Background: The paranasal sinuses serve as a reservoir of nitric oxide (NO), contributing to baseline nasal NO (nNO) levels. nNO has also been shown to increase transiently with humming, a response that may be blunted in severe rhinosinusitis. Blunting of the acoustically-induced nNO transient ("spike") has been proposed as a screening test for osteomeatal complex (OMC) obstruction in sinusitis. Preparatory to conducting a clinical evaluation study, to eliminate variation in patient effort during this maneuver, we evaluated the use of external acoustic energy-in place of humming-to elicit nNO transients, documenting the effects of varying stimulus amplitude and gas sampling rates. Methods: Non-smoking, non-asthmatic subjects with no history of chronic sinusitis or nasal polyposis underwent nNO measurements in triplicate under: (1) control (quiet) conditions, and (2) with 128 Hz external acoustic energy. In Experiment 1, twelve subjects were exposed to two different intensities of external acoustic energy at 3 L/min sampling rate. In Experiment 2, a subset of nine subjects was sampled with and without acoustic stimulation at three different gas sampling rates (1, 2, and 3 L/min). Results: Experiment 1: Subjects, as a group, showed intensity-related increases in nNO with increasing acoustic amplitude (p < 0.01). Experiment 2: independently, both applied acoustic energy and lower nasal gas sampling rates increased measured nNO levels (p < 0.05 to p < 0.0001). Longitudinally, baseline (quiet) nNO obtained on a repeated basis in the two experiments (n = 9) was highly reproducible (R 2 = 0.84; p < 0.001), and acoustically-stimulated nNO was moderately so (R 2 = 0.50; p < 0.05). Conclusions: Application of external acoustic energy is a practical alternative to humming for mobilizing NO from the paranasal sinuses, and could be more objectively applied in any future validation studies involving clinical sinusitis and/or OMC obstruction.