Application of an in-house packed octadecylsilica-functionalized graphene oxide column for capillary liquid chromatography analysis of hormones in urine samples.

Graphene oxide-based LC stationary phases were developed and applied for separating hormones from urine using capillaryLC-MS/MS. Using two analytical approaches - direct injection and column-switching arrangement - it was possible to evaluate the chromatographic parameters and perform tests on the raw biological fluid. Two stationary phases (SPs) were produced, varying the amino silica support particle diameter (Si, 5, and 10 μm). Graphene oxide was covalently bonded to the surface of Si particles, and this material was functionalized by the insertion of octadecylsilica groups, generating the SiGO-C18. Infra-red spectroscopy assays revealed that both steps were successful - supporting GO onto Si and further C18 customization. Scanning electron microscopy showed spherical geometries with minor irregularities and narrow particle size distribution for the produced SPs. The GO-coating rate was higher on the Si particles of 10 μm. As a result, the 10 μm produced column reported better resolution, efficiency, and peak capacity. Therefore, this SiGO-C18 capillary column (100 mm × 0.32 mm i.d., 10 μm dp) was applied successfully in a column-switching method to separate hormones in urine. Linearity (R2 above 0.99), quantification limits (between 1.0 and 5 μg/L), and other figures of merit of the method were determined. It is worth mentioning that the SiGO-C18 capillaryLC column performed adequately, separating the target compounds in less than 6 min. We hope this work could significantly contribute to shedding some light on graphene-based materials as a promising class of stationary phase for miniaturized liquid chromatography.