Simultaneous separation and determination of active components in Cordyceps sinensis and Cordyceps militarris by LC/ESI-MS

A simple and rapid isocratic LC/MS coupled with electrospray ionization (ESI) method for simultaneous separation and determination of adenine, hypoxanthine, adenosine and cordycepin in Cordyceps sinensis (Cs) and its substitutes was developed. 2-Chloroadenosine was used as internal standard for this assay. The optimum separation for these analytes was achieved using the mixture of water, methanol and formic acid (85:14:1, v/v/v) as a mobile phase and a 2.0×150 mm Shimadzu VP-ODS column. Selective ion monitoring (SIM) mode ([M+H]⁺ at m/z 136, 137, 268, 252 and 302) was used for quantitative analysis of above four active components. The regression equations were liner in the range of 1.4–140.0 μg ml⁻¹ for adenine, 0.6–117.5 μg ml⁻¹ for hypoxanthine, 0.5–128.5 μg ml⁻¹ for adenosine and 0.5–131.5 μg ml⁻¹ for cordycepin. The limits of quantitation (LOQ) and detection (LOD) were, respectively 1.4 and 0.5 μg ml⁻¹ for adenine, 0.6 and 0.2 μg ml⁻¹ for hypoxanthine, 0.5 and 0.1 μg ml⁻¹ for adenosine and cordycepin. The recoveries of four constituents were from 93.5 to 107.0%. The nucleoside contents of various types of natural Cs and its substitutes were determined and compared with this developed method.