This article introduces the techniques for isolating metabolites from biological samples using liquid chromatography with sub-2-micron particle columns. The method improves the rate and resolution of metabolite separation without compromising the overall purity or recovery of the metabolites.
The document presents an application note detailing a method to increase the rate of drug metabolite isolation from complex biological matrices like plasma and urine. It focuses on maintaining purity and recovery while reducing analysis time.
The study utilizes the Agilent Rapid Resolution LC (RRLC) system equipped with ZORBAX SB-C18 columns featuring 1.8-micron and 3.5-micron particles. Results demonstrate that switching from conventional 5-micron columns to these smaller particle sizes significantly reduces run times while improving resolution, peak shape, and UV response.
A flow rate of approximately 1 mL/minute is ideal to obtain fractions of a practical volume.
The system back pressure was 464 bar for the 1.5 mL/min analysis.
The operating range of the Agilent 1200 RRLC is 600 bar.