The spread of tetracyclines through agricultural systems is causing the present bacteria to develop antibiotic resistance. The spread of this bacteria, as well as the tetracycline antibiotics in the environment is dangerous because these antibiotics pose health hazards for humans. The overuse of antibiotics, which are added to livestock feed, results in the antibiotics being released into the environment via animal feces. In this research, we have attempted to design an analytical method to isolate antibiotics from agricultural wastes with subsequent detection using liquid chromatography and mass spectrometry (LC-MS). The antibiotics investigated in this study were tetracycline, chlortetracycline, and oxytetracycline. The analytical procedure involves mixing the agricultural samples with an organic solvent, such as methanol, which solubilizes these antibiotics. Next, samples are centrifuged to remove solid particulates. A polymeric weak cation cartridge was used to concentrate and separate the antibiotics from the unwanted organic chemical compounds found in the samples. The antibiotics were released with methanol with small amounts of acid and then detected and quantified using LC-MS and high performance liquid chromatography with a photodiode array detector (HPLC-PDA).