Non-invasive physical plasma (NIPP) achieves biomedical effects primarily through the formation of reactive oxygen and nitrogen species. In clinical use, these species interact with cells of the treated tissue, affecting the cytoplasmic membrane first. The present study investigated the permeability of the cytoplasmic membrane of breast cancer cells with different fluorescent dyes after NIPP treatment and determined the subsequent effects on cell viability. After NIPP treatment and the associated formation of reactive oxygen species, low molecular weight compounds were able to pass through the cytoplasmic membrane in both directions to a higher extent. Consequently, a loss of cellular ATP into the extracellular space was induced. Due to these limitations in cell physiology, apoptosis was induced in the cancer cells and the entire cell population exhibited decreased cell growth. It can be concluded that NIPP treatment disturbs the biochemical functionality of the cytoplasmic membrane of cancer cells, which massively impairs their viability. This observation opens a vast application horizon of NIPP therapy to treat precancerous and malignant diseases beyond breast cancer therapy.