Within this work we have investigated spiro-based tetrathiafulvalenes (TTFs) obtained as mixtures of stereoisomers from racemic spiro[5.5]undeca-1,8-dien-3-one. Compared to previously described spiro-TTFs, enantiomeric and diastereoisomeric forms have been here separated by chiral HPLC and fully characterized both experimentally and theoretically. The two types of spiro-based chiral derivatives contain either one (2) or three (1) chiral centres out of each one is spiro-type. Experimental CD, supported by TD-DFT calculations, shows differences in the optical activity between the 1 and 2 and their intermediates. The low optical activity of 2 and 3 (spiro alone chirality) was attributed to the presence of two conformers in the solution (ax and eq) of opposite Cotton effect whereas in the case of 1 and 5 (spiro and stereogenic centres) the spiro chirality seems to be responsible of the Cotton effect in the high energy region whereas the R and S chirality in the low energy region. Racemic and enantiopure forms have been successfully used for the synthesis of charge transfer complexes with tetracyanoquinodimethane (TCNQ) based acceptors.