The invention provides a method for rapid determination of pharmacokinetics of active ingredients after oral administration of a traditional Chinese medicine rhizoma anemarrhenae; the means of combining microdialysis sampling with ultra performance liquid-triple quadrupole mass spectrometry is specifically adopted, and the pharmacokinetics of the active ingredients comprising neomangiferin, mangiferin and timosaponin B II after a model animal administrates orally a rhizoma anemarrhenae extract are determined. An experiment adopts a reverse dialysis method, a probe is perfused with a perfusion liquid citric acid dextrose solution at a flow rate of 1.0 [mu]l*min<-1>, and samples are collected; chromatographic detection adopts acetonitrile and 0.1% formic acid water for gradient elution. The biological sample analysis requirements of the 'Chinese pharmacopoeia' and the 'Chemical Drugs Non-Clinical Pharmacokinetics Technical Guidance Principle' are met in the aspects of accuracy, precision, specificity, stability, extraction recovery rate, matrix effect and the like. At the same time, continuous sampling of a rat living body after a minimally invasive surgery is achieved, the obtained samples are avoided from being interfered by proteins, enzymes and other biological macromolecular substances, precision instruments are directly used for sample injection analysis, the recovery rate is improved, and the method is especially suitable for analysis of trace active substances of biological samples.