Studies on secondary metabolites contained in virus free plantlet of Rehmannia glutinosa

认为在田间栽培的地黄植株几乎都可能感染病毒。因此，对鲜叶进行了电子显微镜分析，检测出包括TMV在内的三种病毒。为了消除病毒，建立了茎尖组织培养无性系繁殖的方法。根尖组织培养的植株再生和病毒的清除是成功的和常规的。田间栽培2年、3年和4年后，再感染率分别为26%、31%和68%。为确定病毒感染与次生代谢物的关系，建立了环烯醚萜苷的高效液相色谱定量分析方法，并对栽培根的质量进行了评价。除脱毒根的环烯醚萜含量高于脱毒根外，未发现脱毒根与感染病毒根的环烯醚萜含量有明显差异。但变异显著，主要环烯醚萜苷、梓醇在病根中因细菌侵染而消失，而病毒侵染则加速其消失。同样的现象也发生在其他环烯醚萜苷上。此外，从病根中分离到两种酚类苷，结构鉴定为毛蕊糖苷和6′-0-半乳糖糖苷。从真菌接种产生的两种酚类苷及其抗菌活性的实验来看，两种酚类苷可以认为是地黄根中的植物抗菌素。结果表明，病毒感染引起的谷地白蚁异常的次生代谢是通过真菌感染间接发生的。

It is considered that almost all plantlets of Rehmannia glutinosa cultivated in field may be infected with virus. Therefore, the fresh leaves have been analyzed by the electron microscopy and three viruses including TMV are detected. In order to eliminate the viruses, the clonal propatation by shoot tip tissue culture has been set up. The plantlet regeneration from tip tissue culture and the elimination of viruses are successfully and routinely carried out. Experiments on reinfection with TMV are investigated and the reinfection rate is estimated as 26%, 31% and 68% after cultivation in field for 2,3 and 4 years, respectively.To confirm the relation between virus infection and secondary metabolites, the quantitative analysis method by HPLC for iridoid glycosides has been established and estimated the quality of cultivated root. The clear difference between the virus free root and virus infected root was not found except the former containes relatively higher amount of iridoids than the later. However, the remarkable variation is observed, that is, the major iridoid glycoside, catalpol is disappered in the diseased root by bacterial infection which is accelerated by virus infection. The same phenomenon is occurred on other iridoid glycosides. Moreover, two phenolic glycosides have been isolated from the diseased root and the structures are elucidated as acteoside and 6'-0-galactosylacteoside. From the experiments concerning the production of those by fungal inoculation and the antibiotic activity it appears that two phenolic glycosides may be considered as phytoalexin in the root of R. glutinosa. It is concluded that the unusual secondary metabolism in R. glutinosa by virus infection occurres indirectly through fungal infection.