A novel hydrophilic interaction capillary electrochromatographic (HI-CEC) monolith with covalently bonded zwitterionic functional groups was applied for the separation of riboflavins (RF) and its derivatives. With a homemade pressurized capillary electrochromatography-laser induced fluorescence detection (pCEC-LIF) system, trace levels of RF and its derivatives, flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD), can be baseline separated within 8.0 min in isocratic elution mode. The effect of experimental parameters on separation was investigated. Under the optimum conditions, analytes could be determined over nearly three orders of magnitudes with the detection limits (LODs) as low as 5.0 x 10(-11) mol/L (RF), 8.0 x 10(-10) mol/L (FMN), 2.5 x 10(-9) mol/L (FAD), and the relative standard deviations (RSDs) were less than 8.2%. This method is rapid, simple, repeatable and more sensitive than the most of reported methods, and satisfied results has been achieved in serum sample. Furthermore, it can be further applied for trace analysis of RF and its derivatives in biological fluid and cells.