The purpose of the study was to compare selected physiological responses during singles tennis match play and continuous running at a similar mean oxygen uptake (VO2). The study consisted of two main parts, which were separated by 1 week. In the first part, 12 nationally ranked senior tennis players [six females and six males; 47.2 (6.6) years old and 47.0 (5.4) years old, respectively] each completed a 2-h singles tennis match (TE). Mean VO2 during TE [23.1 (3.1) ml.kg(-1). min(-1) for the women and 25.6 (2.8) ml.kg(-1).min(-1) for the men] was measured by a portable spirometry-telemetry system and corresponded to 56% (women) or 54% (men) of their respective maximum VO2. In the second part, the relative VO2 data measured during TE were used to set a similar workload during a 2-h treadmill run at a constant level (RU). At the measured time points, heart rate [140.1 (15.5) beats.min(-1) vs 126.4 (15.1) beats. min(-1)], lactate concentration [1.53 (0.65) mmol.l(-1) vs 1.01 (0.38) mmol.l(-1)] and glucose concentration [5.45 (0.84) mmol.l(-1) vs 4.34 (0.56) mmol.l(-1)] in capillary blood, as well as the respiratory exchange ratio [0.93 (0.03) vs 0.88 (0.03)], were higher (P<0.05) in TE compared to RU. Serum concentrations of free fatty acids increased (P < 0.05) during both work loads [from 0.25 (0.15) mmol.l(-1) to 1.31 (0.44) mmol.l(-1) in TE and from 0.22 (0.17) mmol.l(-1) to 1.24 (0.35) mmol.l(-1) in RU]. Post-exercise urine concentrations of epinephrine [0.17 (0.14) micromol.l(-1) vs 0.08 (0.04) micromol.l(-1)] and norepinephrine [1.27 (0.59) micromol.l(-1) vs. 0.55 (0.33) micromol.l(-1)] were higher in TE (P<0.05). These results indicate a stronger metabolic emphasis on glycolysis and glycogenolysis and an overall enhanced sympathoadrenal activity during tennis match play compared to continuous running exercise at a similar mean VO2.