This study was conducted to investigate the effect of dietary total mixed ration (TMR) supplementation and dry-aging period on the physico-chemical and sensory quality of striploin (longissimus lumborum) from multiparous Hanwoo cows. The experimental cows were divided into two treatments (n=3/treatment) according to the type of feed: in one treatment (slaughter age: 87±6 months old, parity: 5±1) the cows were fed on TMR for 7 months before slaughter, while in the other treatment (slaughter age: 83±9 months old, parity: 5±2) the cows were fed on concentrate and rice straw (CRS) for the whole rearing period. After slaughtering, the striploins were dry-aged for 20 or 40 days at 2±1℃, 85% relative humidity and 2 m/sec air flow velocity and then utilized for quality measurements. The pH value, cooking loss, and hypoxanthine content were higher (p<0.05) for the striploins dry-aged for 40 days than for those dry-aged for 20 days. Notably, the inosine 5’-monophosphate content was decreased (p<0.05) by increasing the dry-aging time. With regard to microbiological quality, the striploins dry-aged for 40 days exhibited lower (p<0.05) lactic acid bacteria counts and higher (p<0.05) coliform counts than those dry-aged for 20 days. Moreover, the tenderness and overall liking scores based on sensory evaluation were higher (p<0.05) for the dry-aged striploins of the TMR treatment and those dry-aged for 40 days than for the CRS treatment and those dry-aged 20 days, respectively. These findings suggest that dietary TMR supplementation leads to an increase in the sensory preference of dry-aged striploins from multiparous Hanwoo cows, and alteration of the CRS into the TMR may improve the quality of dry-aged Hanwoo beef.