Abstract Weaning is a stressful event that affects animal welfare and performance. This study examined the effects of industrial hemp (Cannabis sativa) supplementation on behavior, chute score and physiological responses of cow-calf pairs during weaning. Ten apparently healthy Angus cow-calf pairs, average BCS of 6.0, were randomly allocated to one of two groups for 7 days: Control (CON, 200g/head of commercial grower feed, n=10) and Hemp (HMP, 200g/head containing 80% commercial grower feed containing 20% hemp, n=10), daily. Hay and water were provided ad libitum. Chute and exit scores were recorded and blood samples were collected at d 0, 4 and 7 of the feeding period. Animal activity data was recorded using automated activity monitoring systems Moomonitor and IceQubes for cows and calves, respectively. At weaning, cow-calf pairs were separated abruptly and observed. Data was analyzed using Stata 14.2. Overall, rectal temperature, respiratory rate, rumination duration, rumen fluid pH and activity did not differ (P > 0.05) between treatments. Specifically, hemogram and serum biochemistry results did not differ (P > 0.05) between cow groups. However, resting duration tended to be greater in HEMP compared with CON (383.94±19.3 vs 298.60±16.1 mins/day, P = 0.06) cows. In addition, feeding duration was greater in HEMP compared with CON (473.8±24.5 vs 262.3±31.6 min/day) cows. In calves, heart rate was lower in HEMP compared with CON (62.67 vs 94.7 beats/min, P = 0.03). Hemoglobin concentration was smaller in HEMP compared with CON (12.6 vs 14.1mg/mL, P = 0.02). However, mean corpuscular hemoglobin concentration was greater in HEMP compared with CON (33.04 vs 32.04g/dl, P = 0.04). We concluded that hemp supplementation increased resting and feeding duration of cows, and reduced heart rate of calves during weaning. This is the first report on the use of hemp to improve welfare of cow-calf pairs during weaning.