The complex Cu(II) with thiosemicarbazone of glyoxylic acid was evaluated for biological activity against various microorganisms such as antibacterial Pseudomonas aeruginosa and Mycobacterium phlei and antifungal Aspergillus niger, Penicillium Chrysogenium, Cladosporium resinae. Some of the assayed compounds showed interesting activity against microbes and fungicides. Meat-paste agar (MPA) was used for growing bacterial crops, and wort-agar (CA) was used for mushrooms. These studies showed that the ligand does not show an antibacterial effect but shows an antifungal effect, and the copper complex with this ligand shows both antibacterial and antifungal effects. The antimicrobial action of the complex is much higher than the ligand itself and this action increases with increasing concentration of the complex compound in solution. The complex characterized DFT calculations and Hirsfeld analysis. DFT/TD-DFT calculations were performed to theoretically determine the structural, spectroscopic and crystallographic properties of the synthesized complex. Hirshfeld analysis was performed to determine the interactions between molecular crystals and to display the details. The results have been compared with the corresponding theoretical results