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目的建立茅苍术悬浮细胞系,研究2种内生真菌诱导子对茅苍术悬浮细胞产挥发油的影响。方法用内生真菌诱导子对茅苍术悬浮细胞进行诱导处理后超声提取茅苍术细胞中的挥发油,利用气相色谱法测定挥发油中苍术酮、苍术醇、β-桉叶醇和苍术素的量。结果获得的茅苍术悬浮细胞生长较快,于第21天生物量达到最大,为6.95 g/L。未经诱导处理的茅苍术悬浮细胞中仅检出β-桉叶醇,其在培养21 d的细胞中的量达到最大,为17.469μg/g。添加2种内生真菌(分别属于小克银汉霉属和孔球孢霉属,编号为AL4和AL12)的粗诱导子对茅苍术悬浮细胞生物量及挥发油积累均有影响,其中AL4粗诱导子综合效果更好。添加AL4粗诱导子到14 d的茅苍术悬浮细胞培养基中,诱导子质量浓度为每升培养基含糖20 mg,诱导处理7 d后收集细胞,发现其干质量比同期对照提高3.31%,且检出细胞中苍术酮、苍术醇、β-桉叶醇和苍术素的干质量分别达到14.715、28.395、38.794、8.310μg/g。其中β-桉叶醇的量是对照的2.22倍。结论添加内生真菌诱导子对茅苍术悬浮细胞的生长及挥发油积累均有促进作用。
OBJECTIVE To establish the cell suspension of Atractylodes lancea and study the effects of two endophytic fungi inducer on the volatile oil of Atractylodes macrocephala suspension cells. Methods The endogenous fungal elicitors were used to induce the cell suspension of Atractylodes lancea. The volatile oil was extracted from the cells of Atractylodes macrocephala by ultrasound, and the amount of atractyloside, atractylodin, β-eucalyptol and atractylodes macrocephala were determined by gas chromatography. The results obtained Atractylodes macrocephala suspension cells grew rapidly, the biomass reached the maximum on the 21st day, 6.95 g / L. Only β-eucalyptol was detected in the untreated cells of Atractylodes lancea, which reached the maximum of 17.469μg / g in the cells cultured for 21 days. The crude inducer added with 2 kinds of endophytic fungi (belonging to the genus Micrococynospora and the genus Perfospora, numbered AL4 and AL12 respectively) had the effect on the biomass and the accumulation of volatile oil in the suspension cells of Atractylodes macrocephala, of which the AL4 crude inducible Comprehensive effect is better. After adding AL4 crude inducer to Atractylodes macrocephala suspension cell culture medium for 14 days, the concentration of inducer was 20 mg / liter for each liter of culture medium, and after 7 days of induction treatment, the cells were collected and the dry weight was found 3.31% The dry weight of atractylodin, atractylodin, β-eucalyptol and atractylodes macrocephala were 14.715, 28.395, 38.794 and 8.310 μg / g respectively. The amount of β-eucalyptol was 2.22 times that of the control. Conclusion The addition of endophytic fungi elicitor can promote the growth of Atractylodes lancea and the accumulation of volatile oil.