Mechanism Of Continuous Cropping Obstacle Of Rehmannia Glutinosa

Rehmannia glutinosa Libosch is a perennial herb of Scrophulariaceae, and has been cultivated for more than 1500 years. Its root is used frequently as a crude drug in traditional Chinese medicine. Continuous cropping obstacle recorded firstly in Ming dynasty by Lu zhiyi, it is very serious in the current production, but its reason is not clear up to now. Stachyose and catalpol are the main components of root exudates and plant residua remained in the soil. They may be the main factors resulted in continuous cropping obstacle. In this dissertation, the question was investigated by simplified experiments.First of all, stachyose and catalpol were separated and purified from the fresh roots of R. glutinosa, and soil bacteria were isolated from soil collected from wheat field in Jinan, in which R. glutinosa was never planted. The effects of stachyose and catalpol on bacteria isolates were examined respectively. Cyanide in glucose ammonium liquid medium of each isolate was determined, and the effects of cyanide and bacterial culture fluid on the plantlet of R. glutinosa were also investigated. The main findings are as follows:1. Fifty soil bacteria were isolated, in which forty two were gram-negative, eight were gram-positive, and three isolates belonged to coccus, two clostridium; forty five bacillus. Colonies of the isolates were mainly roundish in shape and yellow in color, two of them could produce water-soluble pigment.2. The soil bacteria isolates were divided into four types according to the absorbance (A₁) at 600nm of the glucose ammonium liquid medium when cultured for 20 h at 28 °C, e.i. type 1 (A₁≥1 .2), type 2 (0.6≤A₁<1.2), type 3 (0.2≤A₁<0.6) and type 4 (A₁<0.2). The absorbance (A2) of the stachyose ammonium liquid medium was usually lower than A₁, so the isolates were further divided into four subtype according to the percentage of A₂ to A₁ (P=A₂/A₁×100%), e.i. subtype 1 (P≥80%), subtype 2 (40%≤P<80%), subtype 3 (20%≤P<40%) and subtype 4 (P<20%). The results indicated that only 2 isolates (type 1-1) can utilize stachyose as carbon source to a degree as glucose, 32 isolates (type 1-2,1-3 and 2-2) have a lower ability in using stachyose, the other 16 (type 1-4, 3 and 4) could hardly use it in the current condition.3. Filter paper-agar diffusion method was used to examine the suppressing effect of catalpol on soil bacteria isolates. The result indicated that catalpol couldn't inhibit the development of the soil bacteria, on the contrary, it seems to promote the propagation of some isolates at low concentration.4. The cyanide contents in the glucose ammonium liquid media was determined spectrophotometrically when the isolates were cultured for 48 h. The content was over 0.5μmol/L in 14 isolates, that of No 37 isolate was up to 17.7μmol/L. Supplement of sodium cyanide into MS medium unveiled that it could depress the growth of plantlet of R. glutinosa.5. When the 7d liquid media of the isolates were supplemented into the MS medium respectively, the plantlets were promoted in No.2, 4, 8, 9, 18 isolates, and inhibited in No.36 and 40 bacteria.In conclusion, soil bacteria were different in the capacity of utilizing stachyose. In the soil survival competition, the bacteria which can utilize stachyose effectively would flourish, and this might be the primary factor resulting in the continuous cropping obstacle of R. glutinosa. Cyanide, one of the metabolites of some soil bacteria, may be a harmful factor to the metabolism and development of the plant. In view of the inhibiting effects of some isolate culture medium on the plantlets, there are some unknown bacteria metabolites which would be deleterious to the productionof the herb, and further investigations are needed. Innovation in this dissertation:1. The continuous cropping obstacle of R. glutinosa was investigated for the firsttime in the view of rhizospheric microecosystem.2. The effects of stachyose and catalpol on soil bacteria were firstly investigated, and stachyose was proposed to be one of the important reasons resulting in the unbalance of soil microecosystem.3. Inhibiting effect of cyanide was firstly proved. The cyanide and some unknown metabolites of some soil bacteria were considered to be harmful factors to the plantation of R. glutinosa.