The ionization mechanism in the novel atmospheric pressure photoionization mass spectrometry (APPI-MS) in negative ion mode was studied thoroughly by the analysis of seven compounds in 17 solvent systems. The compounds possessed either gas-phase acidity or positive electron affinity, whereas the solvent systems had different polarities and gas-phase acidities and some of them positive electron affinities. The analytes that possessed gas-phase acidity formed deprotonated ions in proton transfer; in addition, fragments and solvent adducts were observed. The compounds of positive electron affinity formed negative molecular ions by electron capture or charge exchange and substitution products of formϪ by substitution reactions. The efficiency of deprotonation was decreased if the solvent used possessed higher gas-phase acidity than the analyte. Solvents of positive electron affinity captured thermal electrons and deteriorated the ionization of all the analytes. Also, the proportion of substitution products was affected by the solvent. Finally, the performances of negative ion APPI and negative ion APCI were compared. The sensitivity for the studied compounds was better in APPI, but the formation of substitution products was lower in APCI. ( and aromatic imines and amines [8].The ionization process in dopant assisted APPI in positive ion mode is initiated by the photoionization of a dopant (e.g., toluene) and formation of a dopant radical cation [1,7,9]. Next, the dopant radical cation can ionize the solvent molecule by proton transfer, if the proton affinity (PA) of the solvent molecule is higher than that of the deprotonated radical cation. Protonated solvent molecules can donate a proton to the analyte molecule, in case the PA of the analyte is higher than that of the solvent molecule. Alternatively, the dopant radical cation can ionize the analyte directly by charge exchange, if the ionization energy of the analyte is lower than that of the radical cation. Thus, two routes of ionization are possible in positive ion APPI-charge exchange and proton transfer-the route depends on the ionization energy and proton affinity of the analyte and the solvent. Charge exchange makes ionization of non-polar compounds possible, which may not be possible by using electrospray or APCI.Preliminary results on negative ion APPI indicated that negative ions can be formed by electron capture, charge exchange, proton transfer, or substitution reactions [7]. The aim of this work is to study in detail the ionization mechanism and solvent effect in negative ion APPI. Thus, seven analytes that possess gas-phase acidity or positive electron affinity (EA) were chosen to enable different ionization mechanisms in negative ion APPI. The compounds were analyzed in 17 solvents that have different polarities and gas-phase acidities; in addition two of them have positive EAs. Also, the effect of operational parameters on formation of substitution products was studied. Finally, a comparison to negative ion APCI was made.