## 3 Results and Discussion  
### Synthesis and Characterization  
The initial goal of this work was the synthesis of novel unsymmetrical squaraine cyanine dyes that exhibit properties inherent to those of an ideal photosensitizer molecule, aiming at the discovery of new photosensitizing potential molecules to be applied in cancer PDT. The design of the molecules intended to be synthesized was based on the photophysical and photochemical studies already described in the literature, which reported that quinoline-based squaraines displayed higher wavelength absorption capacity than their benzothiazole- and benzoselenzazole-based analogs, a photophysical property considered advantageous since, it allows greater therapeutic efficacy, as these molecules may be distributed to deeper tissue zones as they will be able to absorb and be stimulated by radiation of higher wavelength [5; 53]. On the other hand, the introduction of heavy atoms into the dye structure, such as iodine, chlorine, or bromine, is widely described to improve its performance relative to its singlet oxygen production ability [54; 55]. The choice of these unsymmetrical dyes also being derived from benzothiazole was due to the fact that this heterocyclic correlates with notable phototherapeutic activity [56; 57]. Given that increasing length of the pendant \(N\)-alkyl chains positively influences photophysical properties such as singlet oxygen production ability and maximum absorption wavelength, \(N\)-hexyl chains have been introduced in both heterocyclic dye moieties [8; 58]. Thus, unsymmetrical squaraine cyanine dyes **10-13** bearing 6-iodoquinoline and benzothiazole moieties were successfully prepared by a multistep procedure, as illustrated in Scheme 1. The dyes preparation was initiated by the quaternization of the commercially 2-methylbenzothazole **1**, through the alkylation of the nitrogen atom with iodohexane, resulting in the synthesis of the quaternary ammonium salt **2**[45]. Then, dibutyl squarate (**4**), obtained by the reaction between squaric acid (**3**) and \(n\)-butanol, reacted with the benzothiazole-based quaternary ammonium salt **2** to afford the monosubstituted intermediate **5**[46; 47]. This latter intermediate was posteriorly hydrolyzed using a 40% sodium hydroxide aqueous solution, and the resulting sodium salt was protonated with a 2 M hydrochloric acid solution, yielding the key monosubstituted intermediate **6**[47]. The zwitterionic unsymmetrical squaraine dye **10** was prepared by a base-catalyzed condensation of intermediate **6** with the iodoquinoline-based quaternary ammonium salt **9**, in a n-butanol/pyridine refluxing mixture. Ammonium salt **9** was prepared by the alkylation of 6-iodo-1-hexylquinoline (**8**) with iodohexane [49], obtained by the described Doebner-Miller reaction between 4-iodoaniline (**7**) and crotonaldehyde in hydrochloric acid [48; 59]. Methylation of the unsubstituted squaraine dye **10** with methyl trifluoromethanesulfonate produced the \(O\)-methyl ether intermediate **11**[4]. Finally, the aminosquaraine cyanine dyes **12** and **13** were obtained by nucleophilic substitution of the methoxy group of the \(O\)-methyl derivative **11** by ammonia and methylamine solutions, respectively [4; 25]. As far as we know, all the synthesized squaraine cyanine dyes are not described in the literature, and their synthesis and spectroscopic characterization are herein presented for the first time. The main features in the \({}^{1}\)H-NMR spectra of dyes **10-13** (Figures S1, S3 and S5) are the signals corresponding to the methine-protons. For **10**, **11** and **12**, the protons above referred appear at \(\delta\) 5.89 and 5.73, 6.08 and 5.55, 6.10 and 5.86 ppm, respectively, as two singlets since these dyes have two different heterocyclic rings.