A two-dimensional liquid chromatography method with optimized pretreatment for detection of guanidine compounds in animal liver and kidney.

This study is centered on optimizing the derivatization reaction conditions and the protein precipitation reagent system for the detection of guanidine compounds (GCs) in animal liver and kidney. It introduces a two-dimensional liquid chromatography combined with ultraviolet detection (2D-LC-UV) method. The optimal derivatization conditions were identified through an orthogonal experiment, with the most favorable reaction conditions being 100 °C for 5 min, and optimal concentrations of benzoin and potassium hydroxide (KOH) determined to be 30 mmol/L and 8 mol/L, respectively. Furthermore, a 50 % methanol-0.5 % hydrochloric acid solution was selected as an effective protein precipitation reagent. Methodological validation demonstrated that, in mouse liver and kidney tissues, the linear range for GCs was 5-500 μmol/L (r2 > 0.99), with a limit of quantification of 5 μmol/L. Analysis of animal samples revealed an organ-specific metabolic distribution of GCs. This study furnishes reliable technical support for the detection of guanidino compounds in animal biological samples.