Two-dimensional liquid chromatography based on different modulations for the determination of Aristolochic Acid Ⅰ in Asari Radix et Rhizoma.

Asari Radix et Rhizoma (ARR) is a traditional Chinese herbal medicine derived from the genus Asarum in the Aristolochiaceae family, which has been widely used for years. Aristolochic acids in it significantly restrict the clinical application of ARR due to its nephrotoxic and carcinogenic properties, of which aristolochic acid I (AA I) is the most representative. The present paper describes two 2D-LC systems modulated by a sample loop and a trap column respectively for the detection of trace aristolochic acid I in ARR. An Inertsil ODS-3 C18 column and a Luna 3u Phenyl-Hexyl column were separately utilized in the first-dimension (1D) and second-dimension (2D) chromatography. A mixed-mode column with reversed-phase and ion-exchange was selected for trap column modulation of 2D-LC system. Method validation results showed that these two 2D-LC methods performed well in terms of repeatability, linearity, intra/interday precision, stability, and accuracy, which were demonstrated qualified for the simultaneous determination of trace AA I in ARR. The AA Ⅰ contents in three batches of ARR were 0.466∼0.812 μg/g by the two 2D-LC methods, which were below the limit requirement in the Chinese Pharmacopoeia. The comprehensive comparison between the two methods showed that a significant solute focusing effect was obtained by trap column modulation, which led to four-time lower LOD compared with loop modulation, while the loop-modulation-based method achieved higher analytical efficiency when analyzing a large batch of samples.