AbstractCu (II), Ni (II), Co (II), and Mn (II) acetato in addition to Cu (II) chloro and nitrato mixed ligand complexes of 4‐([quinolin‐2‐yl]methyleneamino)‐1,2‐dihydro‐2,3‐dimethyl‐1‐phenylpyrazol‐5‐one (L), as a primary ligand and 2,4‐dihydroxy benzaldehyde (H2L), as a secondary ligand have been isolated. The structural features and geometrical arrangements of the synthesized mixed ligand complexes were identified by elemental analysis, Fourier transform infrared spectroscopy (FT‐IR), conductivity measurements, UV‐Vis spectra, magnetic moment measurements, X‐ray diffraction, and electron spin resonance (ESR) spectral and thermogravimetric techniques studies of the mixed ligand complexes confirming the molar ratio 1:1:1 (primary ligand: M: secondary ligand) for Cu (II), Co (II), Mn (II), and UO2(II) chelates. The spectral techniques also confirmed the metal chelates have the formulae (Cu[H2L][L][X]2), where X = Cl−(1); NO3−(2); and (M[L][HL][OAc])YH2O, where M = Cu (II) (3), Co (II) (4); Y = 1, Mn (II) (5); Y = 1 and UO2(II) (6); Y = 2. The kinetic and thermodynamics parameters of the mixed ligand complexes at different decomposition steps were calculated using Coast‐Redfern and Horowiz‐Metzger methods. Also, the antimicrobial activities of ligands and their mixed ligand complexes were screened by Disc Diffusion method. It was found that all mixed ligand complexes (1–6) have high antibacterial activity againstKlebsiella pneumoniaethan penicillin G. By studying the molecular docking between the quinoline‐2‐carbaldehyde, the ligand and the receptors (Escherichia coli(PDB code: 3t88),Staphylococcus aureus(PDB code: 3q8u), lung cancer (PDB code: 1x2j), and colon cancer (PDB code: 2hq6)), it was found that the ligand is efficient compound for all the receptors than the quinoline‐2‐carbaldehyde. The DNA cleavage properties of the two ligands and their complexes have been investigated by gel electrophoresis and the results showed that complex (1) without H2O2and complex (6) with H2O2exhibited a significant partial oxidative cleavage among other complexes.