Improved Chlorhexidine Carryover Performance Using the Alliance iS HPLC System

Importance of Carryover Control in HPLC

Carryover in HPLC analyses can lead to inaccurate quantitation and regulatory failures. Understanding and mitigating both volumetric and adsorptive carryover is essential when transferring methods across different instrument platforms in pharmaceutical and biopharmaceutical laboratories.

Objectives and Study Overview

This study used a scaled USP monograph for chlorhexidine hydrochloride impurities to assess carryover performance on multiple HPLC systems. The aim was to compare default and optimized needle wash strategies and evaluate the Alliance iS HPLC System against competing platforms.

Methodology and Instrumentation Used

The method was scaled from a 4.6×250 mm to a 3×100 mm column (XSelect HSS C18, 2.5 µm) and run at 0.6 mL/min with a 19 min gradient. Key instrumentation included:

• Alliance iS HPLC System
• Vendor X HPLC Systems (Systems 1 & 2)
• Vendor Y HPLC/UHPLC System (System 3)
• Vendor Z HPLC System (System 4)
• UV detection
• Autosampler with flow-through needle design

Sample sequence comprised pre-blanks, standard injections (11.3 µg/mL), challenge injections (1.13 mg/mL), and post-blanks. Carryover was calculated as the ratio of carryover peak area in the first blank to the preceding challenge injection area.

Main Results and Discussion

Default “No Wash” settings on some systems yielded significant carryover, while systems with default needle washes showed improved performance. The Alliance iS System, using tool-free zero-dead-volume fittings and a 1000 µL two-stage needle wash, achieved the lowest carryover at 0.00055%. Adding or extending washes on other systems reduced carryover but often increased cycle time by 30–214 s. In contrast, the Alliance iS design delivered superior carryover control without a significant cycle time penalty.

Benefits and Practical Applications

• Reliable method transfer across instruments
• Minimal volumetric carryover using foolproof fittings
• Efficient adsorptive carryover mitigation via optimized needle wash

Future Trends and Potential Uses

Advances in low-dead-volume fittings and integrated wash mechanisms will further reduce carryover without compromising throughput. Adoption of automated optimization routines for wash parameters may streamline method validation and regulatory compliance.

Conclusion

Effective carryover control requires both robust instrument design and appropriate needle wash strategies. The Alliance iS HPLC System demonstrated exceptional performance in minimizing both volumetric and adsorptive carryover, simplifying method transfer and ensuring analytical accuracy.

References

• USP Monograph Chlorhexidine Hydrochloride. USP–NF. May 1 2022.