By utilising RP-HPLC with UV detection, a straightforward, sensitive, and precise analytical has been established to estimate amphotericin in pharmaceutical effluents that the pharmaceutical industry are discharging into aquatic environments. The devised approach is extremely accurate and sensitive to detect amphotericin at concentrations of less than 0.1ppm-5ppm. For the measurement of Amphotericin in effluents or pharmaceutical industry washouts, a reversed-phase high performance liquid chromatography (RP-HPLC) method was created and validated. On a Symmetry C-18 column (250mm 4.6mm i.d., 5.0m), the separation was accomplished using a citrate buffer with a pH of 4.5 as the buffer, and a combination of 400mL buffer and 600mL acetonitrile in isocratic mode as the mobile phase, all at a flow rate of 1.2 mL/min. A UV detector operating at 383 nm was used for detection. Amphotericin eluted at a retention period of approximately 5.0 min, taking around 12.0 min of total chromatographic analysis time per sample. For accuracy, precision, specificity, linearity, and sensitivity, the procedure was validated. Validation tests proved the accuracy, specificity, speed, dependability, and reproducibility of this HPLC method. The limit of detection (LOD) and limit of quantitation (LOQ) for amphotericin were discovered to be 0.0100 g/ml and 0.0500 g/ml, respectively, and the method was validated in accordance with ICH guidelines. Linearity was observed for amphotericin in the concentration range of 0.05-10 g/mL (R2>0.95). It was discovered that the RSD for intra-day and inter-day precision was less than 5%. The approach is straightforward, definite, exact, and accurate for determining amphotericin in pharmaceutical industry washouts, and the percentage recovery was in good agreement.
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