(M1430-11-63) Going Greener with the USP 621 Globally Harmonized Pharmacopeial Guidance for Liquid Chromatography

Purpose: The U.S. Pharmacopeia (USP) General Chapter < 621 > Chromatography was recently updated. The USP standard text is making efforts to harmonize regulatory texts utilized around the world (i.e. USP, EuPh, JP, ChP).  In this poster we demonstrate how validated gradient LC methods can be modernized to become more efficient, greener methods using the recent chapter updates. This modernization results in reduced hazardous waste generation, while maintaining the LC method's original validation.

Methods: The validated, USP monograph for abacavir sulfate was selected as an example method for modernization. Column hardware was updated to modern dimensions, and the separation method was subsequently, mathematically adjusted according to the formulas and limits specified in USP Chapter 621. The final gradient separation methods were evaluated for reduction in solvent usage and run time. The original monograph system suitability mixture (SST) and criteria were used to investigate the integrity of the original monograph method validation, after hardware and method modernization.

Results: For all modern column dimensions, the original monograph SST resolution requirement was successfully achieved. Although the relative retention time of peaks changed per column chemistry, the resolution of the critical pair passed the original monograph suitability requirements. We did not observe peak deletions or inversions when changing column substituent chemistries within the boundaries of the USP guidance. Greener LC gradient methods i.e. less waste generation, was achieved within the HPLC platform, while significant reductions were noted when methods were moved from the HPLC platform to a UHPLC platform.

Conclusion: The data presented demonstrates that the new USP adjustment guidance, successfully promoted the employment of modern column dimensions and innovative stationary phase substituents. These parameters resulted in significantly less waste generation when high mobile phase volume legacy methods were converted to methods utilizing modern separation hardware. This approach demonstrates the ability to go greener while maintaining the validation of an original method.

References: 1. Achieving Method Modernization with the New Liquid Chromatographic Gradient Allowances Provided by USP General Chapter Chromatography and the Alliance™ iS HPLC System, Catharine E. Layton, Paul D. Rainville, www.waters.com, 720007865, February 2023.
2. General Chapter Chromatography, Official Date: 01-Dec-2022, www.USP.org, referenced 1/7/2023.
3. Abacavir Sulfate Monograph official 01-May-2020, www.uspnf.com, referenced 4/15/2022
4. Preparative OBD Column Calculator, www.waters.com, accessed 1/15/2023.
5. Columns Calculator 2.0, www.waters.com, accessed 1/15/2023


Formula (A) Flow rate (B) injection volume (C) gradient time, and (D) dwell volume offset (FIO) and (E) final gradient table using the dwell volume offset is demonstrated for adjusting the gradient from the monograph, 3.9 x 150 mm, 5 µm HPLC platform with dwell volume 1.145 mL, to a 2.1 x 75 mm, 2.5 µm UHPLC separation platform with dwell volume 0.073 mL.


This graphic shows a comparison of the retention times when modernizing HPLC and UPLC platforms using USP chromatographic gradient method flexibility.


This graphic summaries the resulting savings (i.e. mobile phase solvent, time) when moving validated methods from legacy hardware to modern hardware.