A UPLC Method for Simultaneous Estimation of Trastuzumab and Hyaluronidase in Bulk and Pharmaceutical Dosage Form

Abstract

Objective: The proposed research aimed at establishing a quick, accurate, and exact ultra-performance liquid chromatography (UPLC) methodology for contemporaneously analyzing trastuzumab and hyaluronidas in bulk and pharmaceutical formulation, with a focus on the stability-indicating properties of the assay. Method: Using a Water X-Bridge C18 column (50 mm x 4.6 mm x 2.5 μ) and a mobile phase of ACN: Buffer (pH 2.5) [50% v/v], pushed at 0.5 mL/min, compounds were separated chromatographically. An ultraviolet (UV) detector fixed at 260 nm was employed to identify the isolated compounds. Results: The findings showed that 1.3529 and 2.404 minutes were optimal for separating Trastuzumab and Hyaluronidase, correspondingly. The current procedure has been verified in accordance with ICH standards Q2 R1, and stability-indicating tests have been performed in accordance with ICH standards Q1A R2. Both the intra- and inter-day precisions were determined to be satisfactory. The suggested approach showed linearity between 30 to 180 μg/mL of trastuzumab and between 12.50-75μg/ mL of hyaluronidase. It was determined that the limit of detection (LoD) and limit of quantitation (LoQ) for trastuzumab were 0.36 and 1.2 μg/mL, correspondingly, whereas those for hyaluronidase were 0.15 and 0.5 μg/mL. The approach was shown to have a recovery rate around 98.6 to 99.7%. Conclusion: The suggested approach successfully separated the chemical compounds from their by-products. Therefore, trastuzumab and hyaluronidase routine evaluation and stability-indicating tests were both effectively implemented using the approach.