There are heterogeneous subgroups among those with heartburn, and data on these individuals are relatively scant. We aimed to evaluate the effect of acid challenge on the segmental contractions of esophageal smooth muscle in endoscopy-negative patients with normal esophageal acid exposure. High-resolution esophageal manometry (HRM) was performed on 30 endoscopy-negative patients with heartburn accompanied by normal esophageal acid exposure using 10 water swallows followed by 10 acidic pomegranate juice swallows. Patients were classified into functional heartburn (FH) and hypersensitive esophagus (HE) groups based on the results of 24-hr impedance pH testing. HRM topographic plots were analyzed and maximal wave amplitude and pressure volumes were measured for proximal and distal smooth muscle segments. The pressure volume of the distal smooth muscle segment in the HE group measured during acidic swallows was higher than during water swallows (2224.1 ± 68.2 mmHg/cm per s versus 2105.6 ± 66.4 mmHg/cm per s, P = 0.027). A prominent shift in the pressure volume to the distal smooth muscle segment was observed in the HE group compared with the FH group (segmental ratio: 2.72 ± 0.08 versus 2.39 ± 0.07, P = 0.005). Manometric measurements during acidic swallows revealed that this shift was augmented in the HE group. The optimal ratio of pomegranate juice swallowing for discrimination of FH from HE was 2.82, with a sensitivity of 88.9% and a specificity of 100%. Hypercontractile response of distal smooth muscle segment to acid swallowing was more prominent in the HE group than the FH group.