目的 建立高效液相色譜（HPLC）法同時檢測蟬花培養(yǎng)物4種核苷類成分，分析不同斜面培養(yǎng)基及固體培養(yǎng)基對4種核苷類成分的影響。方法 建立4種核酸類成分——尿苷、鳥苷、腺苷、N6-（2-羥乙基）腺苷的HPLC檢測方法，應(yīng)用Waters Symmetry shield TMRP C₁₈（250 mm×4.6 mm，5 μm）色譜柱；流動相為水-甲醇，梯度洗脫；體積流量1.0 mL/min；柱溫30℃；進樣量10 μL；檢測波長260 nm；外標法測定4種核苷類成分水平。對包括假單胞菌選擇培養(yǎng)基（PSA）、沙氏葡萄糖瓊脂培養(yǎng)基（SDAY，1 000 mL中含葡萄糖量分別為20、30、40 g）在內(nèi)的斜面培養(yǎng)基和包括江蘇農(nóng)墾（SCM）、煙農(nóng)24（YN-24）、煙農(nóng)15（YN-15）、濟麥22（JM-22）在內(nèi)的固體培養(yǎng)基培養(yǎng)的蟬花培養(yǎng)物中4種核酸類成分水平進行測定，利用SPSS16.0統(tǒng)計軟件對測定結(jié)果進行多重統(tǒng)計分析，驗證并找出引起差異的原因。結(jié)果 建立的HPLC法，4種核苷類成分分離度良好，精密度、穩(wěn)定性、重復(fù)性、加樣回收率、耐用性均符合要求；4種不同的斜面培養(yǎng)基對尿苷成分基本無影響（P>0.05），PSA對腺苷、鳥苷、N6-（2-羥乙基）腺苷的水平有明顯促進作用（P<0.05）；固體培養(yǎng)基SCM的4種核苷類成分的水平較高。結(jié)論 培養(yǎng)基種類對蟬花培養(yǎng)物核苷類成分變化有直接的影響，確定蟬花蟲草的生產(chǎn)工藝時，應(yīng)充分考慮培養(yǎng)基的選擇，同時結(jié)合產(chǎn)品成分和生產(chǎn)成本等因素進行綜合評價。

Objective The HPLC method was established to simultaneously determine the contents of four kinds of nucleosides in culture of Cordyceps cicadae, and to analyze the effect of different culture medium on nucleoside component content. Methods The HPLC method was applied to determining the contents of four kinds of nucleosides such as uridine, guanosine, adenosine, and N6-(2-hydroxyethyl) adenosine in C. cicadae. Chromatography was carried out on a Waters Symmetry shield TMRP C₁₈ column (250 mm×4.6 mm, 5 μm) using gradient elution program at the volume flow rate of 1.0 mL/min. Mobile phase was water-methanol, column temperature was 30℃, the sample weight was 10 μL and detection wave length was 260 nm. External standard method was used to calculate the content of nucleoside component. The contents of four kinds of nucleosides were measured in culture of C. cicadae for different culture medium including different slope medium such as Pseudomonas Selection Medium (PSA) and Sabouraud's Glucose Agar Medium (SDAY) containing 20, 30, or 40 g glucose sugar in 1000 mL medium; The different solid medium were Jiangsu state farms (SCM), Yannong 24 (YN-24), Yannong 15 (YN-15), and Jimai 22 (JM-22). The SPSS 16.0 statistical software was applied to verifying and find out the reason causing the difference. Results The separation degree of four kinds of nucleosides was good, and the precision, stability, repetition, sample recovery rate, and durability of HPLC method have been verified to meet the requirements. There is no effect of four different slant medium on the content of uridine. The guanosine, adenosine, and N6-(2-hydroxyethyl) adenosine levels in slant culture medium of PSA were higher than those of other slant culture medium. The levels of four kinds of nucleosides in solid culture medium of SCM were higher than those of others. Conclusion The media types have a direct impact on nucleosides in culture of C. cicadae. Therefore, to determine C. cicadae production process, should give full consideration to the choice of the medium, while the combination of product composition and production costs and other factors comprehensive evaluation.

藥用蟲生真菌中試及產(chǎn)業(yè)化平臺的建立