Synthesis of drugs, biologically active compounds or their derivatives always requires precise and reliable method of their identification, including differentiation of the possible isomers. Pseudodistamines and their precursors became a matter of elevated attention due to their different enzymatic inhibition. This paper deals with one of the groups of the pseudodistamine precursors − trans-3(4)-aminopiperidin-4(3)-ols. Their synthesis brings to a mixture of 2 regioisomers, resulting in the necessity of their reliable recognition. NMR spectroscopy commonly used by organic chemists requires advance knowledge and experience to analyse the spectra of these regioisomers. Therefore, we herein proposed a simpler way to recognize trans-3(4)-aminopiperidin-4(3)-ols using mass spectrometry with electron ionization. Fragmentation of 4 pairs of aminopiperidinol regioisomers with variation of amine moiety was studied. The obtained results allowed defining a group of 3 ions ([M-18]+., [M-19]+, [M-43]+) related only to the structure of trans-4-aminopiperidin-3-ols and 1 ion (m/z 100) related to the structure of trans-3-aminopiperidin-4-ols. Besides, interrogation of intensity of ions common for spectra of both regioisomers allows making differentiation as well.