Analysis of Transport Mechanism of Cyperotundone in Caco-2 Cell Model / 中国实验方剂学杂志

ABSTRACT

Objective: To investigate transport mechanism of cyperotundone in Caco-2 cell model and provide experimental basis for clinical application of Cyperi Rhizoma. Method: The toxicity of cyperotundone with different concentrations to Caco-2 cells was investigated by methyl thiazolyl tetrazolium (MTT) colorimetry, in order to determine the concentration of administration in transport test. The content of cyperotundone was determined by liquid chromatography-mass spectrometry (LC-MS) with apparent permeability coefficient (Papp) and cumulative transport capacity as indexes. The chromatographic conditions were as followingmobile phase of acetonitrile (A)-water (B) for gradient elution (0-1.5 min, 35%A; 1.5-2 min, 35%-90%A; 2-4 min, 90%A; 4-4.1, 90%-35%A; 4.1-8 min, 35%A), the flow rate at 0.3 mL · min-1, injection volume of 1 μL, and temperature of column at 30℃. The mass spectrometric conditions was electrospray ionization (ESI) and positive ion mode, the detection ions of cyperotundone and osthole (internal standard substance) were m/z 219.2-110.9 and m/z 245.0-189.0, respectively. Effect of concentration of cyperotundone, administration time, ethylenediamine tetraacetic acid (EDTA) and P-glycoprotein (P-gp) inhibitor on the transmembrane transport of cyperotundone on in vitro cell model were investigated. Result: Cyperotundone didn't have significant toxicity to Caco-2 cells at 3-90 mg · L-1 after incubation for 4 h. The transportion of cyperotundone in Caco-2 cell model was related to the concentration and time to a certain extent, its Papp was higher than 1×10-6 cm · s-1, which indicated that absorption of cyperotundone was good, the efflux rate (ER) of cyperotundone was 0.5-1.5.There was no significant difference in bidirectional Papp of cyperotundone after the addition of cell bypass transport inhibitor (EDTA) and P-gp transport inhibitor (verapamil). Conclusion: The transport mechanism of cyperotundone in Caco-2 cell model is mainly passive diffusion, and cell bypass transport and P-gp are not involved in its transport.