Abstract: Acetone is one of the highest levels of volatile organic compounds (VOCs) in the gas of human body exhaled breath. Researches show that acetone from human exhaled breath has a certain relationship with a series of diseases, such as diabetes, lung cancer and so on. Acetone mixed gas with different carbon dioxide concentration and relative humidity was prepared by syringe method in Teldar bags. Then proton transfer reaction mass spectrometry (PTR-MS) was used to investigate acetone mixed gas in Teldar bags in order to obtain the acetone concentrations. The experiment aimed at studying the influence of the carbon dioxide concentration, the relative humidity, oxygen ion concentration and nitric oxide ion concentration to acetone concentration measurement. Also the concentration correction formula was deduced. The results show that the increases of carbon dioxide concentration and relative humidity of the mixed gas have no obvious influence on the count of acetone ions, but they can increase concentration ratio of protonated water clusters to protonated water in the drift tube and increase the possibility of acetone molecule and water cluster ions reaction. Oxygen ions will react with acetone molecules, so the increase of oxygen ions concentration can enhance the content of acetone fragment ions, which influence the measurement accuracy of breath acetone by PTR-MS. The measuring concentration of nitric oxide ions is so low that the influence on the result can be negligible. The concentration correction formula can effectively inhibit the effect of carbon dioxide, relative humidity and oxygen ions.