temperature decreases. Although the specimens with higher pearlite fractions have low absorbed energy, on the other hand, the absorbed energy is higher in room temperature than in low temperature. The upper-shelf energy slightly increases with decreasing the pearlite interlamellar spacing. However, the ductile-brittle transition temperature is hardly affected by the pearlite interlamellar spacing because there is an optimum interlamellar spacing dependent on lamellar ferrite and cementite thickness and because the increase in pearlite fraction and the decrease in interlamellar spacing with decreasing transformation temperature have a contradictory role on absorbed energy.