With the facile condensed carbonyl gourp of 2-hydorxybenzaldehyde with the amino group of quinoxalyl-2-carbohydrazide, a new chelating quinoxalyl hydrazone derivative ligand (H2Lpx) was constructed. Its coordination behavior towards three divalent Co2+, Ni2+, and Cu2+ ions was discovered by building up three new metallo-organic complexes (CoLpx, NiLpx, and CuLpx, respectively). Confirmation of their chemical structure was formulated by applying different spectroscopic methods, as well as, the micro-elemental (C,H,N) and thermogravimetric analyses, the conductance, and magnetic evaluations. The inhibition behavior of the free ligand and its metallo-organic complexes against the cell growth of six entitled microorganisms (three fungal and three bacterial types) and three tumors of humans was estimated considering the effective role of the divalent metal ions (Co2+, Ni2+, and Cu2+ ions in their metallo-organic complexes). The interacting nature of H2Lpx, CoLpx, NiLpx, and CuLpx, with calf thymus DNA (ctDNA) was manifested within the viscometric, spectrophotometric titrations and gel electrophoresis.The three complexes (CoLpx, NiLpx, and CuLpx) exposed a respectable inhibition attitude more than that of H2Lpx depending on the inhibition zone areas of the microbial series growth in mm and the half-inhibition concentration in µM (IC50) for the tumor cell lines’ growth. Establishing of the interacted modes between H2Lpx and MLpx with ctDNA was built upon the shifts of the spectral scans for their solutions. In addition, chromism type, Gibb’s free energy, and binding constant values (ΔGb≠ and Kb, respectively) helped to distinguish their interacted modes, which supported DNA damage. The bio-action of MLpx complexes was highly modified compared to that of H2Lpx towards ctDNA.