The aims of this research were analysis of methanolic extract of bioactive compounds of Tribolium castaneum and evaluation of anti-bacterial activity against Streptococcus pneumonia, Pseudomonas eurogenosa, Staphylococcus epidermidis, Escherichia coli, Proteus mirabilis, Streptococcus pyogenes, Staphylococcus aureus, Streptococcus faecalis and Klebsiella pneumonia. GC-MS analysis of Tribolium castaneum revealed the existence of the Oxirane, [(hexadecyloxy)methyl], 5-Methyl-6-phenyltetrahydro-1,3-oxazine-2-thione, Cis,cis,cis-7,10,13-hexadecatrienal, Methyl N-cyclohexyl-3-phenylpropanimidate, Ethyl(1-adamantylamino)carbothioylcarbamate, (-)-Norephedine, 1-Dimethyl(pentafluorophenyl)silyloxycyclopentane, 2-Cyclopropyl-2-nitro-1-phenyl-ethanol, 3-Chloropropionic acid,nonyl ester, Decane, 1-chloro, 2,5-Dimethylhexane-2,5-dihydroperoxide, Ethanamine, 2-(2,6-dimethylphenoxy)-N-methyl, D-(+)-Ribonic acid?-lactone, Methanone, 1,3-dithian-2-ylphenyl, Cyclopentaneacetaldehyde, 2-formyl-3-methyl- ?-methylene, Phenylethyl alcohol, 2,5-dihydroxy-?-methyl, 1-Hexadecanol, 3,6-Diazahomoadamantan-9-one Hydrazone, 3-Pyridinecarboxylic acid, 1,6-dihydro-4-hydroxy-2-methyl-6-?, Spiro[5.5]undecane-1,7-dione, 4-Cyclopropylcarbonyloxytridecane, Eicosanoic acid, phenylmethyl ester, 3,6-Dinitro-4-cyclohexene-1,2-dicarboxylic acid, 1-Propyl-3,6-diazahomoadamantan-9-ol, 2(1H)-Benzocyclooctenone, decahydro-4a-methyl-,trans, Heptanoic acid, 4,5- dimethoxy-2-nitrobenzyl ester, Phthalic acid, butyl undecyl ester, Pterin-6-carboxylic acid, 2(3H)-Naphthalenone,4,4a,5,6,7,8-hexahydro-1-methoxy, Cis-9-hexadecenoic acid, 9,12-Octadecadienoic acid (Z,Z), Octadec-9-enoic acid, Methyl 18-fluoro-octadec-9-enoate, Dasycarpidan-1-methanol, acetate(ester), 3 ,8,8 -Trimethoxy-3-piperidyl-2,2 -binaphthalene-1,1 ,4,4 tetra. The results of anti-bacterial activity produced by Tribolium castaneum showed that the volatile compounds were highly effective to suppress the growth of Escherichia coli.