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A method for the determination of amizone and amixin in the same sample and in their mixture with antibiotics (ceftriaxone, tetracycline, ampicillin and levofloxacin) was developed using reversed-phase highperformance liquid chromatography equipped with a photodiode array detector. A SunFire C18 column, a mobile phase consisting of sodium perchlorate buffer (pH = 2.5) and acetonitrile (75:25), at a flow rate of 0.8 ml/min, was used. The analytes were identified at 205 and 265 nm. The specificity, linearity, precision parameters, LOD and LOQ were evaluated during the validation of the methodology, and the correlation coefficients of amizone and amixin were 0.9992 and 0.9998, respectively. This method can also be used for the determination of amizone and amixin and their presence in a mixture with antibiotics in the environment.

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