EAS: Impact of Instrument Design on Absorptive Carryover

Significance of the Topic

Carryover in HPLC can compromise quantitation and sensitivity due to analyte adsorption on flow path surfaces. Effective control of absorptive carryover is critical in pharmaceutical analysis and QA/QC to ensure data integrity and regulatory compliance.

Study Objectives and Overview

This study evaluates how autosampler injector design and needle washing protocols influence absorptive carryover across five HPLC systems. The goal is to compare default and optimized wash routines to identify mechanisms that minimize sample-to-sample contamination.

Methodology

The method, adapted from the USP monograph for chlorhexidine impurities, used an XSelect HSS C18 SB column (250 × 4.6 mm, 3.5 µm) at 30 °C, with sample cooling at 8 °C and a 10 µL injection volume. Detection was at 254 nm using a TUV detector. A binary gradient of 0.1% TFA in water and acetonitrile was employed. Needle wash solvent was 50:50 acetonitrile:water; seal wash used 10:90 acetonitrile:water.

Used Instrumentation

• Arc HPLC SystemTM
• System V
• System Y
• System Z

Results and Discussion

Among the systems evaluated, lack of a needle wash (System Z) resulted in the highest carryover. Systems incorporating needle wash steps before or after aspiration/injection (System V, System Y, Arc HPLC SystemTM) demonstrated markedly reduced carryover. System Y’s dual wash by dipping the needle and Arc HPLC SystemTM’s post-injection wash provided the most effective removal of residual chlorhexidine. The study highlights wash timing, duration, and mechanism as critical factors governing carryover reduction.

Benefits and Practical Applications

• Enhanced data accuracy in quantitative HPLC assays by mitigating cross-sample contamination.
• Improved sensitivity for trace-level analyses through optimized wash protocols.
• Guidance for method development and validation in regulated environments.

Future Trends and Opportunities

Emerging autosampler designs may incorporate programmable multi-solvent wash sequences, ultrasonic needle cleaning, or disposable injection pathways to further reduce adsorptive carryover. Integration of real-time monitoring of carryover and adaptive wash cycles offers potential for automated contamination control in high-throughput laboratories.

Conclusion

Effective needle washing is essential to control absorptive carryover in HPLC. Injector design and wash parameters significantly influence carryover levels. Implementation of targeted wash steps tailored to the analyte and instrument design ensures reliable quantitative results.

Reference

1. Dlugasch A, Simeone J, McConville P. Alliance Carryover Performance Part 1: Carryover Improvement Achieved Through Instrument Design Changes for the Alliance HPLC System. Waters Application Note 720006386EN, 2018.
2. USP. Chlorhexidine Hydrochloride. United States Pharmacopeia and National Formulary (USP43-NF38), 2022.