Abstract Flexible surface micro-discharge plasma is a non-thermal plasma technique used for treating wounds in a painless way, with significant efficacy for chronic or hard-to-heal wounds. In this study, a confined space was designed to simulate wound conditions, gelatin was used to simulate wound tissue. The distinctions between open and confined spaces were explored, and the effects of temperature, humidity, discharge power, and the gap size within the confined space on the plasma characteristics were analyzed. It was found that, temperature, humidity and discharge power are important factors that affect the concentration distribution of active components and the mode transition between ozone and nitrogen oxides. Compared to open space, the concentration of ozone in confined space was relatively lower, which facilitated the formation of nitrogen oxides. In open space, the discharge was dominated by ozone initially. As temperature, humidity and discharge power increased, nitrogen oxides in the gas-phase products were gradually detected. In confined space, nitrogen oxides can be detected at an early stage and at much higher concentrations than ozone concentration. Furthermore, as the gap of the confined space decreased, the concentration of ozone was observed to decrease while that of nitrate increased, and the rate of this concentration change was further accelerated at higher temperature and higher power. It was shown that ozone concentration decreased from 0.11 µmol to 0.03 µmol and the nitrate concentration increased from 20.5 µmol to 24.5 µmol when the spacing in the confined space was reduced from 5 mm to 1 mm, the temperature of the external discharge was controlled at 40 °C, and the discharge power was 12 W. In summary, this work reveals the formation and transformation mechanisms of active substances in air surface micro-discharge plasma within confined space, providing foundational data for its medical applications.