643 Acta Poloniae Pharmaceutica – Drug Research, Vol. 79 No. 5 pp. 643-659, 2022 DOI: 10.32383/appdr/157503 Received 3 August 2022, Received in revised form 13 November 2022, Accepted 15 December 2022 DRUG SYNTHESIS SYNTHESIS AND STRUCTURAL CHARACTERIZATION OF NOVEL 2-PYRAZOLINE DERIVATIVES: EVALUATION OF THEIR ANTIPROLIFERATIVE ACTIVITY AND FLUORESCENCE PROPERTIES ZEFINE UGRAS 1,2 , FATIH TOK 2 , EMINE SALVA 3 , GOZDE ULTAV 3 , and BEDIA KOCYIGIT-KAYMAKCIOGLU 2,* 1 Institute of Health Sciences, Marmara University, Dragos, 34865 Kartal, Istanbul 2 Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Marmara University, 34854 Maltepe, Istanbul 3 Department of Pharmaceutical Biotechnology, Faculty of Pharmacy, Inönü University, 44280 Battalgazi, Malatya Abstract: In this study, a series of novel 2-pyrazoline derivatives were synthesized and their structures were established by using spectral methods. The antiproliferative activities of compounds were investi- gated against human cell lines A-549 and MCF-7 by MTT assay and L-929 (mouse normal fbroblast) cell cytotoxicity was also examined. Apoptotic effects of the compounds in breast and lung cancer cells were assessed by Annexin V-FITC apoptosis assay using fow cytometry. The antiproliferative effect on lung carcinoma of the synthesized compounds was higher than breast carcinoma. Moreover, it was observed that none of the synthesized compounds have cytotoxic activity in healthy cells. Flow cytometry stud- ies have shown that compounds induced apoptosis at high concentrations. Additionally, fuorescence cell imaging studies were performed for the frst time in A-549 and MCF-7 cancer cell lines to determine the potential of the biosensor compounds by fuorescence microscopy. Compounds 4b, 4d, 4e, and 4f showed fuorescence properties by considering microscopic imaging. Keywords: synthesis, pyrazoline, structural characterization, cytotoxic, fuorescent imaging. * Corresponding author: e-mail: bkaymakcioglu@marmara.edu.tr Fluorescent probes are used in many felds such as genomics and proteogenomics, medical diagnoses like monitoring DNA and RNA abundance, local- ization and dynamics of nucleic acids, drug discov- ery, and microscopy (1, 2). Fluorescent probes are important molecular tools for analytical and opti- cal imaging due to their high sensitivity, specifc- ity, rapid response, and simple applicability (3). To date, countless synthetic fuorescent probes were designed, synthesized, and widely used in the rec- ognition of biomolecules (anions, enzymes, reactive oxygen, nıtrogen, and sulfur species (4). Some ex- ogenous fuorophores such as fuorescein and indo- cyanine green (ICG) which are approved FDA (Food and Drug Administration) are used to analyze and characterize tissues. Pyrazoline scaffold has a non-toxic bioactive property and is well known as a fuorescent probe due to its charge transfer character besides play- ing an important role in biological activities such as antibacterial (5), antidiabetic (6), antifungal (7), anti-infammatory (8), anticancer (9), anticonvulsant (10), antinociceptive (11), antioxidant (12) and anti- pyretic (13). Pyrazolines attract great attention due to their strong fuorescence properties, good mem- brane permeability, low toxicity, and simple synthe- sis methods (14-16). Intramolecular charge transfer (ICT) change is formed in the pyrazoline structure as a result of exci- tation with light. It occurs from the nitrogen atom in the 1st position to the nitrogen atom in the 2nd posi- tion and the carbon atom in the 3rd position, which can cause a conformational change in the molecular structure. Molecular planarity is required for fuo- rescence activity (17). It is known that the pyrazoline ring, which car- ries at least two aryl substituents in the 1st and 3rd positions, exhibits a strong fuorescence activity. With the formation of the pyrazoline ring, the ring has a planar structure with the effect of carbon at- oms in the 4th and 5th positions, and the aromatic rings in the 1st and 3rd positions interact through ISSN 2353-5288 Polish Pharmaceutical Society