Comparative analysis of nucleoside concentrations in water extracts of natural Ophiocordyceps sinensis at 30[degrees celsius] and 100[degrees celsius]

Abstract

Sample preparation is an important aspect in the extraction of potentially pharmaceutical compounds found in traditional Chinese medicines (TCMs). It is valuable to understand which preparation method is ideal for each compound of interest. Cordyceps are among the most sought after TCM products today, with a market value of 32,000 USD per kilogram in 2006. Increasing the efficiency of extraction could potentially reduce the cost to consumers and the ecological impact of their harvesting in Tibet. An accurate, precise, and sensitive reverse-phase high performance liquid chromatography method was developed for the simultaneous separation and quantitation of uridine, inosine, and adenosine in Ophiocordyceps sinensis for boiling (100°C) and ambient temperature (30°C) water extractions. The optimum separation was achieved with a gradient elution of methanol and ammonium acetate through a 2.0 x 250mm Phenomenex Luna 5u C18 column. Linearity of the apparatus was significant with values of R[superscript 2] = 0.9994985, 0.9998885, and 0.9999775 for uridine, inosine, and adenosine respectively. The percent relative standard deviation for intra-day and inter-day precision ranged from 0.88-2.82 %RSD, and 1.88-7.57 %RSD respectively. Accuracy determined through percent recovery tests yielded an accuracy range of 99.32 - 99.91%. Limit of detection and limit of quantification for nucleosides were determined to have ranges of 0.042-0.2240.042[micrograms]/mL and 0.128-0.678[micrograms]/mL respectively. Similar results were obtained for cordyceps obtained from both Qinghai and Xizang at each temperature treatment. Boiling water was more effective for adenosine, while ambient temperature yielded more inosine and uridine. Mean concentration differences between temperature treatments were calculated for adenosine, uridine, and inosine of 9.79, 32.38, and 28.35[micrograms]/mL respectively. One-way ANOVA for the two temperature conditions of uridine, inosine, and adenosine were [F=788.89, df =11, p<0.001], [F=3352.46, df= 11, p<0.001], and [F=1211.17, df=11, p<0.001] respectively. The results suggest that sample preparation has a significant effect on the expression of nucleosides in Cordyceps.