Analysis of Amikacin and Kanamycin in Pharmaceutical Products with 3465 Electrochemical Detector

Importance of the Topic

Amikacin and kanamycin are vital aminoglycoside antibiotics widely used in clinical settings to treat severe bacterial infections and tuberculosis. Accurate quantification of these compounds in pharmaceutical formulations is essential to ensure drug safety, efficacy, and regulatory compliance. High Performance Anion Exchange Chromatography with Pulsed Amperometric Detection (HPAEC-PAD) is recognized by the United States Pharmacopeia (USP) for its sensitivity and selectivity in analyzing aminoglycosides under high-pH conditions.

Objectives and Study Overview

This study aimed to validate an HPAEC-PAD method for simultaneous determination of amikacin (AMI) and kanamycin (KAN) in injection products following USP monograph guidelines. The performance of an ACQUITY™ Arc™ System equipped with a 3465 Electrochemical Detector (ECD) and controlled by Empower™ 3 CDS was evaluated for system suitability, linearity, and assay precision.

Methodology and Instrumentation

The analysis employed a Hamilton® RCX-10 column (250×4.6 mm, 7 µm) maintained at 30 °C. Mobile phases consisted of freshly prepared 115 mM NaOH for KAN and 134 mM NaOH for AMI. A 10 µL sample injection, delivered at 0.5 mL/min, achieved baseline separation within 30 minutes. Pulsed amperometric detection used a FlexCell™ with gold working electrode, HyREF reference, and stainless-steel auxiliary electrode under a multi-step waveform. System maintenance included PEEK tubing, loop offline purge, and daily water flushes to handle high-pH mobile phases.

Main Results and Discussion

System suitability tests for both analytes met all USP criteria for resolution (>1.5), tailing factor (<2.0), and repeatability (<2% RSD). Calibration over six concentration levels (80–140% of target) produced correlation coefficients ≥0.997, confirming linear response through the origin. Assay of commercial amikacin injections yielded recoveries within 95–105% of label claims and RSD <1.5%, demonstrating method accuracy and precision.

Benefits and Practical Applications

• Meets USP monograph requirements for AMI and KAN assay in quality control labs.
• High sensitivity and selectivity for aminoglycosides without derivatization.
• Robust performance under high-pH conditions with minimal maintenance.

Future Trends and Opportunities

Advancements may include coupling HPAEC-PAD with high-resolution mass spectrometry for structural confirmation of impurities. Miniaturized systems and automated sample preparation could reduce solvent use and analysis time. Broader adoption in regulatory labs will drive refinement of waveforms and reference electrodes for enhanced throughput and lower detection limits.

Conclusion

The ACQUITY Arc System with 3465 ECD under Empower 3 control provides a reliable, USP-compliant HPAEC-PAD method for quantifying amikacin and kanamycin in pharmaceutical injections. The approach ensures robust system suitability, linearity, and precision, supporting its routine use in pharmaceutical quality control.

Reference

1. United States Pharmacopeia. Kanamycin Injection, USP43-NF38, Rockville, MD, 2022.
2. United States Pharmacopeia. Kanamycin Sulfate, USP43-NF38, Rockville, MD, 2022.
3. United States Pharmacopeia. Amikacin, USP43-NF38, Rockville, MD, 2022.
4. United States Pharmacopeia. Amikacin Sulfate, USP43-NF38, Rockville, MD, 2022.
5. United States Pharmacopeia. Amikacin Sulfate Injection, USP43-NF38, Rockville, MD, 2022.